ExperimentName: The Cpfl1 mouse harbors loss-of-function variants in the Pde6c gene.

NonHumanModel: Mus musculus

GeneAlteration: The model animals are homozygous for two different loss-of-function variants predicted to introduce premature stop codons into the Pde6c locus. One is a 116-bp insertion between exons 4 and 5, while the other is a 1-bp deletion in exon 7.

ModelPhenotype: HP:0008020 (cone dystrophy), HP:0030465 (Undetectable light-adapted electroretinogram), normal dark-adapted ERG, rapid and progressive loss of cone photoreceptors from the photoreceptor layer

HumanPhenotype: HP:0030465 (Undetectable light-adapted electroretinogram), HP:0030534 (Abnormal best corrected visual acuity test), HP:0000551 (Color vision defect), HP:0000613 (Photophobia), HP:0000639 (Nystagmus), HP:0007401 (Macular atrophy), HP:0000545 (Myopia)

ModelEvidence: This is a naturally occurring mouse model with lack of light-adapted ERG responses and normal dark-adapted ERG responses (Figure 1). The mice also exhibit rapid and progressive loss of cone photoreceptors from retinal sections (Figure 2).

ModelVariant: This model did not introduce a disease-associated variant known from an affected human patient, but rather studied naturally occurring variants disrupting the Pde6c locus.

ExperimentName: Naturally occurring BBS7 loss-of-function primate model

NonHumanModel: Rhesus macaque

GeneAlteration: The model animals harbor a naturally occurring homozygous frameshift variant at nucleotide 160 within exon 3 of the BBS7 locus.

ModelPhenotype: HP:0000608 (Macular degeneration), HP:0030609 (Photoreceptor layer loss on macular OCT), HP:0000580 (Pigmentary retinopathy), HP:0008046 (Abnormal retinal vascular morphology), progressive kidney disease, HP:0030483 (Reduced amplitude of dark-adapted bright flash electroretinogram a-wave), HP:0030469 (Abnormal dark-adapted electroretinogram), HP:0008275 (Abnormal light-adapted electroretinogram), HP:0002171 (Gliosis), HP:0000083 (Renal insufficiency), HP:0012210 (Abnormal renal morphology), HP:0002119 (Ventriculomegaly), HP:0008734 (Decreased testicular size)

HumanPhenotype: HP:0001956 (Truncal obesity), HP:0000618 (Blindness), HP:0001249 (Intellectual disability), HP:0001631 (Atrial septal defect), HP:0001640 (Cardiomegaly), HP:0100598 (Pulmonary edema), HP:0012584 (Bilateral renal hypoplasia), HP:0004719 (Hyperechogenic kidneys), HP:0003774 (Stage 5 chronic kidney disease), HP:0000580 (Pigmentary retinopathy), HP:0000648 (Optic atrophy), HP:0008043 (Retinal arteriolar constriction), HP:0007737 (Bone spicule pigmentation of the retina), HP:0007754 (Macular dystrophy)

ModelEvidence: This is a naturally occurring Rhesus macaque model of biallelic BBS7 disruption. The mutant animals exhibit hypoechoic kidney lesions (Figure 1), retinal atrophy with lack of retinal vasculature (Figure 3A), RPE atrophy (Figure 3B), severe atrophy of retinal layers by OCT with greater severity centrally (Figure 3C), retinal gliosis (Figure 3D), Severely reduced amplitudes and delayed time courses of both dark-adapted and light-adapted flicker ERG (Figure 4), reduced brain volume (Figure 5B), dilation of the lateral ventricle (Figure 5A), cystic kidney tubules with loss of nephrons replaced with fibrosis, tubule atrophy, and ectasia (Figure 7), reduced size and histological maturity of testes (Table 1), and vision deficits (Table 1).

ModelVariant: This model did not introduce a disease-associated variant known from an affected human patient, but rather studied a naturally occurring and apparent null variant disrupting BBS7 expression.

ExperimentName: bbs1-1 Chlamydomonas have full-length flagella and normal IFT function but are nonphototactic.

NonHumanModel: Chlamydomonas reinhardtii

GeneAlteration: The bbs1-1 model animals have been identified as naturally occurring mutants with deletion of approximately half of the BBS1 gene as well as a separate gene unstream of BBS1.

