Research Article

The 3.8 Å structure of the U4/U6.U5 tri-snRNP: Insights into spliceosome assembly and catalysis

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Science  07 Jan 2016:
DOI: 10.1126/science.aad6466

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Splicing of pre-messenger RNA (pre-mRNA) is accomplished by a dynamic mega-complex known as the spliceosome. Assembly of a functional spliceosome requires a pre-assembled U4/U6.U5 tri-snRNP complex, which comprises the U5 small nuclear ribonucleoprotein (snRNP), U4 and U6 small nuclear RNA (snRNA), and a number of protein factors. Here we report the three-dimensional structure of a Saccharomyces cerevisiae U4/U6.U5 tri-snRNP at an overall resolution of 3.8 Å by single particle, electron cryo-microscopy (cryo-EM). The local resolution for the core regions of the tri-snRNP goes to 3.0-3.5 Å, allowing construction of a refined atomic model. Our structure contains U5 snRNA, the extensively base-paired U4/U6 snRNA, and 30 proteins including Prp8 and Snu114, which amount to 8,495 amino acids and 263 nucleotides with a combined molecular mass of approximately 1 mega-Daltons. The catalytic nucleotide U80 from U6 snRNA exists in an inactive conformation, stabilized by its base-pairing interactions with U4 snRNA and protected by Prp3. Remarkably, pre-mRNA is bound in the tri-snRNP through base-pairing interactions with U6 snRNA and Loop I of U5 snRNA. This structure, together with that of the spliceosome, reveals remarkable choreography of the snRNAs in the activation process of the spliceosomal ribozyme.

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