SUMOylation of Mouse p53b by SUMO-1 Promotes Its Pro-Apoptotic Function in Ovarian Granulosa Cells

Li-Jun Huo,Xiao-Ming Liu,Yi-Feng Yuan,Rui Zhai,Fei-Fei Yang,Qing-Yuan Sun

doi:10.1371/journal.pone.0063680

Li-Jun Huo, Xiao-Ming Liu + Show 4 more

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https://doi.org/10.1371/journal.pone.0063680

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Journal: PLoS ONE	Publication Date: May 16, 2013
Citations: 10	License type: CC BY 4.0

Affiliation: Huazhong Agricultural University

Abstract

Follicular atresia is a process of spontaneous degradation of follicles, hindering growth and development in the mammalian ovary. Previous studies showed that follicular atresia was caused by apoptosis of granulosa cells, for which a number of apoptosis-related genes have already been identified. The roles of p53 in apoptosis of mouse granulosa cells and its post-translational modification are still unclear. The main objective of this study was to explore the roles of p53 in mouse granulosa cells. We found that mouse p53b, but not p53a, could be SUMOylated by SUMO-1 at lysine 375, which was essential for the protein stability of p53b in a dose-dependent manner. Immunofluorescent staining showed that wild p53b was located in the nucleus of granulosa cells, while its mutation of SUMOylated site (K375R) was localized in both nucleus and cytoplasm, implying that SUMOylation was necessary for the nuclear localization of p53b in granulosa cells. Overexpression of wild-type p53b, but not the mutation of SUMOylation site (K375R), significantly induced the expression of apoptosis-related gene, Bax, and increased the level of apoptosis in granulosa cells. This suggested that SUMO-1 modification of p53b was essential for inducing apoptosis in granulosa cells. Our results provide strong evidences that modification of p53b by SUMO-1 at lysine 375 was necessary for its activity to induce apoptosis in mouse granulosa cells, and it was involved in the regulation of p53b protein stability and nuclear localization. This implies that modification of p53b by SUMO-1 might regulate follicular atresia by inducing the apoptosis of ovarian granulosa cells in mice.

Highlights

During the process of follicular atresia, follicles spontaneously degrade in the mammalian ovary, hindering growth and development
After transfection with HA-sumo-1 or HA-ubc9 plasmid, a shifting-up band detected by using anti-HA antibody was visible (Fig. 1, lanes 5 and 6), which was consistent with a form of protein that was covalently modified by HA-small ubiquitin-related modifier-1 (SUMO-1) or HA-UBC9
This study clearly showed that mouse p53 protein could be SUMO-1 modified in granulosa cells in vivo

Summary

Introduction

During the process of follicular atresia, follicles spontaneously degrade in the mammalian ovary, hindering growth and development. More than 99% of follicles disappear, primarily due to apoptosis of granulosa cells [1,2,3,4,5,6]. Atresia can occur at any time during follicular development, the majority of follicles become atretic during the early antral stage of development [7]. The transition from preantral to antral follicles occurs after the granulosa cells are exposed to gonadotropin. The gonadotropin initiates differentiation of the granulosa cells making them susceptible to apoptosis. Many apoptosis-related factors have been implicated in follicular atresia, including death ligands and receptors, intracellular pro- and anti-apoptotic molecules, cytokines, growth factors, and a number of apoptosis-related genes, including the p53 gene [9,10,11]

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