title: Structural basis of pre-mRNA recognition by the human cleavage factor Im complex.
abstract: The cleavage factor I(m) (CF I(m)), consists of a 25 kDa subunit (CF I(m)25) and one of three larger subunits (CF I(m)59, CF I(m)68, CF I(m)72), and is an essential protein complex for pre-mRNA 3'-end cleavage and polyadenylation. It recognizes the upstream sequence of the poly(A) site in a sequence-dependent manner. Here we report the crystal structure of human CF I(m), comprising CF I(m)25 and the RNA recognition motif domain of CF I(m)68 (CF I(m)68RRM), and the crystal structure of the CF I(m)-RNA complex. These structures show that two CF I(m)68RRM molecules bind to the CF I(m)25 dimer via a novel RRM-protein interaction mode forming a heterotetramer. The RNA-bound structure shows that two UGUAA RNA sequences, with anti-parallel orientation, bind to one CF I(m)25-CF I(m)68RRM heterotetramer, providing structural basis for the mechanism by which CF I(m) binds two UGUAA elements within one molecule of pre-mRNA simultaneously. Point mutation and kinetic analyses demonstrate that CF I(m)68RRM can bind the immediately flanking upstream region of the UGUAA element, and CF I(m)68RRM binding significantly increases the RNA-binding affinity of the complex, suggesting that CF I(m)68 makes an essential contribution to pre-mRNA binding.
authors.affiliation: Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, University of Science and Technology of China, Hefei, Anhui 230026, China.
authors.forename: Heng
authors.initials: H
authors.lastname: Li
authors.forename: Shuilong
authors.initials: S
authors.lastname: Tong
authors.forename: Xu
authors.initials: X
authors.lastname: Li
authors.forename: Hui
authors.initials: H
authors.lastname: Shi
authors.forename: Zheng
authors.initials: Z
authors.lastname: Ying
authors.forename: Yongxiang
authors.initials: Y
authors.lastname: Gao
authors.forename: Honghua
authors.initials: H
authors.lastname: Ge
authors.forename: Liwen
authors.initials: L
authors.lastname: Niu
authors.forename: Maikun
authors.initials: M
authors.lastname: Teng
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D018360:Crystallography, X-Ray; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011061:Poly A / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D012322:RNA Precursors / Q000737:chemistry / Q000378:metabolism*; D016415:Sequence Alignment; D039221:mRNA Cleavage and Polyadenylation Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D012322:RNA Precursors; C450562:cleavage factor Im, human; D039221:mRNA Cleavage and Polyadenylation Factors; D011061:Poly A