View Program - 63rd Annual Drosophila Research Conference

Ribosome ubiquitylation is a highly regulated process that is essential for maintaining proper protein translation. While much is known about the role of ubiquitin in these pathways, less is known about the function of deubiquitinases. OTUD6 is an ovarian tumor (OTU) family deubiquitinase that is conserved from yeast (OTU2) to humans (OTUD6A & B). To study OTUD6 in Drosophila, CRISPR/Cas9 was used to create epitope-tagged wild-type and catalytically dead (DUB-dead) endogenous OTUD6. DUB-dead and loss-of-function OTUD6 mutants are markedly sensitive to oxidizing (paraquat) and alkylating (mms) agents that impact protein translation and ribosome function. Co-immunoprecipitation coupled with mass spectrometry in OTUD6.DUB-dead brains, where OTUD6 is highly expressed, identified 40S ribosome and RNA exosome proteins. The most significantly enriched 40S proteins, RACK1 and RPS3, play key roles in ribosome quality control and turnover under cellular stress. Mutation of RACK1 rescued the mms sensitivity of OTUD6 mutants, indicating that OTUD6 is likely a negative regulator of RACK1. RACK1 regulates ubiquitylation of several 40S ribosome subunits: a genetic screen of E3 ligases that ubiquitylate the 40S ribosome in a RACK1 dependent manner revealed that OTUD6.DUB-dead interacts epistatically with two E3 ligases. Both E3 ligases are implicated in RpS3 ubiquitylation and in ribosome quality control. In support of OTUD6 regulating ribosome quality control or turnover, OTUD6 mutants have decreased protein translation, delayed development, and dramatically extended lifespan. Thus, we hypothesize that OTUD6 regulates protein translation through a novel deubiquitylation event on the 40S ribosomal subunit in response to stress conditions.

The OTUD6 deubiquitinase associates with the 40S ribosome to regulate translation and the response to stressors in Drosophila