A single point mutation in cyclin T1 eliminates binding to Hexim1, Cdk9 and RNA but not to AFF4 and enforces repression of HIV transcription

Retrovirology

Ran Taube²  Olivier Bensaude¹  Maayan Maatook²  Sigal Galker²  Nina Verstraete¹  Alona Kuzmina² 

[1]Ecole Normale Supérieure, Institut de Biologie de l’ENS (IBENS), and Inserm U1024, and CNRS UMR 8197, Paris F-75005, France

[2]The Shraga Segal Department of Microbiology, Immunology and Genetics Faculty of Health Sciences, Ben-Gurion University of the Negev, P.O. Box 653, Beer-Sheva 84105, ISRAEL

关键词: Super Elongation Complex;    Positive Transcription Elongation Factor b;    Tat;    Transcription repression;    HIV latency;   

Others  :  1131845
DOI  :  10.1186/1742-4690-11-51

 received in 2014-02-17, accepted in 2014-06-10,  发布年份 2014

【 摘 要 】

Background

Human immunodeficiency virus (HIV) gene expression is primarily regulated at the step of transcription elongation. The viral Tat protein recruits the Positive Transcription Elongation Factor b (P-TEFb) and the Super Elongation Complex (SEC) to the HIV promoter and enhances transcription by host RNA polymerase II.

Results

To map residues in the cyclin box of cyclin T1 that mediate the binding of P-TEFb to its interacting host partners and support HIV transcription, a pool of N-terminal cyclin T1 mutants was generated. Binding and functional assays in cells identified specific positions in cyclin T1 that are important for (i) association of P-TEFb with Hexim1, Cdk9 and SEC/AFF4 (ii) supporting Tat-transactivation in murine cells and (iii) inhibition of basal and Tat-dependent HIV transcription in human cells. Significantly, a unique cyclin T1 mutant where a Valine residue at position 107 was mutated to Glutamate (CycT1-V107E) was identified. CycT1-V107E did not bind to Hexim1 or Cdk9, and also could not assemble on HIV TAR or 7SK-snRNA. However, it bound strongly to AFF4 and its association with HIV Tat was slightly impaired. CycT1-V107E efficiently inhibited HIV replication in human T cell lines and in CD4(+) primary cells, and enforced HIV transcription repression in T cell lines that harbor a transcriptionally silenced integrated provirus.

Conclusions

This study outlines the mechanism by which CycT1-V107E mutant inhibits HIV transcription and enforces viral latency. It defines the importance of N-terminal residues of cyclin T1 in mediating contacts of P-TEFb with its transcription partners, and signifies the requirement of a functional P-TEFb and SEC in mediating HIV transcription.

【 授权许可】

   
2014 Kuzmina et al.; licensee BioMed Central Ltd.

【 图 表 】

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