Negative cooperativity between Gemin2 and RNA provides insights into RNA selection and the SMN complex's release in snRNP assembly.

Hongfei Yi,Yan Hou,Xi Kong,Yingzhi Wang,Rundong Zhang,Congcong Shen,Li Mu

doi:10.1093/nar/gkz1135

Abstract

The assembly of snRNP cores, in which seven Sm proteins, D1/D2/F/E/G/D3/B, form a ring around the nonameric Sm site of snRNAs, is the early step of spliceosome formation and essential to eukaryotes. It is mediated by the PMRT5 and SMN complexes sequentially in vivo. SMN deficiency causes neurodegenerative disease spinal muscular atrophy (SMA). How the SMN complex assembles snRNP cores is largely unknown, especially how the SMN complex achieves high RNA assembly specificity and how it is released. Here we show, using crystallographic and biochemical approaches, that Gemin2 of the SMN complex enhances RNA specificity of SmD1/D2/F/E/G via a negative cooperativity between Gemin2 and RNA in binding SmD1/D2/F/E/G. Gemin2, independent of its N-tail, constrains the horseshoe-shaped SmD1/D2/F/E/G from outside in a physiologically relevant, narrow state, enabling high RNA specificity. Moreover, the assembly of RNAs inside widens SmD1/D2/F/E/G, causes the release of Gemin2/SMN allosterically and allows SmD3/B to join. The assembly of SmD3/B further facilitates the release of Gemin2/SMN. This is the first to show negative cooperativity in snRNP assembly, which provides insights into RNA selection and the SMN complex's release. These findings reveal a basic mechanism of snRNP core assembly and facilitate pathogenesis studies of SMA.

Highlights

Small nuclear ribonucleoprotein particles are major building blocks of the spliceosome, which carries out precursor mRNA splicing in eukaryotes
The space between SmD1 and SmG in the 7S complex is too narrow for SmD3/B to fit in and the 5Sm’s RNA-binding pocket could not accommodate the small nuclear RNA (snRNA)’s Sm site conformation from U1- and U4-snRNP core structures [14]
It is unknown whether the narrowness of 5Sm in the 7S complex is a real, physiologically relevant state or just an artefact arising from crystal packing, because in the crystal lattice of the 7S complex a second Gemin2 s C-terminal domain (CTD) is located right in between SmD1 and SmG contacting both (Figure 1C) and crystal packing inducing artificial conformations is well documented [52]

Summary

Introduction

Small nuclear ribonucleoprotein particles (snRNPs) are major building blocks of the spliceosome, which carries out precursor mRNA splicing in eukaryotes. The snRNPs can be divided into two classes: Sm-class snRNPs (U1, U2, U4 and U5 snRNPs for the major spliceosome, and U11, U12, U4atac and U5 for the minor spliceosome) and Sm-like-class snRNPs (U6 and U6atac snRNPs) [1,2] After being transcribed in the nucleus, precursor snRNAs (presnRNAs) are exported into the cytoplasm, where seven Sm proteins are assembled on the Sm site, PuAUUUNUGPu, of the RNAs to form snRNP cores (Sm cores). After import into the nucleus, Sm-class snRNPs are maturated by further modification of RNA and joining of proteins specific to individual snRNP before they participate in pre-mRNA splicing

Methods

Results

Conclusion