Abstract
Discovery of emerging REGγ-regulated proteins has accentuated the REGγ-proteasome as an important pathway in multiple biological processes, including cell growth, cell cycle regulation, and apoptosis. However, little is known about the regulation of the REGγ-proteasome pathway. Here we demonstrate that REGγ can be SUMOylated in vitro and in vivo by SUMO-1, SUMO-2, and SUMO-3. The SUMO-E3 protein inhibitor of activated STAT (PIAS)1 physically associates with REGγ and promotes SUMOylation of REGγ. SUMOylation of REGγ was found to occur at multiple sites, including K6, K14, and K12. Mutation analysis indicated that these SUMO sites simultaneously contributed to the SUMOylation status of REGγ in cells. Posttranslational modification of REGγ by SUMO conjugation was revealed to mediate cytosolic translocation of REGγ and to cause increased stability of this proteasome activator. SUMOylation-deficient REGγ displayed attenuated ability to degrade p21Waf//Cip1 due to reduced affinity of the REGγ SUMOylation-defective mutant for p21. Taken together, we report a previously unrecognized mechanism regulating the activity of the proteasome activator REGγ. This regulatory mechanism may enable REGγ to function as a more potent factor in protein degradation with a broader substrate spectrum.
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Acknowledgements
This work was supported by the National Institutes of Health (1R01CA131914). This manuscript was also funded in part by the National Natural Science Foundation of China (30811120435, 30870503, 81071657), the Science and Technology Commission of Shanghai Municipality (06DZ22923, 08PJ14047, 10JC1404200, 09ZZ41), and the National Basic Research Program (2009CB918402, 2011CB504200).
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( Supplementary information is linked to the online version of the paper on the Cell Research website.)
Supplementary information
Supplementary information, Figure S1
GFP can not be SUMOylated. (PDF 89 kb)
Supplementary information, Figure S2
REGγ can be SUMOylated by SUMO-1, -2, and -3. (PDF 77 kb)
Supplementary information, Figure S3
PIAS1 is the major SUMO-E3 ligase for REGγ. (PDF 147 kb)
Supplementary information, Figure S4
REGγ-SUMO fusion proteins are capable of cytoplasmic translocation. (PDF 91 kb)
Supplementary information, Figure S5
Evolutionary view of the putative SUMOylation sites in REGγ. (PDF 175 kb)
Supplementary information, Figure S6
REGγ SUMOylation occurs on multiple sites. (PDF 146 kb)
Supplementary information, Figure S7
SUMO modification enhances REGγ stability. (PDF 83 kb)
Supplementary information, Figure S8
Endogenous p21 turnover is increased in hyper-SUMOylated cell. (PDF 166 kb)
Supplementary information, Table S1
Predicted REGγ SUMOylation sites with SUMOplot (PDF 1027 kb)
Supplementary information, Table S2
Predicted REGγ SUMOylation sites with SUMOsp2.0 (PDF 1108 kb)
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Wu, Y., Wang, L., Zhou, P. et al. Regulation of REGγ cellular distribution and function by SUMO modification. Cell Res 21, 807–816 (2011). https://doi.org/10.1038/cr.2011.57
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DOI: https://doi.org/10.1038/cr.2011.57
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