Organizational Justice, the Psychological Contract, and Relashionships at work are the most important in making people dissatisfied with their jobs. Person-environment fit, Organizational justice, and Work relashionships are the most important relating to organizational commitment.

At work, two job attitudes have the greatest potential to influence how we behave. These are job satisfaction and organizational commitment.

Job satisfaction refers to the feelings people have toward their job.

Organizational commitment is the emotional attachment people have toward the company they work for.

There is a high degree of overlap between job satisfaction and organizational commitment because things that make us happy with our job often make us more committed to the company as well.

They often are associated with outcomes that are important to the Controlling role, such as performance, helping others, absenteeism, and turnover.

hello

not good

I really think two parts are especially troubling. First, relying on IQ for placement ignores how those tests reflect language, prior access, and test familiarity. It turns a snapshot into destiny. Second, limiting world language courses to the top group withholds a powerful learning tool from everyone else. Studying languages builds literacy, memory, and cultural knowledge. It should be an on-ramp with supports, not a reward after you are already tracked up.

I think that a simple test for authenticity is three questions. Who sets the agenda. What in core practice changes. Who is accountable. If students and families help choose texts and examples, if grading and grouping shift so more students get access to hard work, and if leaders protect time and coaching to make those shifts stick, it stops being tokenism. At a school like Dynamic that might look like co-planning one unit per department with student advisors, replacing a single high-stakes test with multiple ways to show learning, translating every family touchpoint, and tracking access to advanced courses and belonging by subgroup so teachers see impact, not just intentions.

A big May assembly can be celebratory yet still be a festival model: culture as entertainment for a majority audience. If the club’s biggest footprint is during Asian American Month, the burden of representation shifts to students while curriculum and school policies remain unchanged. A stronger model spreads Asian American histories, literatures, and contemporary issues across subjects all year.

Culture shifts show up in discipline and classroom routines. If the old Dynamic felt like students “ruled the school,” what replaced zero tolerance or ad hoc reactions. Consistent schoolwide norms taught like content. Administrators doing learning walks that give bite size feedback on bell to bell instruction. Restorative responses for harm alongside clear boundaries so safety is not left to chance. A predictable on ramp for new teachers so expectations are the same in every room. When those pieces exist, classrooms stop running on personality and start running on shared practice.

This vision gets the everyday pieces right. Food, play, and language are not extras. They are daily signals of who belongs. When the lunch menu matches what families actually eat, when games include cooperative play, and when home languages show up in class, students learn that their lives count as knowledge.

To make this real, plan with families, not for them. Build menus with parent input. Translate every notice and invite replies in any language. Hire and grow bilingual staff. Use students’ first languages for thinking and drafting, then shape the final product for the audience. Track whether more families engage, whether attendance and belonging rise, and whether more students enter advanced classes. That is how curriculum, teaching, and outreach line up with a true multicultural philosophy.

It's right to call out how a narrow canon quietly teaches who counts as a maker of “real” knowledge. When art history means only France and Italy and “classical music” means only Europe, students learn that others are audiences, not authors. That framing flattens the past and limits the kinds of questions students learn to ask.

A better approach is both and, not either or. Keep core analytic skills, then widen the examples. Put Botticelli beside Ife bronzes and Mughal ateliers. Pair Beethoven with Arabic maqam, Hindustani raga, Javanese gamelan, and Andean court traditions that function as classical in their own contexts. In history, center agency beyond Europe. Mansa Musa and Song innovation. Zheng He and Indigenous confederacies. Then match the assessments to the shift. Listening journals across traditions. Curate a mini exhibition that argues for a global canon. DBQs that use primary sources from many languages in translation. Invite local culture bearers to co teach a session. Do all this and “basic education” becomes a set of methods and lenses that work across the world, not a single story.

I believe there is great importance in multicultural education to address topics like racism and discrimination. These issues are not something that disappear in a school setting, especially when they are deeply rooted in the system. Talking about them and opening up the conversation that these actions are not acceptable is crucial for students to understand what they should and shouldn't tolerate in how they are treated by others.

Totally agree with this frame. When multicultural education is treated as school reform, not a festival unit, it gives you tools to attack the conditions that produce underachievement. That means redesigning core structures—who gets into advanced classes, how we assess and group, how language is treated in instruction, how discipline works, and how families participate—so access, challenge, and belonging are routine for every student, not extras for a few.

A quick gut-check for “real change” looks like this: de-track gateway courses with built-in supports, not gatekeeping tests. Use translanguaging and bilingual options so language is an asset in learning. Replace one-shot high-stakes tests with multiple measures tied to strong teaching. Make counseling, attendance, and health supports easiest to reach where need is highest. Share decisions with families in their languages. Then watch the right indicators: AP and STEM access rising across groups, discipline gaps shrinking, attendance and belonging improving, and student work showing more voices and high rigor. That is how multicultural education becomes a climate shift, not a slogan.

I really connected with this quote because it explains why storytelling is so effective. I agree that stories help facts stick better, and thinking about it, most of the meaningful lessons I’ve learned in my own life came through someone’s story, not just raw information. The idea that stories act like “simulators for life” also stood out to me, because it’s true that hearing someone else’s experience can prepare us for situations we haven’t faced yet. This made me realize how much storytelling shapes the way we understand people, which makes it a powerful tool in design and communication.

I really like these two paragraphs. I concur that storytelling is universal and I make the connection with the Informatics admissions process. Overall, the essay counts for about 80% of the application grade and it is basically measuring our storytelling skills. I like that is being approached somewhat more conceptually and in-depth in this chapter.

This sentence really sticks out to me. I never realized how stories have impacted me into making decisions and this sentence got me thinking back and realizing that I have in fact made multiple decisions based on different "experiences" I had through stories. I think one that everyone and anyone can agree one is about love. Everyone has their first loves and for me, I remember solely relying on all the stores from books, movies, TV shows as my reference for my first ever crush back in middle school. Or even walking into college, there are clips in shows or movies that picture "college life" which gave me a good idea of comparison when I first came into college.

I really enjoy reading about the two real-world examples the author gives here for the "And-But-Therefore" template. Seeing how it applied to both a service and a product makes the concept make sense for me in a way the theory alone didn't. I could actually picture how I might use it in a real project. It is also interesting when it is said that these examples aren't the "public-facing language," but the "DNA of your core narrative." That made me think more deeply about how much thought has to go into the foundation of a campaign, not just the fancy sentences we put in ads. It just made me realize that marketing isn’t just about listing features but actually about telling a story that shows how a problem is solved.

I liked this section because it shows how you can tell the same story in different ways just by deciding where to add tension. I think the And-But-Therefore structure is actually pretty useful since it makes you think more about what parts of the story you want people to focus on. It also made me realize that storytelling isn’t just about what happened, but how you choose to frame it.

This line stayed with me because it captures the core argument of the reading with such simplicity. This idea made me reflect on how often information fails to stick not because it isn’t valuable, but because it lacks the emotional pull that tension provides. I found the comparison between the monomyth and life science marketing surprisingly compelling, showing how even technical communication benefits from narrative structure. The And-But-Therefore model stood out as a practical, almost deceptively simple tool that distills storytelling into something immediately usable. Overall, the reading reinforced for me that good storytelling is less about embellishment and more about intentional structure that guides attention and emotion.

refers to Rossetti's poetry which often embodied Victorian gender norms, contributing to limited and idealized notions of women's roles

Woolf highlighted how women lacked access to money, education or even a private space which prevent them to enjoy the same freedom that men has during the early 20th century where significant social changes affected women.

The "beadle" stopping her from walking on the grass is a symbolism of institutional gatekeeping (shows how institutional rules and physical boundaries were used to restrict women's access to academic spaces) shows Oxbridge as a symbol of male-dominated academia.

eLife Assessment

The manuscript by Shukla et al. provides important mechanistic insights into kinesin-1 autoinhibition and cargo-mediated activation. Using a convincing combination of protein engineering, computational modeling, biophysical assays, HDX-MS, and electron microscopy, the authors reveal how cargo binding induces an allosteric transition that propagates to the motor domains and enhances MAP7 binding. Despite limitations arising from conformational heterogeneity and structural resolution, the study presents a unified mechanism for kinesin-1 activation that will be of broad interest to the motor protein, structural biology, and cell biology communities.

Reviewer #1 (Public review):

Summary:

The authors aim to interrogate the sets of intramolecular interactions that cause kinesin-1 hetero-tetramer autoinhibition and the mechanism by which cargo interactions via the light chain tetratricopeptide repeat domains can initiate motor activation. The molecular mechanisms of kinesin regulation remain an important question with respect to intracellular transport. It has implications for the accuracy and efficiency of motor transport by different motor families, for example, the direction of cargos towards one or other microtubules.

Strengths:

The authors focus on the response of inactivated kinesin-1 to peptides found in cargos and the cascade of conformational changes that occur. They also test the effects of the known activator of kinesin-1 - MAP7 - in the context of their model. The study benefits from multiple complementary methods - structural prediction using AlphaFold3, 2D and 3D analysis of (mainly negative stain) TEM images of several engineered kinesin constructs, biophysical characterisation of the complexes, peptide design, hydrogen/deuterium-exchange mass spectrometry, and simple cell-based imaging. Each set of experiments is thoughtfully designed, and the intrinsic limitations of each method are offset by other approaches such that the assembled data convincingly support the authors' conclusions. This study benefits from prior work by the authors on this system and the tools and constructs they previously accrued, as well as from other recent contributions to the field.

