Tat & Rev

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 Project Leaders: James HurleyAlan Frankel & Matija Peterlin

The HIV regulatory protein Tat enhances transcription from the viral promotor by increasing processivity of RNA polymerase II (RNAP II) complexes. Using the proteomics pipeline, we found that Tat selectively recruits P-TEFb as part of a large stoichiometric complex containing additional transcription elongation factors, termed the super elongation complex (SEC) (Figure 1).

Structural and functional studies of complexes containing P-TEFb, AFF4, Tat and other key components are revealing how Tat recruits the elongation factors required for efficient HIV transcription.

Figure 1. Schematic showing assembly and recruitment of Super Elongation Complex to elongating Pol II via different functional modules

 

 

The Rev project, headed by Alan Frankel, is aimed at determining structures of Rev in complex with its various binding partners, using NMR, crystallography and cryo-EM. Among other studies, we have published a native preparation method that yields discrete hexameric Rev-RRE complexes amenable to biophysical, biochemical and structural characterization, in contrast to previous filamentous Rev-RRE complexes.

The native Rev oligomer forms an exquisitely specific complex with the RRE, having 500-fold higher affinity than any monomeric Rev-RNA complex.

A structure of the Rev dimer revealed a dimer arrangement that organizes the molecule for RNA binding/recognition. The structure also suggests a model for how Rev oligomers can be arranged to act as an adaptor to the host export machinery (Figure 1).

Studies of Rev in its various complexes are ongoing in the center, with a particular emphasis on Crm1-RanGTP nuclar export complexes and other novel factors being identified in the HARC Center proteomics effort