Characterization of Cdk9 T-loop phosphorylation in resting and activated CD4(+) T lymphocytes

The cellular kinase complex P-TEFb is composed of Cdk9 and cyclin T, and it is required for expression of most pro-tein-coding genes by RNAP II. Cdk9 has been shown re-cently to be activated in cis by autophosphorylation of Thr186 in its T-loop. Using a phosphospecific Cdk9 anti-body, we examined the level of Cdk9 T-loop phosphoryla-tion in resting and activated CD4 T lymphocytes. Cdk9 T-loop phosphorylation was found to be low-to-undetect-able in resting CD4 T lymphocytes, and upon activation by distinct stimuli, there is a rapid (<1 h) increase in pCdk9 that does not require protein synthesis. The low level of Cdk9 T-loop phosphorylation was not to be a re-sult of the absence of an associated regulatory cyclin partner. These observations suggest that autophosphor-ylation of the Cdk9 T-loop is repressed in resting CD4 T lymphocytes. The low level of T-loop phosphorylation in resting cells is also reflected in a low level of phosphory-lation of Ser2 in the carboxyl terminal domain of RNAP II, suggesting that lack of Cdk9 T-loop autophosphorylation may limit RNAP II elongation in quiescent CD4 T lymphocytes. J. Leukoc. Biol. 86: 1345–1350; 2009