Weaknesses:

It is not always straightforward to follow the design logic of a particular set of experiments, with the result that the internal consistency of the data appears unconvincing in places. For example, i) the Figure 1 AlphaFold3 models do not include motor domains whereas the nearly all of the rest of the data involve constructs with the motor domains; ii) the kinesin constructs are chemically cross-linked prior to TEM sample preparation - this is clear in the Methods but should be included in the Results text, together with some discussion of how this might influence consistency with other methods where crosslinking was not used. Can those cross-links themselves be used to probe the intramolecular interactions in the molecular populations by mass spec? In general, the information content of some of the figure panels can also be improved with more annotations (e.g. angular relationship between views in Figure 1B, approximate interpretations of the various blobs in Fig 3F, and more thought given to what the reader should extract from the representative micrographs in several figures - inclusion of the raw data is welcome but extraction and magnification of exemplar particles (as is done more effectively in Fig S5) could convey more useful information elsewhere.

Reviewer #2 (Public review):

Summary:

In this paper, Shukla, Cross, Kish, and colleagues investigate how binding of a cargo-adaptor mimic (KinTag) to the TPR domains of the kinesin-1 light chain, or disruption of the TPR docking site (TDS) on the kinesin-1 heavy chain, triggers release of the TPR domains from the holoenzyme. This dislocation provides a plausible mechanism for transition out of the autoinhibited lambda-particle toward the open and active conformation of kinesin-1. Using a combination of negative-stain electron microscopy, AlphaFold modeling, biochemical assays, hydrogen-deuterium exchange mass spectrometry (HDX-MS), and other methods, the authors show how TPR undocking propagates conformational changes through the coiled-coil stalk to the motor domains, increasing their mobility and enhancing interactions with the microtubule-bound cofactor MAP7. Together, they propose a model in which the TDS on CC1 of the heavy chain forms a "shoulder" in the compact, autoinhibited state. Cargo-adaptor binding, mimicked here by KinTag, dislodges this shoulder, liberating the motor domains and promoting MAP7 association, driving kinesin-1 activation.

Strengths:

Throughout the study, the authors use a clever construct design - e.g., delta-Elbow, ElbowLock, CC-Di, and the high-affinity KinTag - to test specific mechanisms by directly perturbing structural contacts or affecting interactions. The proposed mechanism of releasing autoinhibition via adaptor-induced TPR undocking is also interrogated with a number of complementary techniques that converge on a convincing model for activation that can be further tested in future studies. The paper is well-written and easy to follow, though some more attention to figure labels and legends would improve the manuscript (detailed in recommendations for the authors).

Weaknesses:

These reflect limits of what the current data can establish rather than flaws in execution. It remains to be tested if the open state of kinesin-1 initiated by TPR undocking is indeed an active state of kinesin-1 capable of processive movement and/or cargo transport. It also remains to be determined what the mechanism of motor domain undocking from the autoinhibited conformation is, and perhaps this could have been explored more here. The authors have shown by HDX-MS that the motor domains become more mobile on KinTag binding, but perhaps molecular dynamics would also be useful for modelling how that might occur.

Reviewer #3 (Public review):

Summary:

The manuscript by Shukla and colleagues presents a comprehensive study that addresses a central question in kinesin-1 regulation - how cargo binding to the kinesin light chain (KLC) tetratricopeptide repeat (TPR) domains triggers activation of full-length kinesin-1 (KHC). The authors combine AlphaFold3 modeling, biophysical analysis (fluorescence polarization, hydrogen-deuterium exchange), and electron microscopy to derive a mechanistic model in which the KLC-TPR domains dock onto coiled-coil 1 (CC1) of the KHC to form the "TPR shoulder," stabilizing the autoinhibited (λ-particle) conformation. Binding of a W/Y-acidic cargo motif (KinTag) or deletion of the CC1 docking site (TDS) dislocates this shoulder, liberating the motor domains and enhancing accessibility to cofactors such as MAP7. The results link cargo recognition to allosteric structural transitions and present a unified model of kinesin-1 activation.

Strengths:

(1) The study addresses a fundamental and long-standing question in kinesin-1 regulation using a multidisciplinary approach that combines structural modeling, quantitative biophysics, and electron microscopy.

(2) The mechanistic model linking cargo-induced dislocation of the TPR shoulder to activation of the motor complex is well supported by both structural and biochemical evidence.

(3) The authors employ elegant protein-engineering strategies (e.g., ElbowLock and ΔTDS constructs) that enable direct testing of model predictions, providing clear mechanistic insight rather than purely correlative data.

(4) The data are internally consistent and align well with previous studies on kinesin-1 regulation and MAP7-mediated activation, strengthening the overall conclusion.

Weaknesses:

(1) While the EM and HDX-MS analyses are informative, the conformational heterogeneity of the complex limits structural resolution, making some aspects of the model (e.g., stoichiometry or symmetry of TPR docking) indirect rather than directly visualized.

(2) The dynamics of KLC-TPR docking and undocking remain incompletely defined; it is unclear whether both TPR domains engage CC1 simultaneously or in an alternating fashion.

(3) The interplay between cargo adaptors and MAP7 is discussed but not experimentally explored, leaving open questions about the sequence and exclusivity of their interactions with CC1.

eLife Assessment

This important study describes a new link between nutrient signaling and chromosome regulation, providing compelling evidence that reduced activity in the central nutrient-sensing pathway governed by TORC1 improves chromosome stability and alters gene expression in S. pombe through effects on cohesin. While the biological importance of this newly described circuit is not yet fully known, and some data would benefit from further clarification, the overall body of evidence supports the main conclusions.

Reviewer #1 (Public review):

Summary:

In this study, Besson et al. investigate how environmental nutrient signals regulate chromosome biology through the TORC1 signaling pathway in Schizosaccharomyces pombe. Specifically, the authors explore the impact of TORC1 on cohesin function - a protein complex essential for chromosome segregation and transcriptional regulation. Through a combination of genetic screens, biochemical analysis, phospho-proteomics, and transcriptional profiling, they uncover a functional and physical interaction between TORC1 and cohesin. The data suggest that reduced TORC1 activity enhances cohesin binding to chromosomes and improves chromosome segregation, with implications for stress-responsive gene expression, especially in subtelomeric regions.

Strengths:

This work presents a compelling link between nutrient sensing and chromosome regulation. The major strength of the study lies in its comprehensive and multi-disciplinary approach. The authors integrate genetic suppression screens, live-cell imaging, chromatin immunoprecipitation, co-immunoprecipitation, and mass spectrometry to uncover the functional connection between TORC1 signaling and cohesin. The use of phospho-mutant alleles of cohesin subunits and their loader provides mechanistic insight into the regulatory role of phosphorylation. The addition of transcriptomic analysis further strengthens the biological relevance of the findings and places them in a broader physiological context. Altogether, the dataset convincingly supports the authors' main conclusions and opens up new avenues of investigation.

Weaknesses:

While the study is strong overall, a few limitations are worth noting. The consistency of cohesin phosphorylation changes under different TORC1-inhibiting conditions (e.g., genetic mutants vs. rapamycin treatment) is unclear and could benefit from further clarification. The phosphorylation sites identified on cohesin subunits do not match known AGC kinase consensus motifs, raising the possibility that the modifications are indirect. The study relies heavily on one TORC1 mutant allele (mip1-R401G), and additional alleles could strengthen the generality of the findings. Furthermore, while the results suggest that nutrient availability influences cohesin function, this is not directly tested by comparing growth or cohesin dynamics under defined nutrient conditions.

Reviewer #2 (Public review):

Summary:

In this study, the authors follow up on a previous suppressor screen of a temperature-sensitive allele of mis4 (mis4-G1487D), the cohesin loading factor in S. pombe, and identify additional suppressor alleles tied to the S. pombe TORC1 complex. Their analysis suggests that these suppressor mutations attenuate TORC1 activity, while enhanced TORC1 activity is deleterious in this context. Suppression of TORC1 activity also ameliorates chromosome segregation and spindle defects observed in the mis4-G1487D strain, although some more subtle effects are not reconstituted. The authors provide evidence that this genetic suppression is also tied to the reconstitution of cohesin loading. Moreover, disrupting TORC1 also enhances Mis4/cohesin association with chromatin (likely reflecting enhanced loading) in WT cells, while rapamycin treatment can enhance the robustness of chromosome transmission. These effects likely arise directly through TORC1 or its downstream effector kinases, as TORC1 co-purifies with Mis4 and Rad21; these factors are also phosphorylated in a TORC1-dependent fashion. Disrupting Sck2, a kinase downstream of TORC1, also suppresses the mis4-G1487D allele while simultaneous disruption of Sck1 and Sck2 enhances cohesin association with chromatin, albeit with differing effects on phosphorylation of Mis4 and Psm1/Scm1. Phosphomutants of Mis4 and Psm1 that mimic observed phosphorylation states identified by mass spectrometry that are TORC1-dependent also suppressed phenotypes observed in the mis4-G1487D background. Last, the authors provide evidence that the mis4-G1487D background and TORC1 mutant backgrounds display an overlap in the dysregulation of genes that respond to environmental conditions, particularly in genes tied to meiosis or other "stress".

