Mgr. Dalibor Blažek, Ph.D.

Project: Regulation and function of P-TEFb complexes

Person in Charge: Assoc. prof. RNDr. Šárka Pospíšilová, Ph.D.

Host institution: Department of Internal Hematooncology, Faculty of Medicine, Masaryk University

Country of Origin: Czech Republic

Country of scientific activity: USA

Project duration: 35 months

Scientific panel: Life sciences

Abstract:

Transcription of the RNA Polymerase II (RNAPII)–dependent genes is a process consisting of several steps and is tightly regulated at many levels. Recent genome-wide ChIP-seq studies have clearly demonstrated that the transition of RNAPII from the paused state to productive elongation is a major regulatory step, especially for genes responsive to different stimuli and development. The decision about productive elongation is regulated by the phosphorylation of serine 2 in the C-terminal domain (CTD) of RNAPII by the positive transcription elongation factor b (P-TEFb). P-TEFb is a heterodimer consisting of cyclin-dependent kinase 9 (cdk9) and a cyclin subunit (i.e. cyclinT1, cyclinT2 or cyclinK). P-TEFb exists in two forms: small where “free” P-TEFb is enzymatically active, and large where kinase is inactive. The large complex (LC) consists of Hexim1, MEPCE and LARP7 proteins and 7SK small nuclear RNA. Release of the P-TEFb from its large, inactive form is tightly regulated. The mechanism by which P-TEFb is released from its LC in not very well understood, nor is the specific function of different P-TEFb kinases consisting of different cyclin subunits. In this proposal, I will study the regulation of P-TEFb complexes and role of the individual cyclin subunits in the regulation of cdk9.

The ongoing project summary

Set up of the laboratory, laboratory staff hired and guided through the experiments. Basic characterization of methylation of Heximr and clarification of the regulation of P-TEFb complex.
Brief summary of the main so far merits:
Characterization of methylation at glutamate of Heximr in steady-state situation and after treatment with agents inducing stress. Preparation of cell lines with stably integrated cyclins and discovery of the binding partners for cyclin K.

Summary of expected outputs of the project and the potential for the South Moravian region

Transfer of new methods and project with potentially clinically interesting proteins to the South Moravian region.