Depletion of the poly(C)-binding proteins αCP1 and αCP2 from K562 cells leads to p53-independent induction of cyclin-dependent kinase inhibitor (CDKN1A) and G1 arrest

Abstract

The α-globin poly(C)-binding proteins (αCPs) comprise an abundant and widely expressed set of K-homolog domain RNA-binding proteins. αCPs regulate the expression of a number of cellular and viral mRNAs at the levels of splicing, stability, and translation. Previous surveys have identified 160 mRNAs that are bound by αCP in the human hematopoietic cell line, K562. To explore the functions of these αCP/mRNA interactions, we identified mRNAs whose levels are altered in K562 cells acutely depleted of the two major αCP proteins, αCP1 and αCP2. Microarray analysis identified 27 mRNAs that are down-regulated and 14 mRNAs that are up-regulated in the αCP1/2-codepleted cells. This αCP1/2 co-depletion was also noted to inhibit cell proliferation and trigger a G1 cell cycle arrest. Targeted analysis of genes involved in cell cycle control revealed a marked increase in p21WAF mRNA and protein. Analysis of mRNPcomplexes in K562 cells demonstrates in vivo association of p21WAF mRNA with αCP1 and αCP2. In vitro binding assays indicate that a 127-nucleotide region of the 3′-untranslated region of p21WAF interacts with both αCP1 and αCP2, and codepletion of αCP1/2 results in a marked increase in p21WAF mRNA half-life. p21WAF induction and G1 arrest in the αCP1/2-co-depleted cells occur in the absence of p53 and are not observed in cells depleted of the individual αCP isoforms. The apparent redundancy in the actions of αCP1 and αCP2 upon p21WAF expression correlates with a parallel redundancy in their effects on cell cycle control. These data reveal a pivotal role for αCP1 and αCP2 in a p53-independent pathway of p21WAF control and cell cycle progression. © 2009 by The American Society for Biochemistry and Molecular Biology, Inc.