PP2B And PP1α Cooperatively Disrupt 7SK SnRNP To Release P-TEFb For Transcription In Response To Ca²⁺ Signaling

In eukaryotic cells,the transcription of messenger RNA(mRNA) from protein-coding genes is performed by RNA polymeraseâ…¡.Two major forms of RNA Polâ…¡exist in eukaryotic cells:the hypophosphorylated Polâ…¡a or the hyperphosphorylated RNA Polâ…¡o.In Polâ…¡o,the carboxy-terminal domain(CTD) of the largest subunit(Rpb1) of Polâ…¡is extensively phosphorylated.In eukaryotes,two major cyclin-dependent kinases,CDK7,and CDK9 specifically target and phosphorylate the CTD of RNA Polâ…¡.While CDK7 phosphorylates Ser5 of the CTD,CDK9 targets Ser2 for phosphorylation.To accommodate different growth conditions and transcriptional demands,a reservoir of P-TEFb is kept in an inactive state in the multisubunit 7SK snRNP.Under certain stress or disease conditions,P-TEFb is released to activate transcription,although the signaling pathway(s) that controls this is largely unknown.Here,through analyzing the UV- or hexamethylene bisacetamide(HMBA)-induced release of P-TEFb from 7SK snRNP,an essential role for the calcium ion(Ca²⁺)-calmodulin-protein phosphatase 2B(PP2B) signaling pathway is revealed.However,Ca²⁺ signaling alone is insufficient,and PP2B must act sequentially and cooperatively with protein phosphatase 1α(PPα) to disrupt 7SK snRNP.Activated by UV/HMBA and facilitated by a PP2B-induced conformational change in 7SK snRNP,PP1αreleases P-TEFb through dephosphorylating phospho-Thr186 in the Cdk9 T-loop.This event is also necessary for the subsequent recruitment of P-TEFb by the bromodomain protein Brd4 to the preinitiation complex,where Cdk9 remains unphosphorylated and inactive until after the synthesis of a short RNA.Thus,through cooperatively dephosphorylating Cdk9 in response to Ca²⁺ signaling,PP2B and PP1αalter the P-TEFb functional equilibrium through releasing P-TEFb from 7SK snRNP for transcription.