Overall, the authors provide compelling evidence from genetics, biochemistry, and cell biology to support a previously unknown mechanism by which nutrient sensing regulates cohesin loading with implications for the stress response. The technical approaches are generally sound, well-controlled, and comprehensive.

Specific Points:

(1) While the authors favor the model that the enhanced cohesin loading upon diminished TORC1 activity helps cells to survive harsh environmental conditions, as starvation of S. pombe also drives commitment to meiosis, it seems as plausible that enhanced cohesin loading is related to preparing the chromosomes to mate.

(2) Related to Point 1, the lab of Sophie Martin previously published that phosphorylation of Mis4 characterizes a cluster of phosphotargets during starvation/meiotic induction (PMID: 39705284). This work should be cited, and the authors should interrogate how their observations do or do not relate to these prior observations (are these the same phosphosites?).

(3) It would be useful for the authors to combine their experimental data sets to interrogate whether there is a relationship between the regions where gene expression is altered in the mis4-G1487D strain and changes in the loading of cohesin in their ChIP experiments.

(4) Given that the genes that are affected are predominantly sub-telomeric while most genes are not affected in the mis4-G1487D strain, one possibility that the authors may wish to consider is that the regions that become dysregulated are tied to heterochromatic regions where Swi6/HP1 has been implicated in cohesin loading.

(5) It would be helpful to show individual data points from replicates in the bar graphs - it is not always clear what comprises the data sets, and superplots would be of great help.

eLife Assessment

Mitochondrial DNA (mtDNA) exhibits a degree of resistance to mutagenesis under genotoxic stress, and this study on the mitochondrial Transcription Factor A (TFAM) presents valuable data concerning the possible mechanisms involved. The presented data are solid, technically rigorous, and consistent with established literature findings. The experiments are well-executed, providing reliable evidence on the change of TFAM-DNA interactions following UVC irradiation. However, the evidence is inadequate to support the primary claims.

Reviewer #1 (Public review):

Summary:

The authors investigate how UVC-induced DNA damage alters the interaction between the mitochondrial transcription factor TFAM and mtDNA. Using live-cell imaging, qPCR, atomic force microscopy (AFM), fluorescence anisotropy, and high-throughput DNA-chip assays, they show that UVC irradiation reduces TFAM sequence specificity and increases mtDNA compaction without protecting mtDNA from lesion formation. From these findings, the authors suggest that TFAM acts as a "sensor" of damage rather than a protective or repair-promoting factor.

Strengths:

(1) The focus on UVC damage offers a clean system to study mtDNA damage sensing independently of more commonly studied repair pathways, such as oxidative DNA damage. The impact of UVC damage is not well understood in the mitochondria, and this study fills that gap in knowledge.

(2) In particular, the custom mitochondrial genome DNA chip provides high-resolution mapping of TFAM binding and reveals a global loss of sequence specificity following UVC exposure.

(3) The combination of in vitro TFAM DNA biophysical approaches, combined with cellular responses (gene expression, mtDNA turnover), provides a coherent multi-scale view.

(4) The authors demonstrate that TFAM-induced compaction does not protect mtDNA from UVC lesions, an important contribution given assumptions about TFAM providing protection.

Weaknesses:

(1) The authors show a decrease in mtDNA levels and increased lysosomal colocalization but do not define the pathway responsible for degradation. Distinguishing between replication dilution, mitophagy, or targeted degradation would strengthen the interpretation

(2) The sudden induction of mtDNA replication genes and transcription at 24 h suggests that intermediate timepoints (e.g., 12 hours) could clarify the kinetics of the response and avoid the impression that the sampling coincidentally captured the peak.

(3) The authors report no loss of mitochondrial membrane potential, but this single measure is limited. Complementary assays such as Seahorse analysis, ATP quantification, or reactive oxygen species measurement could more fully assess functional integrity.

(4) The manuscript briefly notes enrichment of TFAM at certain regions of the mitochondrial genome but provides little interpretation of why these regions are favored. Discussion of whether high-occupancy sites correspond to regulatory or structural elements would add valuable context.

(5) It remains unclear whether the altered DNA topology promotes TFAM compaction or vice versa. Addressing this directionality, perhaps by including UVC-only controls for plasmid conformation, would help disentangle these effects if UVC is causing compaction alone.

(6) The authors provide a discrepancy between the anisotropy and binding array results. The reason for this is not clear, and one wonders if an orthogonal approach for the binding experiments would elucidate this difference (minor point).

Assessment of conclusions:

The manuscript successfully meets its primary goal of testing whether TFAM protects mtDNA from UVC damage and the impact this has on the mtDNA. While their data points to an intriguing model that TFAM acts as a sensor of damaged mtDNA, the validation of this model requires further investigation to make the model more convincing. This is likely warranted for a follow-up study. Also, the biological impact of this compaction, such as altering transcription levels, is not clear in this study.

Impact and utility of the methods:

This work advances our understanding of how mitochondria manage UVC genome damage and proposes a structural mechanism for damage "sensing" independent of canonical repair. The methodology, including the custom TFAM DNA chip, will be broadly useful to the scientific community.

Context:

The study supports a model in which mitochondrial genome integrity is maintained not only by repair factors, but also by selective sequestration or removal of damaged genomes. The demonstration that TFAM compaction correlates with damage rather than protection reframes an interesting role in mtDNA quality control.

Reviewer #2 (Public review):

Summary:

King et al. present several sets of experiments aimed to address the potential impact of UV irradiation on human mitochondrial DNA as well as the possible role of mitochondrial TFAM protein in handling UV-irradiated mitochondrial genomes. The carefully worded conclusion derived from the results of experiments performed with human HeLa cells, in vitro small plasmid DNA, with PCR-generated human mitochondrial DNA, and with UV-irradiated small oligonucleotides is presented in the title of the manuscript: "UV irradiation alters TFAM binding to mitochondrial DNA". The authors also interpret results of somewhat unconnected experimental approaches to speculate that "TFAM is a potential DNA damage sensing protein in that it promotes UVC-dependent conformational changes in the [mitochondrial] nucleoids, making them more compact." They further propose that such a proposed compaction triggers the removal of UV-damaged mitochondrial genomes as well as facilitates replication of undamaged mitochondrial genomes.

Strengths:

(1) The authors presented convincing evidence that a very high dose (1500 J/m2) of UVC applied to oligonucleotides covering the entire mitochondrial DNA genome alleviates sequence specificity of TFAM binding (Figure 3). This high dose was sufficient to cause UV lesions in a large fraction of individual oligonucleotides. The method was developed in the lab of one of the corresponding authors (reference 74) and is technically well-refined. This result can be published as is or in combination with other data.

(2) The manuscript also presents AFM evidence (Figure 4) that TFAM, which was long known to facilitate compaction of the mitochondrial genome (Alam et al., 2003; PMID 12626705 and follow-up citations), causes in vitro compaction of a small pUC19 plasmid and that approximately 3 UVC lesions per plasmid molecule result in a slight, albeit detectable, increase in TFAM compaction of the plasmid. Both results can be discussed in line with a possible extrapolation to in vivo phenomena, but such a discussion should include a clear statement that no in vivo support was provided within the set of experiments presented in the manuscript.

Weaknesses:

Besides the experiments presented in Figures 3 and 4, other results do not either support or contradict the speculation that TFAM can play a protective role, eliminating mitochondrial genomes with bulky lesions by way of excessive compaction and removing damaged genomes from the in vivo pool.

To specify these weaknesses:

(1) Figure 1 - presents evidence that UVC causes a reduction in the number of mitochondrial spots in cells. The role of TFAM is not assessed.

(2) Figure 2 - presents evidence that UVC causes lesions in mitochondrial genomes in vivo, detectable by qPCR. No direct assessment of TFAM roles in damage repair or mitochondrial DNA turnover is assessed despite the statements in the title of Figure 2 or in associated text. Approximately 2-fold change in gene expression of TFAM and of the three other genes does not provide any reasonable support to suggestion about increased mitochondrial DNA turnover over multiple explanations on related to mitochondrial DNA maintenance.

(3) Figure 5. Shows that TFAM does not protect either mitochondrial nucleoids formed in vitro or mitochondrial DNA in vivo from UVC lesions as well as has no effect on in vivo repair of UV lesions.

(4) Figure 6: Based on the above analysis, the model of the role of TFAM in sensing mtDNA damage and elimination of damaged genomes in vivo appears unsupported.

(5) Additional concern about Figure 3 and relevant discussion: It is not clear if more uniform TFAM binding to UV irradiated oligonucleotides with varying sequence as compared to non-irradiated oligonucleotides can be explained by just overall reduced binding eliminating sequence specific peaks.

Reviewer #3 (Public review):

Summary:

The study is grounded in the observations that mitochondrial DNA (mtDNA) exhibits a degree of resistance to mutagenesis under genotoxic stress. The manuscript focuses on the effects of UVC-induced DNA damage on TFAM-DNA binding in vitro and in cells. The authors demonstrate increased TFAM-DNA compaction following UVC irradiation in vitro based on high-throughput protein-DNA binding and atomic force microscopy (AFM) experiments. They did not observe a similar trend in fluorescence polarization assays. In cells, the authors found that UVC exposure upregulated TFAM, POLG, and POLRMT mRNA levels without affecting the mitochondrial membrane potential. Overexpressing TFAM in cells or varying TFAM concentration in reconstituted nucleoids did not alter the accumulation or disappearance of mtDNA damage. Based on their data, the authors proposed a plausible model that, following UVC-induced DNA damage, TFAM facilitates nucleoid compaction, which may serve to signal damage in the mitochondrial genome.

