Department of Molecular and Cell Biology, University of California at Berkeley, Berkeley, CA 94720, USA.
Nucleic Acids Res. 2010 Jan;38(2):360-9. doi: 10.1093/nar/gkp977. Epub 2009 Nov 11.
The 7SK snRNP represents a major reservoir of activity where P-TEFb, a general transcription factor key for RNA polymerase II elongation, can be withdrawn to promote gene expression, cell growth and development. Within this complex, 7SK snRNA is a central scaffold that coordinates key protein-protein interactions and maintains P-TEFb in an inactive state. Although the stability of 7SK directly affects the amount of active P-TEFb in vivo, relatively little is known about how it is maintained and how the 7SK methylphosphate capping enzyme MePCE and LARP7, a La-related protein associated with the 3'-poly(U) of 7SK, contribute to this process. Here, we show that 7SK is capped by the LARP7-free MePCE and in probably a co-transcriptional manner prior to its sequestration into 7SK snRNP. However, upon interacting with LARP7 within 7SK snRNP, MePCE loses its capping activity, probably due to the occlusion of its catalytic center by LARP7. Despite its lack of capping activity in 7SK snRNP, MePCE displays a capping-independent function to promote the LARP7-7SK interaction, which in turn stabilizes 7SK and facilitates the assembly of a stable MePCE-LARP7-7SK subcomplex. Our data indicate that MePCE and LARP7 act cooperatively to stabilize 7SK and maintain the integrity of 7SK snRNP.
7SK snRNP 代表了一个主要的活性储库，其中 P-TEFb（一种关键的通用转录因子，可促进 RNA 聚合酶 II 的延伸）可以被撤回以促进基因表达、细胞生长和发育。在这个复合物中，7SK snRNA 是一个中央支架，协调关键的蛋白质-蛋白质相互作用，并使 P-TEFb 保持非活性状态。尽管 7SK 的稳定性直接影响体内活性 P-TEFb 的数量，但对于它是如何维持的以及 7SK 甲基磷酸化帽酶 MePCE 和 LARP7（一种与 7SK 的 3'-poly(U) 相关的 La 相关蛋白）如何促进这一过程，相对知之甚少。在这里，我们表明 7SK 被无 LARP7 的 MePCE 帽化，并且可能是在其被隔离到 7SK snRNP 之前以共转录的方式进行的。然而，在 7SK snRNP 内与 LARP7 相互作用后，MePCE 失去了其帽化活性，可能是由于 LARP7 阻塞了其催化中心。尽管 MePCE 在 7SK snRNP 中没有帽化活性，但它显示出一种不依赖帽化的功能，以促进 LARP7-7SK 的相互作用，这反过来又稳定了 7SK，并促进了稳定的 MePCE-LARP7-7SK 亚复合物的组装。我们的数据表明，MePCE 和 LARP7 协同作用以稳定 7SK 并维持 7SK snRNP 的完整性。
