The interplay between p16 serine phosphorylation and arginine methylation determines its function in modulating cellular apoptosis and senescence

Abstract

Cyclin-dependent kinase inhibitor p16INK4a (p16) primarily functions as a negative regulator of the retinoblastoma protein (Rb) -E2F pathway, thus plays critical role in cell cycle progression, cellular senescence and apoptosis. In this study, we showed that the methylation of Arg 138 and the phosphorylation of Ser 140 on p16 were critical for the control of cell proliferation and apoptosis. Compared to wild type p16, mutant p16R138K possessed improved function in preventing cell proliferation and inducing apoptosis, while the Ser 140 mutation (p16S140A) exhibited the opposite alteration. We also demonstrated that H2O2 was able to induce the phosphorylation of p16, which facilitated the interaction between CDK4 (Cyclin-dependent protein kinase) and p16, in 293T (human emborynic kidney) cells. Furthermore, the elevated arginine methylation in p16S140A mutant and increased serine phosphorylation in p16R138K mutant suggest that a antagonizing mechanism coordinating Arg 138 methylation and Ser 140 phosphorylation to regulates p16 function as well as cellular apoptosis and senescence. These findings will therefore contribute to therapeutic treatment for p16-related gene therapy by providing theoretical and experimental evidence.

Molecular mapping of interstitial lung disease reveals a phenotypically distinct senescent basal epithelial cell population

“…Many markers that distinguish the Basal-2 cluster from the epithelial populations as a whole continued to distinguish this population when compared directly with another basal cell population, Basal-1. Induction of CDKN2A gene expression was not observed in this population, which is aligned with the lack of clear and consistent transcriptional induction previously described in in vitro senescence models and which may reflect a difference between transcript and protein levels ( 35 , 36 ), considering the immunoreactivity for p16 protein observed in Figure 1 . Notably, there was lower expression of genes associated with regulation of both apoptosis and senescence ( LGALS1 , CAV1 , FOS , JUN , CYR61 ) in the Basal-2 cluster ( Figure 6B ).…”

“…This yielded a total of 228 coregulated senescence-associated genes shared among the different epithelial cell types (Supplemental Table 1). Of note, CDKN2A does not appear in this signature as induction was not consistent across models (data not shown), but p16 protein levels and activity can be further regulated post-transcriptionally (35,36). We employed a similar approach to generate a consensus signature for senescent fibroblasts, serving as a comparator to epithelial senescence ( Figure 3G, Supplemental Table 2).…”

Snail suppresses cellular senescence and promotes fibroblast‐led cancer cell invasion

“…Importantly, even though p16INK4A is mutated, Snail siRNAs induced cellular senescence in cancer cells and diminished their invasive properties. Moreover, we could not exclude a possible involvement of p16 phosphorylation in cellular senescence regulated by Snail .…”

Protein arginine methylation: an emerging regulator of the cell cycle

“…Interestingly, recent publications have brought to light the interplay between arginine methylation and serine phosphorylation, adding an extra layer of complexity to the regulation of the cell cycle. For example, methylation of p16 at arginine 138 by PRMT1 increases as phosphorylation of p16 at serine 140 decreases, and vice versa in 293T cells [ 107 ]. These modifications work antagonistically to regulate the interaction of p16 with CDK4 [ 107 ], thus regulating cell proliferation and apoptosis.…”

“…For example, methylation of p16 at arginine 138 by PRMT1 increases as phosphorylation of p16 at serine 140 decreases, and vice versa in 293T cells [ 107 ]. These modifications work antagonistically to regulate the interaction of p16 with CDK4 [ 107 ], thus regulating cell proliferation and apoptosis.…”