ModelPhenotype: abnormal flagellar motility (inability to perform phototaxis), flagellar absence of BBSome components (specifically BBS4)

HumanPhenotype: HP:0000639 (Nystagmus), HP:0000510 (Rod-cone dystrophy), HP:0007663 (Reduced visual acuity), HP:0000662 (Nyctalopia), HP:0000518 (Cataract), HP:0008323 (Abnormal light- and dark-adapted electroretinogram), HP:0000107 (Renal cyst), HP:0001513 (Obesity), HP:0010442 (Polydactyly), HP:0002463 (Language impairment), HP:0000026 (Male hypogonadism)

ModelEvidence: The mutants lack the ability to perform phototaxis, which is common among all BBSome mutants tested (described in text but data not shown). Flagella from the mutant show loss of BBS4 localization (Figure 4C). However, flagellar length was normal (Figure 2B) and flagellar regeneration was normal as well (described in text but data not shown).

ModelVariant: This model had a naturally occurring deletion disrupting BBS1 but did not feature knock-in of a known variant from the human patient population.

ExperimentName: Zebrafish knockdown of the GPR143 ortholog OA1

NonHumanModel: Danio rerio

GeneAlteration: A zebrafish model of OA1 loss-of-function was generated by morpholino-based knockdown.

ModelPhenotype: HP:0007703 (Abnormality of retinal pigmentation)

HumanPhenotype: HP:0000639 (Nystagmus), HP:0003593 (Infantile onset), HP:0007663 (Reduced visual acuity), HP:0001361 (Nystagmus-induced head nodding), HP:0000646 (Amblyopia), HP:0000483 (Astigmatism), HP:0000613 (Photophobia), normally pigmented skin and hair, HP:0005592 (Giant melanosomes in melanocytes), HP:0007750 (Hypoplasia of the fovea), HP:0007894 (Hypopigmentation of the fundus), HP:0025551 (Optic nerve misrouting)

ModelEvidence: The zebrafish match one feature of the human patients in that they exhibit abnormalities of retinal pigmentation consistent with the ability of GPR143 loss-of-function to trigger defects in melanosome regulation (Figure 1).

ModelVariant: This model features disruption of OA1 expression by knockdown, and does not harbor a mutation equivalent to a variant from a human patient.

ExperimentName: Canine model of Adam9 loss-of-function

NonHumanModel: Canis lupus familiaris

GeneAlteration: The affected dogs have homozygous disruption of the Adam9 gene by a deletion of regions from exons 15 and 16.

ModelPhenotype: HP:0001105 (Retinal atrophy), HP:0000529 (Progressive visual loss), HP:0000580 (Pigmentary retinopathy), HP:0000662 (Nyctalopia)

HumanPhenotype: HP:0011463 (Childhood onset), HP:0000548 (Cone/cone-rod dystrophy), HP:0007663 (Reduced visual acuity), HP:0001141 (Severely reduced visual acuity), HP:0007994 (Peripheral visual field loss), HP:0000545 (Myopia), HP:0000483 (Astigmatism), HP:0000523 (Subcapsular cataract)

ModelEvidence: The affected dogs have been described as having generalized progressive retinal atrophy. Complete loss of all retinal layers was observed in affected animals (Figures 1H-1P). This matches the mode of inheritance and progressive nature of the human disease state, as well as the retinal abnormalities and visual defects.

ModelVariant: The model harbors a targeted deletion that does not have an equivalent / orthologous variant reported in human patients.

ExperimentName: CRISPR/Cas9-based disruption of mouse Impg1

NonHumanModel: Mus musculus

GeneAlteration: Model animals were subjected to CRISPR/Cas9-based deletion of 10bp from exon 2 of Impg1, and analyzed in the homozygous state. Absence of Impg1 signal was confirmed by immunofluorescent staining. Mice were studied at ages 5 months and 8 months.