Strengths:

The presented data are solid, technically rigorous, and consistent with established literature findings. The experiments are well-executed, providing reliable evidence on the change of TFAM-DNA interactions following UVC irradiation. The proposed model may inspire future follow-up studies to further study the role of TFAM in sensing UVC-induced damage.

Weaknesses:

The manuscript could be further improved by refining specific interpretations and ensuring terminology aligns precisely with the data presented.

(1) In line 322, the claim of increased "nucleoid compaction" in cells should be removed, as there is a lack of direct cellular evidence. Given that non-DNA-bound TFAM is subject to protease digestion, it is uncertain to what extent the overexpressed TFAM actually integrates into and compacts mitochondrial nucleoids in the absence of supporting immunofluorescence data.

(2) In lines 405 and 406, the authors should avoid equating TFAM overexpression with compaction in the cellular context unless the compaction is directly visualized or measured.

(3) In lines 304 and 305 (and several other places throughout the manuscript), the authors use the term "removal rates". A "removal rate" requires a direct comparison of accumulated lesion levels over a time course under different conditions. Given the complexity of UV-induced DNA damage-which involves both damage formation and potential removal via multiple pathways-a more accurate term that reflects the net result of these opposing processes is "accumulated DNA damage levels." This terminology better reflects the final state measured and avoids implying a single, active 'removal' pathway without sufficient kinetic data.

(4) In line 357, the authors refer to the decrease in the total DNA damage level as "The removal of damaged mtDNA". The decrease may be simply due to the turnover and resynthesis of non-damaged mtDNA molecules. The term "removal" may mislead the casual reader into interpreting the effect as an active repair/removal process.

eLife Assessment

This study investigates the folding and unfolding behavior of the doubly knotted protein TrmD-Tm1570, providing insight into the molecular mechanisms underlying protein knotting. The findings reveal multiple unfolding pathways and suggest that the formation of double knots may require chaperone assistance, offering valuable insights into topologically complex proteins. The evidence is solid, supported by consistent agreement between simulation and experiment, though some aspects of the presentation and experimental scope could be clarified or expanded.

Reviewer #1 (Public review):

Summary:

This paper investigates the thermal and mechanical unfolding pathways of the doubly knotted protein TrmD-Tm1570 using molecular simulations, optical tweezers experiments, and other methods. In particular, the detailed analysis of the four major unfolding pathways using a well-established simulation method is an interesting and valuable result.

Strengths:

A key finding that lends credibility to the simulation results is that the molecular simulations at least qualitatively reproduce the characteristic force-extension distance profiles obtained from optical tweezers experiments during mechanical unfolding. Furthermore, a major strength is that the authors have consistently studied the folding and unfolding processes of knotted proteins, and this paper represents a careful advancement building upon that foundation.

Weaknesses:

While optical tweezers experiments offer valuable insights, the knowledge gained from them is limited, as the experiments are restricted to this single technique.

The paper mentions that the high aggregation propensity of the TrmD-Tm1570 protein appears to hinder other types of experiments. This is likely the reason why a key aspect, such as whether a ribosome or molecular chaperones are essential for the folding of TrmD-Tm1570, has not been experimentally clarified, even though it should be possible in principle.

Reviewer #2 (Public review):

Summary:

In this manuscript, the authors combined coarse-grained structure-based model simulation, optical tweezer experiments, and AI-based analysis to assess the knotting behavior of the TrmD-Tm1570 protein. Interestingly, they found that while the structure-based model can fold the single knot from TrmD and Tm1570, the double-knot protein TrmD-Tm1570 cannot form a knot itself, suggesting the need for chaperone proteins to facilitate this knotting process. This study has strong potential to understand the molecular mechanism of knotted proteins, supported by many experimental and simulation evidence. However, there are a few places that appear to lack sufficient details, and more clarification in the presentation is needed.

Strengths:

A combination of both experimental and computational studies.

Weaknesses:

There is a lack of detail to support some statements.

(1) The use of the AI-based method, SOM, can be emphasized further, especially in its analysis of the simulated unfolding trajectories and discovery of the four unfolding/folding pathways. This will strengthen the statistical robustness of the discovery.

(2) The manuscript would benefit from a clearer description of the correlation between the simulation and experimental results. The current correlation, presented in the paragraph starting from Line 250, focuses on measured distances. The authors could consider providing additional evidence on the order of events observed experimentally and computationally. More statistical analyses on the experimental curves presented in Figure 4 supplement would be helpful.

(3) How did the authors calibrate the timescale between simulation and experiment? Specifically, what is the value \tau used in Line 270, and how was it calculated? Relevant information would strengthen the connection between simulation and experiment.

(4) In Line 342, the authors comment that whether using native contacts or not, they cannot fold double-knotted TrmD-Tm1570. Could the authors provide more details on how non-native interactions were analyzed?

(5) It appears that the manuscript lacks simulation or experimental evidence to support the statement at Line 343: While each domain can self-tie into its native knot, this process inhibits the knotting of the other domain. Specifically, more clarification on this inhibition is needed.

I like that this puts the work on adults first. A one-off workshop will not change practice. Teachers need real experiences that help them know, care, and act. That means learning with communities, reading across traditions, and trying to change something concrete in their own setting. When teachers feel that shift in themselves, they are better at building it with students.

To make it real, give teachers time and support to do community walks led by families, visit local cultural centers, and co-plan a unit with students that brings in multiple voices. Ask them to audit one syllabus and replace or pair texts so more perspectives sit at the center. Build in reflection on bias and on who speaks and who is seen in class. Back it with structure: release time, coaching, translation for family events, and small funds for new materials. Do this over years, not weeks, and you get a curriculum that helps students learn the content and practice democratic participation.

This frames the work in the right way. The push to widen the canon and share power is not a fight of people of color against White people. It is a coalition effort to build institutions that reflect who we actually are. Expanding the canon is additive. It raises rigor and relevance by putting classic texts in conversation with authors and histories that many students have never seen in class.

To make that real, schools need shared decision making on curriculum, time and funding to review syllabi with students and community members, professional learning on inclusive teaching, and clear data on who gets access to advanced courses and who feels they belong. When many groups help choose what counts as knowledge, the result is a stronger common culture, not a smaller one.

This passage gets it right. Multicultural education is not a unit in February. It is a rebuild. If tracking and narrow test interpretations sort kids into different futures, then equity work means de-tracking core courses with strong supports, opening gates to advanced work, and using multiple measures rather than one score to judge talent. It also means teaching with the assumption that ability grows and that knowledge is made by people in contexts, so students learn to question whose voices are centered and why.

Make it real through structures, not slogans. Audit placements and discipline by subgroup and fix the patterns. Use MTSS so help arrives before referral. Pair rigorous curriculum with culturally sustaining texts. Give teachers weekly collaboration time and coaching on equitable talk moves and feedback. Fund by student need and hire, mentor, and retain a diverse staff. Plan on years, not weeks, and measure progress with belonging, access, and outcomes, not just intentions.

I find this concept interesting. I think that people who believe and advocate for multicultural education are all fighting towards a simple common goal, equity and inclusion. So I think it's interesting that they say that in order to further this field scholars need to agree on a definition. Even if individuals are not working toward the same specifics I would think that working toward multiculturalism in education in a broad sense would still further the field and bring us closer to the goal.

Local partnerships are crucial for effective near-term conservation.

AMMs symbolize climate impacts globally and culturally.

Legal limitations restrict addressing the root cause of habitat loss.

Conservation law is now relying on future climate projections.

Highlights the vulnerability of Arctic ecosystems to oil development.

Climate change alters ecological interactions and competition.

Explains that earlier melt accelerates further warming and delays refreezing.

Sea ice seasons are shifting significantly across nearly all Arctic regions.

Shows the strong reliance of Arctic communities on AMMs.

Monitoring AMMs is challenging due to their remote range and elusive behavior.

Notes that Arctic climate impacts are severe but understudied compared to other biomes.

Emphasizes the cultural dependence on these species; conservation must account for Indigenous subsistence needs.

Highlights that incomplete data and inconsistent survey methods make it hard to understand how Arctic marine mammals are doing, reflects major scientific uncertainty.

Species richness varies strongly by region, with the Atlantic sector most diverse.

eLife Assessment

This study used a conditional knockout mouse line to remove Ptbp1 in retinal progenitors and demonstrated that its deletion has no effect on retinal neurogenesis or cell fate specification, thereby challenging the prevailing view of Ptbp1 as a master regulator of neuronal fate. The data are convincing, supported by transcriptomic analysis, histology, and proliferation assays. This study is important, and the broader implications for other CNS regions warrant further investigation.

