Article

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

Department of Molecular Virology and Microbiology, Baylor College of Medicine, Houston, Texas 77030, USA.
Journal of leukocyte biology (Impact Factor: 4.99). 09/2009; 86(6):1345-50. DOI: 10.1189/jlb.0509309
Source: PubMed

ABSTRACT The cellular kinase complex P-TEFb is composed of Cdk9 and cyclin T, and it is required for expression of most protein-coding genes by RNAP II. Cdk9 has been shown recently to be activated in cis by autophosphorylation of Thr186 in its T-loop. Using a phosphospecific Cdk9 antibody, we examined the level of Cdk9 T-loop phosphorylation in resting and activated CD4(+) T lymphocytes. Cdk9 T-loop phosphorylation was found to be low-to-undetectable 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 result of the absence of an associated regulatory cyclin partner. These observations suggest that autophosphorylation 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 phosphorylation 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.

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Available from: Rajesh Ramakrishnan, Aug 12, 2015
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Questions & Answers about this publication

  • Rajesh Ramakrishnan added an answer in Protein Purification:
    Suggestions for immunoprecipitation kits/protocols?
    I'm somewhat new to IP and am looking for a good kit (or bead type) to isolate my proteins (large family of related proteins; we have an antibody specific for a conserved region).
    My antibody is a polyclonal IgG from rabbit. My proteins of interest range from 20kD-200kD in size (I don't know if that matters). The antibody has a decent number of primary amines.
    I'm looking to elute my proteins without the antibody, and would prefer not to denature my proteins.
    I don't have any experience with commercial IP kits, so any suggestions would be appreciated.
    Thanks!