ModelPhenotype: HP:0030612 (Abnormal retinal morphology, specifically mislocalization of chondroitin sulfate in the IPM, with none in the outer segment and more in the inner segment)

HumanPhenotype: HP:0007754 (Macular dystrophy), HP:0007769 (Peripheral retinal degeneration), HP:0032037 (Mildly reduced visual acuity, progressive), HP:0008179 (Decreased Arden ratio of electrooculogram)

ModelEvidence: This is the first mouse model of Impg1 disruption, with confirmed absence of Impg1 in the homozygous animals. The only observed abnormality is mislocalization of chondroitin sulfate in the IPM, with none in the outer segment and more in the inner segment (Figure 7). A mild disruption was observed in the localization of PNA, a cone-specific marker, to the outer segment (Fig. 2F). Impg2 appears to be normally localized (Figure 2C). ERG responses were normal (data not shown). Please note that an independent study generating a different model of homozygous Impg1 disruption reported different findings (PMID: 36140676).

ModelVariant: The model does not introduce a variant previously observed in human patients, but rather disrupts the expression of the Impg1 ortholog.

ExperimentName: Mouse model of homozygous Cln5 loss

NonHumanModel: Mus musculus

GeneAlteration: Mice are homozygous for Cln5 loss of function, caused by Insertion of a neomycin resistance cassette eliminating exon 3, resulting in a premature stop codon in exon 4. The presence or absence of a truncated gene product was not assessed.

ModelPhenotype: HP:0003208 (Fingerprint intracellular accumulation of autofluorescent lipopigment storage material), HP:0003205 (Curvilinear intracellular accumulation of autofluorescent lipopigment storage material), HP:0000572 (Visual loss)

HumanPhenotype: HP:0003208 (Fingerprint intracellular accumulation of autofluorescent lipopigment storage material), HP:0003205 (Curvilinear intracellular accumulation of autofluorescent lipopigment storage material), HP:0000572 (Visual loss)

ModelEvidence: The model phenotypes closely match the human patient phenotypes at the microscopic level (characteristic autofluorescent inclusion bodies ) as well as the loss of vision, but do not reproduce neurological features such as seizures, ataxia, loss of motor coordination, mental regression, and behavioral changes.

ModelVariant: While it is unclear whether the mice express a truncated form of Cln5 or whether this form is degraded, the predicted protein product does not represent a particular human variant. The authors speculate that it may be most similar to the NM_006493.4(CLN5):c.522dup (p.Trp175LeufsTer30) variant (PMID:10953198).

NonHumanModel Mus Musculus

GeneAlteration The knock-in strain was generated by homologous recombination of the targeting vector with 129Sv/J embryonic stem cells genome. A 450-bp DNA segment encompassing 5′ UTR and exon 1 of the CASQ2 was used to screen RPCI-21 PAC mouse genomic library. Positive clones were sequenced to define the structure of an 8.5-Kb region encompassing promoter, exon 1, and part of intron 1 of mouse CASQ2. This region was cloned in 3 parts and assembled into the targeting vector pFlrt (supplemental Figure I). The linearized targeting vector was electroporated into 129Sv/J embryonic stem cells. The clone selected with G418 and gancyclovir was injected into C57BL/6NCrL blastocyts and transferred to pseudopregnant CD-1 females. Genotype was determined by sequencing of DNA extracted from tail biopsy specimens (DNasy Tissue Kit, Qiagen).

ModelPhenotype HP:0004758

HumanPhenotype HP:0004758

ModelEvidence Fig. 1 ECG during stimuli shows VT in only the mutant mice

ModelVariant NM_001232.3(CASQ2):c.97C>T (p.Arg33Ter), 41043 link

NonHumanModel Mus Musculus

GeneAlteration The knock-in strain was generated by homologous recombination of the targeting vector with 129Sv/J embryonic stem cells genome. A 450-bp DNA segment encompassing 5′ UTR and exon 1 of the CASQ2 was used to screen RPCI-21 PAC mouse genomic library. Positive clones were sequenced to define the structure of an 8.5-Kb region encompassing promoter, exon 1, and part of intron 1 of mouse CASQ2. This region was cloned in 3 parts and assembled into the targeting vector pFlrt (supplemental Figure I). The linearized targeting vector was electroporated into 129Sv/J embryonic stem cells. The clone selected with G418 and gancyclovir was injected into C57BL/6NCrL blastocyts and transferred to pseudopregnant CD-1 females. Genotype was determined by sequencing of DNA extracted from tail biopsy specimens (DNasy Tissue Kit, Qiagen).

