Abstract
The 7SK small nuclear RNA is a highly conserved non-coding RNA that regulates transcriptional elongation. 7SK utilizes the HEXIM proteins to sequester the transcription factor P-TEFb by a mechanism similar to that used by retroviral TAR RNA to engage Tat and P-TEFb. Tat has also recently been shown to bind 7SK directly and recruit P-TEFb to TAR. We report here the solution structures of the free and arginine-bound forms of stem loop 4 of 7SK (7SK-SL4). Comparison of the 7SK-SL4 and TAR structures demonstrates the presence of a common arginine sandwich motif. However, arginine binding to 7SK-SL4 is mechanistically distinct and occurs via docking into a pre-organized pocket resulting in a 1000-fold increased affinity. Furthermore, whereas formation of the binding pocket in TAR requires a critical base-triple, hydrogen-bond formation between the equivalent bases in 7SK-SL4 is not essential and the pocket is stabilized solely by a pseudo base-triple platform. In addition, this theme of preformed protein binding motifs also extends into the pentaloop. The configuration of the loop suggests that 7SK-SL4 is poised to make ternary contacts with P-TEFb and HEXIM or Tat. These key differences between 7SK-SL4 and TAR present an opportunity to understand RNA structural adaptation and have implications for understanding differential interactions with Tat.
Highlights
RNA polymerase II is highly regulated at all phases of transcription
A preformed protein-binding pocket in the bulge is composed of a pseudo-triple platform and it is noteworthy that hydrogen bond formation is not required for complex formation
It is subjected to an entropic penalty that reduces its binding affinity by three orders of magnitude compared to 7SK-SL4
Summary
RNA polymerase II is highly regulated at all phases of transcription. Recent work has elucidated interactions of Tat with various cellular host factors, one of which is a direct interaction between 7SK and Tat.[12,13] More significantly, Tat has been shown to bind 7SK RNA by expelling HEXIM from the ternary complex, prompting a re-evaluation of our understanding of P-TEFb equilibrium between host and viral transcription. Recent work has elucidated interactions of Tat with various cellular host factors, one of which is a direct interaction between 7SK and Tat.[12,13] More significantly, Tat has been shown to bind 7SK RNA by expelling HEXIM from the ternary complex, prompting a re-evaluation of our understanding of P-TEFb equilibrium between host and viral transcription. 13; 14
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