Reviewer #1 (Public review):

Summary:

The researchers sought to determine whether Ptbp1, an RNA-binding protein formerly thought to be a master regulator of neuronal differentiation, is required for retinal neurogenesis and cell fate specification. They used a conditional knockout mouse line to remove Ptbp1 in retinal progenitors and analyzed the results using bulk RNA-seq, single-cell RNA-seq, immunohistochemistry, and EdU labeling. Their findings show that Ptbp1 deletion has no effect on retinal development, since no defects were found in retinal lamination, progenitor proliferation, or cell type composition. Although bulk RNA-seq indicated changes in RNA splicing and increased expression of late-stage progenitor and photoreceptor genes in the mutants, and single-cell RNA-seq detected relatively minor transcriptional shifts in Müller glia, the overall phenotypic impact was low. As a result, the authors conclude that Ptbp1 is not required for retinal neurogenesis and development, thus contradicting prior statements about its important role as a master regulator of neurogenesis. They argue for a reassessment of this stated role. While the findings are strong in the setting of the retina, the larger implications for other areas of the CNS require more investigation. Furthermore, questions about potential reimbursement from Ptbp2 warrant further research.

Strengths:

This study calls into doubt the commonly held belief that Ptbp1 is a critical regulator of neurogenesis in the CNS, particularly in retinal development. The adoption of a conditional knockout mouse model provides a reliable way for eliminating Ptbp1 in retinal progenitors while avoiding the off-target effects often reported in RNAi experiments. The combination of bulk RNA-seq, scRNA-seq, and immunohistochemistry enables a thorough examination of molecular and cellular alterations at both embryonic and postnatal stages, which strengthens the study's findings. Furthermore, using publicly available RNA-Seq datasets for comparison improves the investigation of splicing and expression across tissues and cell types. The work is well-organized, with informative figure legends and supplemental data that clearly show no substantial phenotypic changes in retinal lamination, proliferation, or cell destiny, despite identified transcriptional and splicing modifications.

Weaknesses:

The retina-specific method raises questions regarding whether Ptbp1 is required in other CNS locations where its neurogenic roles were first proposed. Although the study performs well in transcriptome and histological analyses, it lacks functional assessments (such as electrophysiological or behavioral testing) to determine if small changes in splicing or gene expression affect retinal function.

Reviewer #2 (Public review):

Summary:

Ptbp1 has been proposed as a key regulator of neuronal fate through its role in repressing neurogenesis. In this study, the authors conditionally inactivated Ptbp1 in mouse retinal progenitor cells using the Chx10-Cre line. While RNA-seq analysis at E16 revealed some changes in gene expression, there were no significant alterations in retinal cell type composition, and only modest transcriptional changes in the mature retina, as assessed by immunofluorescence and scRNAseq. Based on these findings, the authors conclude that Ptbp1 is not essential for cell fate determination during retinal development.

Strengths:

Despite some effects of Ptbp1 inactivation (initiated around E11.5 with the onset of Chx10-Cre activity) on gene expression and splicing, the data convincingly demonstrate that retinal cell type composition remains largely unaffected. This study is highly significant since it challenges the prevailing view of Ptbp1 as a central repressor of neurogenesis and highlights the need to further investigate, or re-evaluate, its role in other model systems and regions of the CNS.

Weaknesses:

A limitation of the study is the use of the Chx10-Cre driver, which initiates recombination around E11. This timing does not permit assessment of Ptbp1 function during the earliest phases of retinal development, if expressed at that time.

Comments on revisions:

The authors have thoroughly and satisfactorily addressed all my previous comments.

Author response:

The following is the authors’ response to the original reviews.

Reviewer #1 (Public review): 

Summary: 

The researchers sought to determine whether Ptbp1, an RNA-binding protein formerly thought to be a master regulator of neuronal differentiation, is required for retinal neurogenesis and cell fate specification. They used a conditional knockout mouse line to remove Ptbp1 in retinal progenitors and analyzed the results using bulk RNA-seq, single-cell RNA-seq, immunohistochemistry, and EdU labeling. Their findings show that Ptbp1 deletion has no effect on retinal development, since no defects were found in retinal lamination, progenitor proliferation, or cell type composition. Although bulk RNA-seq indicated changes in RNA splicing and increased expression of late-stage progenitor and photoreceptor genes in the mutants, and single-cell RNA-seq detected relatively minor transcriptional shifts in Müller glia, the overall phenotypic impact was low. As a result, the authors conclude that Ptbp1 is not required for retinal neurogenesis and development, thus contradicting prior statements about its important role as a master regulator of neurogenesis. They argue for a reassessment of this stated role. While the findings are strong in the setting of the retina, the larger implications for other areas of the CNS require more investigation. Furthermore, questions about potential reimbursement from Ptbp2 warrant further research. 

Strengths: 

This study calls into doubt the commonly held belief that Ptbp1 is a critical regulator of neurogenesis in the CNS, particularly in retinal development. The adoption of a conditional knockout mouse model provides a reliable way for eliminating Ptbp1 in retinal progenitors while avoiding the off-target effects often reported in RNAi experiments. The combination of bulk RNA-seq, scRNA-seq, and immunohistochemistry enables a thorough examination of molecular and cellular alterations at both embryonic and postnatal stages, which strengthens the study's findings. Furthermore, using publicly available RNA-Seq datasets for comparison improves the investigation of splicing and expression across tissues and cell types. The work is wellorganized, with informative figure legends and supplemental data that clearly show no substantial phenotypic changes in retinal lamination, proliferation, or cell destiny, despite identified transcriptional and splicing modifications. 

We thank the Reviewer for their evaluation of the strengths of the study.

Weaknesses: 

The retina-specific method raises questions regarding whether Ptbp1 is required in other CNS locations where its neurogenic roles were first proposed. The claim that Ptbp1 is "fully dispensable" for retinal development may be toned down, given the transcriptional and splicing modifications identified. The possibility of subtle or transitory impacts, such as ectopic neuron development followed by cell death, is postulated, but not completely investigated. Furthermore, as the authors point out, the compensating potential of increased Ptbp2 warrants additional exploration. Although the study performs well in transcriptome and histological analyses, it lacks functional assessments (such as electrophysiological or behavioral testing) to determine if small changes in splicing or gene expression affect retinal function. While 864 splicing events have been found, the functional significance of these alterations, notably the 7% that are neuronalenriched and the 35% that are rod-specific, has not been thoroughly investigated. The manuscript might be improved by describing how these splicing changes affect retinal development or function. 

We have revised the text to address these points as requested.

Reviewer #2 (Public review): 

Summary: 

Ptbp1 has been proposed as a key regulator of neuronal fate through its role in repressing neurogenesis. In this study, the authors conditionally inactivated Ptbp1 in mouse retinal progenitor cells using the Chx10-Cre line. While RNA-seq analysis at E16 revealed some changes in gene expression, there were no significant alterations in retinal cell type composition, and only modest transcriptional changes in the mature retina, as assessed by immunofluorescence and scRNAseq. Based on these findings, the authors conclude that Ptbp1 is not essential for cell fate determination during retinal development. 

Strengths: 

Despite some effects of Ptbp1 inactivation (initiated around E11.5 with the onset of Chx10-Cre activity) on gene expression and splicing, the data convincingly demonstrate that retinal cell type composition remains largely unaffected. This study is highly significant since it challenges the prevailing view of Ptbp1 as a central repressor of neurogenesis and highlights the need to further investigate, or re-evaluate, its role in other model systems and regions of the CNS. 

We thank the Reviewer for their evaluation of the strengths of the study.

Weaknesses: 

A limitation of the study is the use of the Chx10-Cre driver, which initiates recombination around E11. This timing does not permit assessment of Ptbp1 function during the earliest phases of retinal development, if expressed at that time.  

We have revised the text to address the potential limitations of the use of the Chx10-Cre driver in this study.

Reviewer #1 (Recommendations for the authors):

(1) The author only selected scRNA-Seq datasets to examine the expression patterns of Ptbp1 in the retina; incorporating immunostaining analysis in the mouse retina is necessary.

Ptbp1 expression patterns in the mouse retina were performed in Fig. 1b-1d, where Ptbp1 expression was analyzed via immunostaining for Ptbp1 protein in Chx10-Cre control and Ptbp1KO retinas at E14, P1, and P30, and are quantified in Fig. 1e. 

(2) In Figure 1, Ptbp1 signals were still detected in the KO mice, with the author suggesting that this may indicate cross-reactivity with an unknown epitope. Why is this unknown epitope only detected in the ganglion cell layer? Additional antibodies are needed to confirm the staining results. Furthermore, it is essential to verify the KO at the mRNA level using PCR. 

We are unsure of the identity of this cross-reacting epitope, although it might be Ptbp2, which is enriched expressed in immature retinal ganglion cells (Fig. S1).  In any case, we do not believe that the identity of this epitope is not relevant to assessing the efficiency of Ptbp1 deletion, as it is not detectably expressed in retinal ganglion cells in any case (Fig. S1).

Although the heatmap in Figure 2B indicates a decrease in Ptbp1 levels in the KO mice, the absence of statistical data makes it difficult to evaluate the KO efficiency. 

Respectfully, we believe that Ptbp1 knockout efficiency is adequately addressed using immunohistochemistry, and that further statistical analysis is not essential here. 

Cre staining of the Chx10-Cre;Ptbp1lox/lox mice or using reporter lines is also suggested to indicate the theoretically knockout cells. Providing high-power images of the Ptbp1 staining would help readers clearly recognize the staining signals.

