Molecular architecture of the human U4/U6.U5 tri-snRNP.

The U4/U6.U5 triple small nuclear ribonucleoprotein (tri-snRNP) is a major spliceosome building block. Here we describe a 7 Å 3D structure of the 1.8 MDa human tri-snRNP obtained by single-particle electron cryomicroscopy. We fit all known high-resolution structures of tri-snRNP components into the EM density map and validated them by protein crosslinking. Our model reveals how the spatial organization of Brr2 RNA helicase prevents pre-mature U4/U6 RNA unwinding in isolated human tri-snRNPs and how the Sad1 protein likely tethers Brr2 to its pre-activation position. Comparison of our model with cryo-EM 3D structures of the S. cerevisiae tri-snRNP and S. pombe spliceosome indicates that Brr2 undergoes a dramatic conformational change during spliceosome activation, and that Prp8 is also rearranged to accommodate the spliceosome's catalytic RNA network