Review coordinated by Life Science Editors. Reviewed by: Dr. Helen Pickersgill, Life Science Editors Potential Conflicts of Interest: None

Main point of the paper: The authors discover a new function for a nucleolar protein, a hydroxylase called MINA (a Myc target gene) involved in ribosome biogenesis and linked with lots of diseases (cancer, allergy), at the plasma membrane of epithelial cells. Here, it promotes epithelial tight junction integrity and barrier function, likely together with a guanylate kinase, MPP6.

Why this is interesting: This may be the first link specifically between ribosome biogenesis and adhesion, which occur in two different subcompartments of the cell, and it is mediated by one protein, MINA. The linking of these processes would enable the careful coordination that is needed between cell division and adhesion, and could thereby help cells transition between two states (i.e., from quiescence to proliferation). So, it’s the multifunctional property of MINA in two different parts of the cell that could help promote an important state transition, which may be why it’s linked to different diseases.

Background: MINA is primarily known as a nucleolar-localized protein that promotes ribosome biogenesis/function. It catalyzes histidine hydroxylation of the large ribosomal subunit protein RPL27A thereby promoting its activity. Consistent with this, MINA is upregulated in rapidly growing cells, including tumors. However, it is expressed also in normal human tissues, and has also been linked with seemingly ribosome-independent functions, specifically in suppressing cell migration, invasion, and EMT. This suggests it has other, unknown functions, which may or may not be related to each other.

Transitioning between distinct cell states such as differentiation and proliferation requires extensive coordination of diverse processes. For example, transitioning from quiescence to growth requires increasing metabolic activity and involves coupling processes like the cell cycle and adhesion to enable the cell to successfully divide.

Results: • MINA depletion in intestinal epithelial cells (Caco-2 cells stably expressing MINA-targeting shRNA) caused flattening and other phenotypes indicative of a role in tight junction integrity and barrier function. This pointed to an unexpected role for a nucleolar protein in epithelial membrane biology. • Pull-downs with a MINA mutant that was unable to localize to the nucleolus in three cell lines (Caco-2, U2OS, and MEFs) together with proteomic screens in Caco-2 cells of the WT and nucleolar-interacting mutant identified MPP6, a member of the MAGUK superfamily of cell adhesion and polarity proteins, as a candidate interacting partner of MINA. • In vitro interaction assays with endogenous and tagged proteins verified the MINA-MPP6 interaction in the nucleoplasm, and showed that it was promoted by RNApol1 inhibition (which increases the pool of extra-nucleolar MINA in subconfluent cells).<br /> • MINA knockdown redistributes HA-tagged-MPP6 from the plasma membrane to the nucleus, suggesting a functional consequence of this interaction. • shRNA knockdown of MPP6 phenocopied MINA depletion with respect to altered barrier function. i.e., both are implicated in tight junction integrity and epithelial barrier function.

Remaining questions/points: • It’s not yet demonstrated definitively that MINA works via MPP6 in tight junction integrity and epithelial barrier function. It would help to have more insight into this mechanism – it appears to not be related to MINA’s catalytic activity, and may ‘just’ involve MINA promoting MPP6 membrane localization, but whether this is direct or indirect (and if/how it’s regulated) is unclear. • Whether there is a regulatory link between ribosomal biogenesis and membrane biology via MINA is unclear – i.e., if/how MINA’s two functions are physiologically regulated to coordinate proliferation and adhesion (this may require going beyond cell lines). It could be that these two functions are normally independent (but if you experimentally interfere in vitro, then you see a dependency).

This is a single strand on an RNA molecule that leaves the the nucleus of a cell in order to relocate to the cytoplasm. This is where the mRNA can help create the protein for the cell in a process known as protein synthesis. The mRNA takes in information passed into it by DNA and decode it for the ribosomes to make more protein for the cell to live on.