To clarify the identity of the knockout cells, we have updated Figure 1 to include the Chx10-CreEGFP staining which more clearly delineates the cells in which Ptbp1 is deleted. Regarding verification of the knockout, we believe additional PCR assays are not necessary, as we have already demonstrated efficient loss of Ptbp1 in Chx10-Cre-expressing cells at the RNA level by both single-cell RNA-sequencing and bulk RNA-sequencing, and also at the protein level by immunohistochemistry. Sun1-GFP Cre reporter lines are also used in Figures 1 and S2 to visualize patterns of Cre activity, a point which is now highlighted in the text. Together, these approaches provide sufficient evidence for effective Ptbp1 knockout. 

(3) The possibility of ectopic neuron formation followed by cell death is intriguing but underexplored. Consider adding apoptosis assays (e.g., TUNEL staining) at early developmental stages to test this hypothesis.

While apoptosis assays such as TUNEL staining would be helpful to address this hypothesis, we feel incorporating these additional experiments is currently beyond the scope of this study. We agree the possibility of cell death is intriguing and plan to explore this in future work.

(4) On page 4, the statement "We did not observe any significant differences ... Chx10Cre;Ptbp1lox/lox mice (Fig. 2b,c)" should refer to Fig. 3b,c instead.

We have changed the text to refer to Fig. 3b,c.

(5) The labeling in Figure 3 as "Cre-Ptbp1" is inconsistent with the figure legend "Ptbp1-Ctrl.".

This language was used because the samples for EdU staining in Figure 3 were Chx10-Cre negative Ptbp1^lox/lox mice. We have updated the language in the manuscript and figure to reflect the genotypes more clearly. 

(6) P30 mice are still sexually immature; the term "adolescent" or "juvenile" should be used instead of "adult."

We have updated the language in the text from “adult” to “adolescent” to describe P30 mice, although the retina itself is mature by this age.

Reviewer #2 (Recommendations for the authors):

(1) As mentioned in the public review, a limitation of the study is that Ptbp1 KO is not induced prior to E11. The authors should acknowledge this limitation and include in the Discussion that the use of the Chx10-Cre line does not permit evaluation of a potential role for Ptbp1 during very early stages of retinal development, should it be expressed at that time (an aspect that would be important to determine).

We and have added this limitation to the Discussion in the sentence highlighted below.

Furthermore, the use of the Chx10-Cre transgene in this study does not exclude a potential role for Ptbp1 during very early stages of retinal development prior to E11 (pg. 6).

(2) While the data convincingly show no significant changes in retinal cell type distribution in Ptbp1 mutants, the claims in the abstract and introduction that Ptbp1 is "dispensable for retinal development" or "dispensable for the process of neurogenesis" may be overstated. Indeed, the results indicate that loss of Ptbp1 function influences retinal development by promoting neurogenesis through induction of a neuronal-like splicing program in neural progenitors. Concluding solely that Ptbp1 is dispensable for retinal cell fate specification, rather than for retinal development as a whole, would thus seem more accurate.

We have updated the language in the text to reflect Ptbp1’s role in regulating retinal cell fate specification more clearly.

(3) The authors conclude from Figure 5 that "No changes in the identity or composition of any retinal cell type were observed." Which statistical test was applied to support this conclusion? The figure indicates that Müller cells comprise 10.5% of the total cell population in controls versus 8.2% in Ptbp1-KO retinas. It may be important to consider the overall distribution of glia versus all neurons (rather than each neuron subtype individually). While the observed difference (~2% more glia at the expense of neurons) appears modest, it would be important to determine whether this trend is consistent and statistically significant.

To evaluate cell type composition, we performed differential expression analysis across all major retinal cell types and compared proportional cell type representation between control and Ptbp1 KO retinas. While these analyses did not reveal marked differences in any specific cell type, we acknowledge that the scRNA-Seq dataset includes a single experimental replicate, containing two retinas in each replicate. Therefore, we cannot draw firm statistical conclusions regarding the relative distribution of glia versus neurons, and the modest difference observed in glia cell proportion should be interpreted with caution. We agree that assessing glia-to-neuron ratios across additional replicates will be important in future studies.

(4) Referringx to Figure S1 (scRNA-seq data), the authors state that Ptbp1 mRNA is robustly expressed in retinal progenitors and Müller glia in both mouse and human retina. While the immunostaining in Figure 4 indeed clearly shows strong expression in Müller cells, the scRNAseq data presented in Figure S1 do not support the claim of "robust" expression in Müller glia in the mouse retina. This is even more striking in the human data, where panels F and H show that Ptbp1 is expressed at extremely low, certainly not "robust", levels in Müller cells. The corresponding sentence in the Results section should therefore be revised to more accurately reflect the data presented in Figure S1, or be supported by complementary immunofluorescence evidence.

We thank the reviewer for this comment. We have revised this section of the Results to better reflect Fig S1, as follows:

We observe high expression levels of Ptbp1 mRNA in primary retinal progenitors in both species and Müller glia in mouse retina, with weaker expression in neurogenic progenitors, and little expression detectable in neurons at any developmental age.

(5) When mentioning potential compensation by Ptbp2, the authors may also consider discussing the possibility that compensatory mechanisms can differ between knockdown and knockout approaches. In this context, it is noteworthy that a recent study by Konar et al., Exp Eye Res, 2025 (published after the submission of the present manuscript) reports that Ptbp1 knockdown promotes Müller glia proliferation in zebrafish.

We thank the reviewer for this suggestion. To address this, we have included a section considering this possibility in the discussion section highlighted below.

It is also possible that compensatory mechanisms differ between knockdown and knockout approaches. Notably, a recent study (Konar et al. 2025) reported that Ptbp1 knockdown promotes Müller glia proliferation in zebrafish, suggesting that effects of acute reduction of Ptbp1 may not fully mirror those of complete loss-of-function. 

(6) The statistical analyses were performed using a t-test. However, this parametric test is not appropriate for experiments with low sample sizes. A non-parametric test, such as the MannWhitney test, would be more suitable in this context. Furthermore, performing statistical analysis on n = 2 (Figure 3C) is not statistically valid.

We thank the reviewer for this comment. We agree that with a small n, non-parametric tests are more appropriate. We have added additional retinas (now n=5) for the Ptbp1-KO condition in Figure 3C and reanalyzed with the appropriate non-parametric Mann-Whitney test. For all other datasets with sufficient replicates (n≥ 4/genotype), parametric tests such as unpaired t-tests remain valid, and the results are consistent with non-parametric testing. 

(7) Figure S3 is accompanied by only a brief explanation in the Results section (a single sentence despite the figure containing six panels), which makes it difficult for readers unfamiliar with this type of data to interpret.

We thank the reviewer for the suggestion. To address this, we have included a more detailed explanation of Supplementary Figure S3 to better clarify our analysis of mature neuronal and glial cell types in both Ptbp1-deficient and wild-type animals. The relevant text now reads:

Notably, splicing patterns in Ptbp1-deficient retinas showed stronger correlation with Thy1positive neurons— which exhibit low Ptbp1 expression—and minimal overlap with microglia and auditory hair cells, the adult cell types with the highest Ptbp1 levels (Fig. S3).

Gene expression and splicing changes were compared across several reference tissues: heart tissue and Thy1-positive neurons, mature hair cells, microglia, and astrocytes (Fig. S3a,b). A heatmap of differentially expressed genes showed that while Ptbp1-deficient retinas diverged from WT retinas, their expression profiles did not resemble those of fully differentiated cell types like rods, astrocytes, or adult WT retina (Fig. S3c). Consistently, Pearson correlation analysis revealed that Ptbp1-deficient and WT retinas were more similar to each other than to fully differentiated neuronal or glial populations (Fig. S3d). Splicing profile analysis further revealed that while there was high correlation of PSI between Ptbp1-deficient and WT retinas, Ptbp1deficient retinas more closely resembled Thy1-positive neurons, whereas WT retinas aligned more strongly with mature cells such as astrocytes, microglia, and auditory hair cells (Fig. S3ef). Together, these results suggest that although Ptbp1 loss induces hundreds of alternative splicing events, the magnitude of PSI changes in the KO retinas remains considerably lower than that seen in fully differentiated cell types (Extended Data 3). Thus, while a subset of splicing events overlaps with those characteristic of mature neurons or rods, the overall splicing and expression profiles of KO retinas are more similar to those of developing retinal tissue rather than terminally differentiated neuronal or glial populations.

(8) To assess progenitor proliferation, the authors performed EdU labeling experiments in P0 retinas. Is there a rationale for not examining earlier developmental time points to evaluate potential effects on early RPCs?

We thank the reviewer for this comment. We chose to perform EdU labeling experiments at P0 for several reasons. P0 represents a developmental stage where RPCs are actively proliferating and represent ~35% of all retina cells, and the retina is transitioning to intermediate-late-stage development, providing sufficient time to ensure efficient and widespread disruption of Ptbp1. Earlier embryonic timepoints were not examined here, as addressing all stages of development was beyond the scope of this current study. However, we agree that investigating whether Ptbp1 plays stage-specific roles during development on early RPCs is an important question and potential future direction.

(9) In Figure S2, panel D shows staining in GCL under the Ptbp1 condition that does not make sense and is inconsistent with panel C. If possible, the authors should provide an alternative image to prevent any confusion.

