Comparative Oncology Laboratory, University of California, Davis, California 95616, USA.
J Biol Chem. 2012 Oct 5;287(41):34410-8. doi: 10.1074/jbc.M112.384990. Epub 2012 Aug 20.
The cyclin-dependent kinase inhibitor p21(Waf1/Cip1) is a major regulator of the cell cycle and plays an important role in many cellular processes, including differentiation, stress response, apoptosis, and tumorigenesis. We previously cloned the gene encoding dog p21 and found that unlike its human ortholog, dog p21 is expressed as two isoforms, one high molecular mass band of 23 kDa and one low molecular mass band of 19 kDa. In the current study, we found that the high molecular mass band is phosphorylated, whereas the low molecular mass band is hypophosphorylated. Moreover, by generating multiple mutants of dog p21, we found that serine 123 and proline 124, which form a consensus site for proline-directed phosphorylation, are required for expression of the high molecular mass p21 isoform through phosphorylation at serine 123. Most importantly, we showed that serine 123 phosphorylation inhibits ubiquitin-independent proteasomal degradation of p21 protein and subsequently, prolongs p21 protein half-life and enhances the ability of p21 to suppress cell proliferation. Taken together, these data reveal that serine 123 phosphorylation modulates p21 protein stability and activity by suppressing ubiquitin-independent proteasomal degradation.
细胞周期蛋白依赖性激酶抑制剂 p21(Waf1/Cip1) 是细胞周期的主要调节剂，在许多细胞过程中发挥重要作用，包括分化、应激反应、细胞凋亡和肿瘤发生。我们之前克隆了编码犬 p21 的基因，发现与人类同源物不同，犬 p21 表达为两种同工型，一种是 23kDa 的高分子质量带，另一种是 19kDa 的低分子质量带。在本研究中，我们发现高分子质量带被磷酸化，而低分子质量带被低磷酸化。此外，通过生成犬 p21 的多个突变体，我们发现丝氨酸 123 和脯氨酸 124 形成脯氨酸定向磷酸化的保守位点，对于通过丝氨酸 123 磷酸化表达高分子质量 p21 同工型是必需的。最重要的是，我们表明丝氨酸 123 的磷酸化通过抑制泛素非依赖性蛋白酶体降解来抑制 p21 蛋白，随后延长 p21 蛋白半衰期并增强 p21 抑制细胞增殖的能力。总之，这些数据表明丝氨酸 123 的磷酸化通过抑制泛素非依赖性蛋白酶体降解来调节 p21 蛋白的稳定性和活性。
