USP31 acetylation at Lys1264 is essential for its activity and cervical cancer cell growth.

Abstract

Ubiquitin-specific protease 31 (USP31) is a member of deubiquitinase family that is involved in nuclear factor-κB activation and sarcomagenesis. However, little is known about posttranslational modification in the regulation of its activity and cervical cancer cell growth. In our study, we found that the Lys1264 residue of USP31 can be modified with an acetyl group by high-resolution mass spectrometry in HeLa cell line, and site-specific mutagenesis can significantly increase USP31 ubiquitin hydrolase activity and decrease the expression of p65. When being transfected with a plasmid expressing mutated USP31, the number of cancer cells was significantly decreased. We also observed that mutated USP31 could promote apoptosis but not cell cycle by flow cytometer analysis. Overexpression of mutated USP31 could reverse the effect in USP31 knockdown cell line. To further investigate its activity in tumorigenesis, deacetylase sirtuin 1 (Sirt1) was shown to interact with USP31 by co-immunoprecipitation and blocking the function of Sirt1 by knockdown or the inhibitor nicotinamide could increase the acetylation of USP31. When Lys1264 of USP31 mutated, Sirt1 could not remove its acetylation and alter the expression level of p65. Finally, inhibition or knockdown of Sirt1 suppressed USP31 activity in HeLa cell line, leading to cisplatin-induced apoptosis resistance. Therefore, acetylation at Lys1264 suppresses USP31 activity and plays a protective role in cancer cell growth. Our study contributes to understanding the mechanism of USP31 activity regulation and its role in tumorigenesis.