2016
DOI: 10.7554/elife.15910
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Insights into HIV-1 proviral transcription from integrative structure and dynamics of the Tat:AFF4:P-TEFb:TAR complex

Abstract: HIV-1 Tat hijacks the human superelongation complex (SEC) to promote proviral transcription. Here we report the 5.9 Å structure of HIV-1 TAR in complex with HIV-1 Tat and human AFF4, CDK9, and CycT1. The TAR central loop contacts the CycT1 Tat-TAR recognition motif (TRM) and the second Tat Zn2+-binding loop. Hydrogen-deuterium exchange (HDX) shows that AFF4 helix 2 is stabilized in the TAR complex despite not touching the RNA, explaining how it enhances TAR binding to the SEC 50-fold. RNA SHAPE and SAXS data w… Show more

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Cited by 48 publications
(68 citation statements)
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“…It is important to note that in the 3BLQ.pdb structure, Glu96 of CycT1 had been replaced with a glycine. Since Arg65 of CDK9 is observed to form a salt bridge interaction with Glu96 based on more recent P-TEFb structures (38,64) and our MD simulations, it is quite likely that the glycine substitute may have caused Arg65 to shift away from both pThr186 and CycT1. Consistent with this idea we observed that, although the E96A CycT1 mutation caused a modest dissociation of P-TEFb, the E96K CycT1 mutant abrogated dimer assembly Hsp90 stabilizes CDK9 in the cytoplasm of resting memory T cells.…”
Section: Formation Of the Heterodimer Interface Between Cdk9 And Cyctmentioning
confidence: 73%
“…It is important to note that in the 3BLQ.pdb structure, Glu96 of CycT1 had been replaced with a glycine. Since Arg65 of CDK9 is observed to form a salt bridge interaction with Glu96 based on more recent P-TEFb structures (38,64) and our MD simulations, it is quite likely that the glycine substitute may have caused Arg65 to shift away from both pThr186 and CycT1. Consistent with this idea we observed that, although the E96A CycT1 mutation caused a modest dissociation of P-TEFb, the E96K CycT1 mutant abrogated dimer assembly Hsp90 stabilizes CDK9 in the cytoplasm of resting memory T cells.…”
Section: Formation Of the Heterodimer Interface Between Cdk9 And Cyctmentioning
confidence: 73%
“…It also increases Tat-P-TEFb affinity for TAR by 30-fold favoring TAR recognition and binding [111]. Finally, the structure of TAR in complex with Tat/P-TEFb/AFF4 was recently solved albeit a 5.9-Å low-resolution [114]. Backed-up by various structural approaches such as small angle X-ray scattering, hydrogen-deuterium exchange, and 2′-hydroxyl acylation analyzed by primer (SHAPE), the structural study revealed a direct interaction of the TAR central loop with CycT1 TRM and the Tat Cys-rich motif containing the Zn 2+ coordinating loop (Tat 24–29 ), but detected no direct interaction with AFF4 [114].…”
Section: The Super Elongation Complexmentioning
confidence: 99%
“…Instead, AFF4 indirectly enhances TAR binding to the SEC by stabilizing the Tat/CycT1 interaction. While not directly observed due to poor resolution, modeling allowed positioning of the amino acid main chain of the Tat ARM within the major groove of TAR, in contact with both the TAR bulge and the RNA phosphate backbone [114]. …”
Section: The Super Elongation Complexmentioning
confidence: 99%
“…SAXS, like EM, is often used in combination with other techniques for integrative modelling. Recently, SAXS, HDX and RNA SHAPE (selective 2’-hydroxyl acylation analysed by primer extension) data based restraints were used to provide a dynamic model of a machinery involved in HIV-1 pro-viral transcription [46]. The SHAPE data highlights RNA segments that are potentially involved in interaction based on changes in reactivity of nucleotides upon assembly.…”
Section: Approaches To Integrative Modellingmentioning
confidence: 99%