Abstract 1429: C-terminal phosphorylation of p27 promotes cell cycle dependent changes through transcriptional coregulation of cJun and STAT3

Abstract

Abstract p27 is a cyclin dependent kinase (CDK) inhibitor that binds cyclin-CDKs to arrest cell cycle. p27 undergoes PI3K/AKT-mediated C-terminal phosphorylation that alters its protein-protein interactions and function. Our prior work showed C-terminally phosphorylated p27 (p27pT157pT198) promotes its association with cJun. Genomic profiling showed p27 is co-recruited with cJun to over half of cJun chromatin binding sites to either activate or repress target genes. p27/cJun activated gene targets include TGFB2, and are associated with EMT, and programs that upregulate stem cells and alter cell adhesion and migration. Here, we pursued the hypothesis that cyclic changes in C-terminally phosphorylated p27 abundance govern p27-regulated transcriptional activity across the cell cycle, switching p27 from a corepressor of target genes in G0, to a transactivator upon PI3K/AKT activation in mid-G1. In synchronized NIH3T3 cells, we made the novel observation that p27pT197 phosphorylation is periodic, peaking in mid G1 after peak in Akt activation. Furthermore, phospho-activated cJunpS63, and phosphorylated STAT3 (STAT3pY705) levels are periodic across the cell cycle with maximal presence in mid-G1. cJun is minimally expressed in quiescence, and is strongly upregulated in early G1. While total cJun expression level remains constant at all time points after exiting G0, cJunpS63 rises sharply in G1 and then declines with cell cycle progression. Similarly, while STAT3 levels rise modestly on G0 exit, phosphorylated STAT3pY705 levels show a similar pattern to that of cJun pS63, minimal in quiescence, then increasing in mid G1. Moreover, both STATpY705 and cJunpS63 co-precipitation with total p27 and p27pT197 peak in early G1. Since complex formation is period, the action of p27/cJun/STAT3 on transcriptional regulation might also be periodic. Myc is upregulated early in G1 and is essential for G1 progression. ChIP-PCR indicate recruitment of p27 and H3K27 acetylation at MYC increased after exit from quiescence. RNA-seq of NIH3T3 cells across the cell cycle showed significant change of total transcript profiles very early after G0 exit and are distinct at each time point reflecting different phases of cell cycle. Genes governing ribosome biogenesis and protein synthetic machinery are rapidly upregulated in early G1. DNA replication/repair genes are rapidly downregulated in early G1 but rise with S/G2-M. Ongoing work investigates global target gene selection by p27/STAT3/cJun and their expression across the cell cycle, and will be presented. The potential role of p27-regulated gene expression in the coordinated progression of cells across the cell cycle has not been explored to date and might prove to be critical in cell cycle progression. Citation Format: Amir Bagheri, Seyedeh Fatemeh Razavipour, Madison Sharp, Joyce Slingerland. C-terminal phosphorylation of p27 promotes cell cycle dependent changes through transcriptional coregulation of cJun and STAT3 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1429.