UC San Diego

The process of protein targeting to mitochondria is complex and relies on both polypeptide and mRNA mediated pathways in a partially redundant manner. Mitochondria associated RNA-binding proteins (RBPs) have emerged as key facilitators of post-transcriptional regulation of nuclear-encoded mitochondrial mRNAs (NEMmRNAs), often by enhancing protein targeting to mitochondria. With only a few examples described in humans, I set out to identify RBPs with a role in post-transcriptional regulation of NEMmRNAs. Through a systematic analysis of the ENCODE collection of eCLIP datasets mapping the binding sites of over a 150 RBPs, I identified several RBPs with RNA-target sets enriched in NEMmRNAs, of these I selected LARP4 for further analysis. From data produced in multiple human cell lines, I show that the RNA-target set of LARP4 is particularly enriched in mRNAs that encode oxidative phosphorylation proteins (OXPPs) and mitochondrial ribosome proteins (MRPs). I generate LARP4 knockout cell lines to study the consequences of LARP4 depletion. By immunoblot analysis and quantitative proteomics analysis, I show that OXPPs and MRPs are present in reduced abundance in LARP4 knockout cells. Interestingly, cell fractionation experiments showed that the depletion of OXPPS and MRPs was more pronounced in mitochondrial fractions than in whole cell fractions, suggesting defect in protein targeting to mitochondria. Additionally, I show that the depletion of these proteins essential to oxidative phosphorylation has a functional impact on oxidative phosphorylation rates as well as energy dependent cellular processes such as cell proliferation and translation. Furthermore, I show these phenotypes are ameliorated by LARP4 re-expression. Together these data support a novel functional role of LARP4 in post-transcriptionally regulating NEMmRNAs to promote their expression and support oxidative phosphorylation function.