A complex containing the CPSF73 endonuclease and other polyadenylation factors associates with U7 snRNP and is recruited to histone pre-mRNA for 3'-end processing.
Animal replication-dependent histone pre-mRNAs are processed at the 3' end by endonucleolytic cleavage that is not followed by polyadenylation. The cleavage reaction is catalyzed by CPSF73 and depends on the U7 snRNP and its integral component, Lsm11. A critical role is also played by the 220-kDa protein FLASH, which interacts with Lsm11. Here we demonstrate that the N-terminal regions of these two proteins form a platform that tightly interacts with a unique combination of polyadenylation factors: symplekin, CstF64, and all CPSF subunits, including the endonuclease CPSF73. The interaction is inhibited by alterations in each component of the FLASH/Lsm11 complex, including point mutations in FLASH that are detrimental for processing. The same polyadenylation factors are associated with the endogenous U7 snRNP and are recruited in a U7-dependent manner to histone pre-mRNA. Collectively, our studies identify the molecular mechanism that recruits the CPSF73 endonuclease to histone pre-mRNAs, reveal an unexpected complexity of the U7 snRNP, and suggest that in animal cells polyadenylation factors assemble into two alternative complexes-one specifically crafted to generate polyadenylated mRNAs and the other to generate nonpolyadenylated histone mRNAs that end with the stem-loop.
Department of Biochemistry and Biophysics, Program in Molecular Biology and Biotechnology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.
Xiao-Cui
XC
Yang
Ivan
I
Sabath
Jan
J
Dębski
Magdalena
M
Kaus-Drobek
Michał
M
Dadlez
William F
WF
Marzluff
Zbigniew
Z
Dominski
D020816:Amino Acid Motifs; D000818:Animals; D051017:Apoptosis Regulatory Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism; D039223:Cleavage And Polyadenylation Specificity Factor / Q000235:genetics / Q000378:metabolism*; D039241:Cleavage Stimulation Factor; D006367:HeLa Cells; D006657:Histones / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D026721:RNA 3' End Processing; D012322:RNA Precursors / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D020885:Ribonucleoprotein, U7 Small Nuclear / Q000235:genetics / Q000378:metabolism*; D039221:mRNA Cleavage and Polyadenylation Factors / Q000235:genetics / Q000378:metabolism*
D051017:Apoptosis Regulatory Proteins; D002135:Calcium-Binding Proteins; D039223:Cleavage And Polyadenylation Specificity Factor; D039241:Cleavage Stimulation Factor; D006657:Histones; D009687:Nuclear Proteins; D012322:RNA Precursors; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D020885:Ribonucleoprotein, U7 Small Nuclear; D039221:mRNA Cleavage and Polyadenylation Factors; C485604:CASP8AP2 protein, human; C474466:CSTF2T protein, human; C506080:LSM11 protein, human; C101043:SYMPK protein, human