Hypo-phosphorylation of the retinoblastoma protein (pRb) by cyclin D:Cdk4/6 complexes results in active pRb

Publisher Website
Google Scholar
Abstract
In cycling cells, the retinoblastoma protein (pRb) is un- and/or hypo-phosphorylated in early G 1 and becomes hyper-phosphorylated in late G 1 . The role of hypo-phosphorylation and identity of the relevant kinase(s) remains unknown. We show here that hypo-phosphorylated pRb associates with E2F in vivo and is therefore active. Increasing the intracellular concentration of the Cdk4/6 specific inhibitor p15 INK4b by transforming growth factor β treatment of keratinocytes results in G 1 arrest and loss of hypo-phosphorylated pRb with an increase in unphosphorylated pRb. Conversely, p15 INK4b -independent transforming growth factor β-mediated G 1 arrest of hepatocellular carcinoma cells results in loss of Cdk2 kinase activity with continued Cdk6 kinase activity and pRb remains only hypo-phosphorylated. Introduction of the Cdk4/6 inhibitor p16 INK4a protein into cells by fusion to a protein transduction domain also prevents pRb hypo-phosphorylation with an increase in unphosphorylated pRb. We conclude that cyclin D:Cdk4/6 complexes hypo-phosphorylate pRb in early G 1 allowing continued E2F binding.