Abstract
Cyclin kinase inhibitor p21 inhibits or arrests the cell cycle progression in response to DNA damage. In addition, p21 regulates the progression of apoptosis negatively or positively depending of the situation. The stability of p21 is regulated by its phosphorylation or binding with partner molecules. When cells grow without DNA damage, p21 is degraded with proteasome. In this study, we analyzed the mechanism to stabilize p21. The transient expression of various p21 deletion mutants revealed that the mutant with deletion of 15-48aa was extremely unstable. And the proteasome inhibitor, lactacystin, stabilized the del15-48. The del15-48 was unstable in the cells stably expressing 1-60aa region, indicating 1-60 did not function in trans. Fusion of the 1-60 fragment to N terminal of del15-48 stabilized the product, indicates 1-60 region in the molecule is effective for the stabilization. We next constructed the cells stably expressing del15-48. The del15-48 was unstable but stabilized by lactacystin. The gamma irradiation (6 Gy) enhanced the expression of del15-48 without elevation of mRNA level, and increased the binding with cyclinA or CDK2. Taken together, we hypothesize two mechanisms for the stabilization of p21. First, DNA damage stabilizes the p21 by phosphorylation or binding with partner molecules. Second, for the basal expression of p21, 15-48 region is essential to prevent the degradation by the proteasome.