ModelPhenotype HP:0004758

HumanPhenotype HP:0004758

ModelEvidence Fig. 1 ECG during stimuli shows VT in only the mutant mice

ModelVariant NM_001232.3(CASQ2):c.97C>T (p.Arg33Ter), 41043 link

ExperimentName: Plaa G23V/G23V homozygous mice

NonHumanModel: mus musculus

GeneAlteration: Crispr/Cas9 gene editing was used to successfully mutate the mouse Plaa gene with a patient derived variant, G23V (mouse c. 68G>T) (Figure 3 and Figure S4). From the first round of injections, the obtained 33 mice, in which 3 had the c.68G>T variation alone/ solely. Confirmation of the change was performed by Sanger sequencing in the founder mice.

ModelPhenotype: HP:0004325, HP:0002808, HP:0001337, HP:0012448

HumanPhenotype: HP:0012448

ModelEvidence: Plaa G23V homozygous mice are viable past birth, but cannot survive past 6 months without assistance of feeding, etc (Figure 3B). The mutant show microcephaly (reduced brain volume, Figure 3C). MRI performed on the G23V homozygous mice compared with control wildtype littermates show significantly reduced corpus callosum and cerebellum volumes, indicating reduced white matter (Figure 3E). The cerebellum is noted to have disrupted foliation (figure S5C). The authors note early onset neurodysfunction, including tremor and muscle weakness , showing severe hindlimb clasping, reduced grip strength and altered gait detectable from P14 (Figure 3 and Figure S5).

They authors even went on to cross the Plaa G23V heterozygous mice with Plaa homozygous null mice to create Plaa G23V/- mice that have a more severe phenotype that the homozygous Plaa G23V homozygous, but less severe that the Plaa homozygous null (plaa-/-).

EvidenceShown: Figure 3, Figure S3-6,

ModelVariant: NM_001031689.2(PLAA):c.68G>T (p.Gly23Val)

ClinVar ID: 427941

Score:

Reason: This mouse model represents a patient derived mutation that was knocked directly into the mouse Plaa locus, through use of Crispr/Cas9 gene editing. The mice were shown to have reduced Plaa protein expression (reduced by 70%), and the development of phenotypes consistent with phenotypes observed in individuals. I have therefore, increased the score based on the evidence provided.

Score: 3

ExperimentName: Plaa G23V/G23V homozygous mice

NonHumanModel: mus musculus

GeneAlteration: Crispr/Cas9 gene editing was used to successfully mutate the mouse Plaa gene with a patient derived variant, G23V (mouse c. 68G>T) (Figure 3 and Figure S4). From the first round of injections, the obtained 33 mice, in which 3 had the c.68G>T variation alone/ solely. Confirmation of the change was performed by Sanger sequencing in the founder mice.

ModelPhenotype: HP:0004325, HP:0002808, HP:0001337, HP:0012448

HumanPhenotype: HP:0012448

ModelEvidence: Plaa G23V homozygous mice are viable past birth, but cannot survive past 6 months without assistance of feeding, etc (Figure 3B). The mutant show microcephaly (reduced brain volume, Figure 3C). MRI performed on the G23V homozygous mice compared with control wildtype littermates show significantly reduced corpus callosum and cerebellum volumes, indicating reduced white matter (Figure 3E). The cerebellum is noted to have disrupted foliation (figure S5C). The authors note early onset neurodysfunction, including tremor and muscle weakness , showing severe hindlimb clasping, reduced grip strength and altered gait detectable from P14 (Figure 3 and Figure S5).

They authors even went on to cross the Plaa G23V heterozygous mice with Plaa homozygous null mice to create Plaa G23V/- mice that have a more severe phenotype that the homozygous Plaa G23V homozygous, but less severe that the Plaa homozygous null (plaa-/-).

EvidenceShown: Figure 3, Figure S3-6,

ModelVariant: NM_001031689.2(PLAA):c.68G>T (p.Gly23Val)

ClinVar ID: 427941

Score:

Reason: This mouse model represents a patient derived mutation that was knocked directly into the mouse Plaa locus, through use of Crispr/Cas9 gene editing. The mice were shown to have reduced Plaa protein expression (reduced by 70%), and the development of phenotypes consistent with phenotypes observed in individuals. I have therefore, increased the score based on the evidence provided.

Score: 3