An Atomic Structure of the Human Spliceosome

Abstract

Mechanistic understanding of pre-mRNA splicing requires detailed structural information on various states of the spliceosome. Here we report the cryo electron microscopy (cryo-EM) structure of the human spliceosome just before exon ligation (the C complex) at an average resolution of 3.76 Å. The splicing factor Prp17 stabilizes the active site conformation. The step II factor Slu7 adopts an extended conformation, binds Prp8 and Cwc22, and is poised for selection of the 3'-splice site. Remarkably, the intron lariat traverses through a positively charged central channel of RBM22; this unusual organization suggests mechanisms of intron recruitment, confinement, and release. The protein PRKRIP1 forms a 100-Å α helix linking the distant U2 snRNP to the catalytic center. A 35-residue fragment of the ATPase/helicase Prp22 latches onto Prp8, and the quaternary exon junction complex (EJC) recognizes upstream 5'-exon sequences and associates with Cwc22 and the GTPase Snu114. These structural features reveal important mechanistic insights into exon ligation.

The Exon Junction Complex Undergoes a Compositional Switch that Alters mRNP Structure and Nonsense-Mediated mRNA Decay Activity

“…While haploinsufficiency of EIF4A3, RBM8A, and MAGOH lead to premature neuronal differentiation and apoptosis in the mutant brains (Mao et al, 2016; McMahon et al, 2016), similar reduction in CASC3 levels (or even its near complete depletion) does not cause the same defects but leads to a more general developmental delay (Mao et al, 2017). We note that a view contrary to our findings is presented by the recently reported human spliceosome C* structure, where CASC3 is seen bound to the trimeric EJC core (Zhang et al., 2017). As the spliceosomes described in these structural studies were assembled in vitro in nuclear extracts, it is possible that CASC3 present in extracts can enter pre-assembled spliceosomes and interact with EJC.…”

New moonlighting functions of mitochondrial cytochrome c in the cytoplasm and nucleus

“…The coordinates files of SET/TAF‐Iβ and NRP1 were obtained using homology models based on PDB ID 2E50 , as described previously . The following protein structures were constructed as homology models with SWISS‐MODEL: STRAP (based on 3JAP ), eiF2γ (based on 6FEC, ), BiP1 (based on 6HAB, ), RD21 (based on 5EGW, ), TCL (based on 6ES4, ), and Sm/D1 (based on 5XJC, ).…”

The cryo-EM structure of the SF3b spliceosome complex bound to a splicing modulator reveals a pre-mRNA substrate competitive mechanism of action

“…Proteomics studies show that SF3b proteins are depleted by 70% in the C complex relative to the B act complex (Agafonov et al 2011). Moreover, the cryo-EM structure of the human C * complex revealed that the SF3b proteins were absent (Zhang et al 2017), pointing to their departure. These data are all consistent with the paradigm that SF3B1 splicing modulators act early in spliceosome assembly by weakening interactions between the U2 snRNP and the pre-mRNA, likely impeding the formation of the "A complex" (Roybal and Jurica 2010;Corrionero et al 2011;Folco et al 2011;Effenberger et al 2014;Vigevani et al 2017).…”