Thank you for bringing this to our attention. The image shown for Ptbp1-KO in Figure 2d shows Sun1-eGFP labeling, which labels every cell affected by the Cre condition. The genotype for this mouse was Chx10-Cre;Ptbp1lox/lox;Sun1-GFP. We apologize for any confusion and have updated the genotype in the figure legend.

(10) The authors should revise the following sentence at the end of the Discussion section, as its meaning is unclear: "...and conditions for in vitro analysis may have accurately replicated conditions in the native CNS."

We thank the reviewer for this comment and have revised this sentence in the discussion for the sentence below.

Previous studies using knockdown may have been complicated by off-target effects (Jackson et al. 2003), and conditions for in vitro analysis may not have accurately replicated conditions in the native CNS.

What causes them to close at different timings since they are both derived from the same neural tube?

Fishing pressure threatens ecosystems and CCAMLR’s political cohesion.

Success on krill management will show whether CCAMLR can rebuild trust.

Breakdown of consensus could lead to unregulated or conflicting fishing actions.

Breakdown of consensus could lead to unregulated or conflicting fishing actions.

Defining “best science” is essential for progress on krill and toothfish management.

The organization must choose between renewed cooperation or fragmentation.

CCAMLR must adapt quickly or risk failing its conservation mandate.

Record-low sea ice highlights the urgency of climate impacts on the region.

Emphasizes CCAMLR’s global significance for marine conservation.

Krill fishing shouldn’t expand until climate impacts are better understood.

Whale rebound also alters krill and penguin trends, complicating analysis.

Not all species will respond the same; some decline while others expand.

Climate change threatens both ecosystems and commercially important species in the Southern Ocean.

Strong scientific consensus links 30% protection to ecosystem recovery and climate resilience.

Data-poor, low-interest regions are ideal candidates for precautionary protection.

High-impact regions urgently need MPAs before krill catch limits are increased.

Demanding proof of harm contradicts CCAMLR’s legal precautionary mandate.

High ecological uncertainty strengthens the argument for MPAs and strict safeguards.

Despite early momentum, CCAMLR’s MPA progress has stalled due to political disagreement.

This MPA set a global precedent for protection in areas beyond national jurisdiction.

CCAMLR recognized early that MPAs were essential for biodiversity protection and ecosystem-based management.

Safeguards include monitoring, biodiversity protection, spatial management, and adaptation to environmental change.

Multinational cooperation is necessary to ensure adequate data collection and ecosystem management.

Rapid climate-driven changes complicate management decisions and require updated monitoring.

Increased catch proposals face political and scientific uncertainties; consensus is required to avoid ecosystem risk.

Pilot studies demonstrate the feasibility of spatially and temporally distributed precautionary catch limits.

Monitoring is essential to identify negative effects of fishing before they become severe.

Consensus is necessary to implement precautionary measures and ensure ecosystem resilience in a changing environment.

Effective krill management depends on adopting precautionary measures before catches increase.

Delays or disputes over science weaken CCAMLR’s core ecosystem-based management approach.

Sustainable management is expensive; relies on both governments and industry contributions.

Highlights the balance between sustainable fishing expansion and ecosystem protection.

Reminds that cooperation and science are foundational principles of the Antarctic Treaty.

Warns that cooperation is weakening, threatening the effectiveness of the Convention.

Political disagreements slowing or blocking conservation action.

Explains why krill is economically important and heavily targeted for future food resources.

CCAMLR is not just a fisheries body but part of a larger environmental governance framework.

Krill overfishing threatens predators and ecosystem stability.

Human pressure in the region is currently low, but proposed catch increases would significantly raise impact on wildlife and ecosystems.

Scientific evidence is becoming secondary to national policy priorities. This shift undermines evidence-based fisheries management and increases risk of unsustainable decisions.

Reading this chapter about crowdsourcing and “power users vs lurkers” actually make me a little uncomfortable, because I suddenly realize I am part of the problem. On platforms like Reddit, StackOverflow, or even course discussion boards, I usually just read other people’s posts and almost never answer or edit anything. I still get so much benefit from the 1% of people who do most of the work, but they don’t really get equal reward for that labor, except maybe some reputation points or social status. It feels a bit unfair that so much “invisible work” is done by a tiny group, and platforms basically depend on their free time and motivation. At the same time, I understand why lurkers exist: sometimes we are shy, or afraid to be wrong in public, or just tired. I wonder if platforms should design more gentle “on-ramps” for contribution, so it’s less scary to move from lurker to low-key contributor instead of this huge jump.

For the bibliography, I was really interested in [p31], “Disinformation as Collaborative Work: Surfacing the Participatory Nature of Strategic Information Operations.” Just from the title and description, it already changes how I think about fake news. Before, I imagined disinformation like one bad actor or one troll farm pushing lies. This paper instead frames it as a kind of “collaborative work,” where many different people and tools are involved, sometimes even regular users who don’t realize they are part of the campaign. That idea is kind of scary, because it means disinformation is not only top-down, but also bottom-up and participatory. It also connects nicely with the chapter’s point that crowdsourcing can be used both for good (like Foldit or crisis help) and for harmful goals. It makes me feel we really need better education on how to not accidentally help spread these operations.

getting closer to closure

able to share Indy0Pad Page via IPFS with hypothesis.embdeded

https://bafybeihtxbr3mkagapvdagtjrllwmoptxu6tuiufgrxmlrdqxyqwueirwq.ipfs.dweb.link/?filename=index.html&urn=/hyperpost/🌐/♖/indy0/🌐/1/gyuri/⭕/0/

able to share indy0pad created pages for annotations without needing to rely on via.hypothesis https://hypothes.is/a/1RO0RsQzEfCTlrN6ao7HOA

if you foow the above link

it would say that the page

https://hyperpost.peergos.me/1/gyuri/⭕/0

does not exists

Now create HyperPost Page that would provide access to the IPFS hosted page

either for open access

or via InerPersonal channels

wo that sharing is recorded and creates an space for

instant co-laboration

co-munnication

real time or

maintain continuity of conversations without needeing to be synchronous

You have to wonder if Costello is aware that no one believes her.

People rejected the study that went against their belief as methodologically inferior and ended up believing in their original position even more (Lord, et. al., 1979)! In other words, using data to debunk people’s beliefs or previously established opinions may not necessarily work, a tendency to guard against when conducting Planning and Controlling activities. I have experienced that a first impression from others lasted long.

You can protect yourself against this tendency by being aware of it and making a conscious effort to open your mind to new information.

Self-fulfilling prophecy. This happens when an established stereotype causes one to behave in a certain way, which leads the other party to behave in a way that confirms the stereotype For ex, because you are treating the other person more nicely, the response you get may also be nicer, which confirms your original belief that Asians are friendly

When human beings perceive themselves, they are also subject to the false consensus error. Simply put, we overestimate how similar we are to other people

1. Our visual perception definitely goes beyond the physical information available to us. Our perception is affected by our values, needs, and emotions. There are many biases that affect human perception of objects, self, and others.

you want to summarize your topic in the conlcusion paragraph and try to get the readers to want to learn more about your topic

use the 5 W's and H questions on that body paraghrap

body paragraph is to give detail about your topic, like the background

these are the 5 steps on how your intro paragraph should be organized

your introduction should have a hook sentence generally to bring audience attention

this are non-academic sources that can also help you

this is what you should look at for your research

the main focus about the essay is to get people to understand what you are trying to say

informative essay answers the five W's and H questions

it is written in the perspective of someone who is trying to find specific information about a topic

a report that relays on the results of a central research question in a very organized way

Digital archeological tools are not only used for how cool they are or look, but about how they are used each day, on the field or lab. It is about how easy they make entering finds into the database through a tab and organizing the database.

Did Robert C. Russell and Margaret King Odell specifically use the Major System as the model for their Soundex phonetic algorithm for indexing?

https://en.wikipedia.org/wiki/Soundex

There is a close relationship between the Soundex indexing scheme and the scheme behind the Major System!!!

paraphrase

paraphrase

against the argument of impeding on the amendment

Quoted

digital tools like 3D models and mapping software change the way archeologist create visual locations or sites. Rather than using hands to draw locations, these digital tools help them create accurate visual versions of the places and artifacts.

The digital tools may look or sound easy but they have built in rules and system to follow that may influence the manner archeologist think about and use the data they collect.

This also highlights the relationship between human expertise and digital algorithm, displaying it as a partnership and how this digital tool has shaped the archeological work.

This highlights the impact of algorithm in archeology. These algorithms handle large amount of archeology data and can in turn uncover patterns of human error that may have been missed.

The Mekong Delta is basically a giant rice machine. It’s super fertile and floods predictably, which is why Vietnam is one of the top rice exporters in the world. So cool!!

This really shows how Southeast Asia is built around water—rivers, coasts, islands, all of it. It explains why so much of life, culture, and even jobs revolve around rivers and the ocean.

Even though India’s tech economy is taking off and making billions, the wealth doesn’t reach everyone. It’s wild how you can have luxury tech campuses and extreme poverty existing side-by-side in the same country.

This shows how India’s tech boom didn’t just change jobs—it changed the whole vibe of cities like Bangalore. Coffee shops, night shifts, and busy streets at 8 PM all reflect how the IT industry is shaping everyday life for young people.

Said basically says naming regions from a European point of view is biased. Calling it Southwest Asia/North Africa is way more accurate and not centered around Europe.

