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bioRxiv · 10.1101/2024.11.01.621267

The short conserved region-2 of LARP4 interacts with ribosome-associated RACK1 and promotes translation

Ranjan, A.·Mattijssen, S.·Charlly, N.·Cruz Gallardo, I.·Pitman, L. F.·Coleman, J. C.·Conte, M. R.·Maraia, R. J.
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Abstract

LARP4 interacts with poly(A)-binding protein (PABP) to protect mRNAs from deadenylation and decay, and recent data indicate it can direct the translation of functionally related mRNA subsets. LARP4 was known to bind RACK1, a ribosome-associated protein, although the specific regions involved, and relevance had been undetermined. Here, yeast two-hybrid domain mapping followed by other methods identified positions 615-625 in conserved region-2 (CR2) of LARP4 (and LARP4B) as directly binding RACK1 region 200-317. Consistent with these results, AlphaFold2-multimer predicted high confidence interaction of CR2 with RACK1 propellers 5-6. CR2 mutations strongly decreased LARP4 association with cellular RACK1 and ribosomes by multiple assays, whereas less effect was observed for PABP association, consistent with independent interactions. CR2 mutations decreased LARP4 ability to optimally stabilize a {beta}-globin mRNA reporter containing an AU-rich element (ARE) more significantly than a {beta}-globin and other reporters lacking this element. While polysome profiles indicate the {beta}-glo-ARE mRNA is inefficiently translated, consistent with published data, we show that LARP4 increases its translation whereas the LARP4-CR2 mutant is impaired. Analysis of nanoLuc-ARE mRNA for production of luciferase activity confirmed LARP4 promotes translation efficiency while CR2 mutations are disabling. Thus, LARP4 CR2-mediated interaction with RACK1 can promote translational efficiency of some mRNAs.

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Ranjan, A., Mattijssen, S., Charlly, N., Cruz Gallardo, I., Pitman, L. F., Coleman, J. C., Conte, M. R., Maraia, R. J.. 2024-11-01. The short conserved region-2 of LARP4 interacts with ribosome-associated RACK1 and promotes translation. https://doi.org/10.1101/2024.11.01.621267

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