These artists weren’t documenting reality—they were painting what Europeans wanted the “East” to look like. It helped build a really stereotypical, exotic vibe around the region.

机器学习系统本身就代表着复杂的工程挑战。然而，它们是由一些基本构建模块组成的，这些模块必须被透彻理解才能进行更复杂的实现。这种教学方法与既定的教育进程相呼应：学生在掌握分布式系统之前先精通基本算法，或者在学习高级机器学习理论之前先熟练掌握线性代数。机器学习系统同样拥有一些必要的底层组件，这些组件构成了所有后续学习的基础

expect huge differences

knowledge when shared mutiplies!

potentially enriching both the sharer and the recipients in such an exhange of sharing

Knowledge Commons

is an ide that deeply resonates with me

I prefer the term Mutual Learning (Symmathesy)

or Open Leaarning Commons

and vernturing to create the Indy (Mutual) Learning Commons

Will be rading and annotating with great interest

Stalin was like a God to them, or a savior. They put him up on a pedestal and build giant statues of him, and now they sing his praises like he is their king.

Independent projects. students apply AI ethically and strategically in revisions.

Students analyze AI’s impact. identifies over-reliance, enhances critical evaluation of revisions.

Students explain choices. builds metacognitive awareness in revision decisions.

Gradual removal of support and students practice independent revision with AI tools.

Feedback on prompt writing → teaches students to refine AI suggestions and integrate them into revisions.

Teachers demonstrate how to use AI for drafting and revising, helps students see examples of ethical and effective revision strategies.

Thesis

This passage just highlights one of the challenges digital archeology faces when introduced to the public. While the digital technology has it's benefit it can also come with risk like exaggerating the message behind certain artifacts which may be misleading.

Shows that delayed police response directly endangered women.

Reveals how hard it was for women in the 1970s to disclose abuse, even to trusted people.

Shows the frustration survivors feel when society blames them rather than the abuser.

Direct evidence contradicts the myth that women “choose” to stay.

This important statistic shows the scale of domestic violence and the need for shelters.

Shows the movement pushing for systemic reforms, not just individual solutions.

Demonstrates how feminists challenged institutions that justified or excused male violence.

This is an example of sexist media narratives that blame women for their own abuse.

This shows how the movement used education and consciousness-raising as strategies to fight domestic violence.

This links wife-battering to patriarchal structures and shows how feminists were pushing for change in this area.

The article argues that women’s domestic labour is devalued by society, limiting their economic options.

This shows how women's unpaid labour creates economic dependence, making it hard to leave their abusive partners.

This highlights how silence and shame kept women from speaking out and further reinforces the myth that abuse was rare.

This shows that domestic violence wasn't considered a problem until feminists exposed it.

experiment

added this by hand

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4 way to embed hypothesis by the indy0pad

just change the template injected and save to IPFS

AI has significant benefits but requires caution and teacher guidance.

AI gives immediate, specific feedback.

AI-generated examples teach structure, tone, and evidence.

AI helps identify and evaluate text structure.

Students compare their evaluations with AI feedback.

AI creates example texts instantly for classroom use.

AI helps with consistent rubric scoring and targeted instruction.

Balanced reading/writing improves performance. AI can help analyze texts and generate examples.

Teachers model writing, guide practice, and encourage reflection. AI speeds up modeling.

Students synthesize skills + develop voice. AI provides mentorship resources.

Students explore writing with curiosity. AI helps with inspiration.

Students master technical writing skills and AI serves as editor.

Thesis Statement

This is relevant to my project because it shows us how digital archeology is not only recorded data but it allows the archeologist imagine and communicate new stories from the past, evolving from traditional tools to modern method.

successful

talk about banking

this is a good point. hadn't thought about the inconvenience of change from a user POV

could libraries use an Indigenous system to classify their whole collection, or do we always have to separate Indigenous materials from the rest and have a special Indigenous collection elsewhere? like how AV materials are separate from paper books

Making sure threatened species are not effected by bycatch.

This conservation method effectively boosts wild populations by rescuing chicks affected by heatwaves and food shortages.

Artificial shelters could help offset lost habitat, offering shade and protection from predators, a key to improving chick survival.

Collaboration with fishers provides a practical solution to reduce accidental penguin deaths from nets.

Highlights the urgency, climate change and human interference have pushed penguins toward a mass extinction risk.

Solution that stopped further effects on penguins but still has impacts that can be felt today.

Habitat degradation increases heat stress and predation risk.

Overfishing creates food shortages, reducing chick survival and endangering entire colonies.

Human fishing removes key food such as sardines, anchovies, and krill.

Bycatch is a massive human-caused problem. Penguins drown because they cannot escape fishing nets.

Food sources that are becoming scarce due to overfishing and other factors.

Climate change and human activity disrupt the predictability of marine food sources, creating a major survival challenge.

This is relevant to my project because it displays hoe digital technology makes it possible for archeology to present large-scale data to the public.

quote

the different laws and regulations !!!

This highlights how digital tools improve archeology collection. It points out how the shift from the analogue method, which is manual, to the digital method has made data collection more accurate and efficient. This emphasizes the benefits of digital archeology.

to tell (a story, especially a story that you create by using your imagination) He was spinning yarns [=telling stories] about his adventures in the navy. She spun a tale [=made up a story] about her car breaking down to explain why she was late.

Cancel culture had become a big part of life especially in adolescence in my opinion. I first got a phone during covid so I had never been exposed to social media before that. During covid social media was kind of all we had as teenagers and so I spent a ton of time creating posts and consuming posts. I saw a lot of creators get canceled and shamed for things that they had done. Some had really done bad things and did not deserve to be famous on the platform, some were pretty minor and go blown out of proportion for the sake of entertainment on social media. This always led me to be afraid of being canceled even though I wasn't even famous, but I still would feel like my every step on social media could be criticized. I don't think that overall cancel culture is a bad thing, but it sure has led to a cancel culture in real life as well. In high school if a rumor spread about someone then that was who they were. They were cancelled. Then years later it would turn out that they didn't actually do the bad thing. It's just created a culture that no one wants to look bad or support anything bad, which is a good thing I guess because people want to be good. But it is also not facilitating very much investigation into these accusations and leading to people being cut off from society more easily and often.

This shows that with digital archeology people have easy access to collect and study data. This is because it uses free and reachable tools that everyone can use. Archeologist also tend to do their works better because of the smart systems in play. This is relevant to my topic because it shows how much better archeology is because of the introduction of digital tools.

• Add the above script tag to the top-level document
• Do not add the script tag to the iframed documents themselves, the client will do this itself.
• Opt iframes into annotation by adding the "enable-annotation" attribute:

• Author initially struggled with social skills, feeling awkward, excitable, and bullied due to abrasive behavior and sensitivity.

• Six stages of social learning show a progression from self-focused to deeply embodied connection strategies.

• Stage 1: Tried to be a dazzling, interesting, intellectual person to gain approval.

  • Emulated admired cultural figures, memorized poetry, and read complex literature.
  • Told dramatic personal stories and developed scholarly opinions.
  • This approach earned polarized approval, often distancing others because of its presentational quality.

• Stage 2: Learned to play the social game by adapting to others' social styles, especially in restaurant work.

  • Observed and mimicked the social behaviors that successful servers used.
  • Adopted flexible social roles matching the table's mood (efficient, flirtatious, etc.).
  • Made people feel comfortable by playing their "game," though still somewhat role-driven.

• Stage 3: Loosened grip on social scripts; used quirky, authentic behaviors to relax social interactions.

  • Added benign strangeness and surreal quirkiness to interactions to signal social playfulness.
  • Created moments of unexpected connection by breaking scripts sideways.
  • Learned that how things are said can be more important than what is said.

• Stage 4: Developed bodily awareness and real-time non-verbal communication like dancing with others.

  • Became attuned to subtle body language and unspoken emotional states.
  • Reacted fluidly and spontaneously to ongoing social and emotional cues.
  • This stage deepened presence beyond verbal skill to sensory social attunement.

• Stage 5: Practiced projecting love and acceptance, creating emotional openness and deep connection.

  • Embraced a meditative state of spacious openness inspired by energy healing.
  • Provided a nervous system capacity for emotional distress, enabling catharsis.
  • Found that deep listening and presence facilitated rapid emotional intimacy.

• Stage 6: Learned to moderate emotional connection, balancing openness with boundaries.

  • Recognized that constant emotional openness can be draining and misinterpreted.
  • Managed energetic boundaries and adjusted connection intensity according to context.
  • Balanced desire for connection with comfort in solitude and acceptance of varied social interaction levels.

• Author reflects on effects of spiritual openness in various social roles and the challenge of maintaining boundaries.

• Final insight: Genuine social connection evolves from crafted performance to embodied presence, balancing authenticity, empathy, and emotional limits.

In the movie "Hidden Figures", this is demonstrated; Katherine Johnson, a black woman who did the necessary calculations, was called out for adding her name to the calculations. Instead of simply putting Paul Stafford's name (who did the calculations, but did not write the report - she was also the person who ensured all calculations were correct because of her amazing intelligence), she added her name (rightfully so), but got in trouble for it. Though it was not directly stated that it was because she was a black woman, it is not hard to understand that this was the main issue for the white men. They could not fathom that a black woman signed her name on her work. Throughout the film, it is very apparent that black women's contributions were seen as irrelevant to the men.