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Göttingen, Germany

Schneider M.,MPI of Biophysical Chemistry | Will C.L.,MPI of Biophysical Chemistry | Anokhina M.,MPI of Biophysical Chemistry | Tazi J.,Montpellier University | And 2 more authors.
Molecular Cell | Year: 2010

The first step in splicing of pre-mRNAs with long introns is exon definition, where U1 and U2 snRNPs bind at opposite ends of an exon. After exon definition, these snRNPs must form a complex across the upstream intron to allow splicing catalysis. Exon definition and conversion of cross-exon to cross-intron spliceosomal complexes are poorly understood. Here we demonstrate that, in addition to U1 and U2 snRNPs, cross-exon complexes contain U4, U5, and U6 (which form the tri-snRNP). Tri-snRNP docking involves the formation of U2/U6 helix II. This interaction is stabilized by a 5′ splice site (SS)-containing oligonucleotide, which can bind the tri-snRNP and convert the cross-exon complex into a cross-intron, B-like complex. Our data suggest that the switch from cross-exon to cross-intron complexes can occur directly when an exon-bound tri-snRNP interacts with an upstream 5′SS, without prior formation of a cross-intron A complex, revealing an alternative spliceosome assembly pathway. © 2010 Elsevier Inc. All rights reserved.

Verbeeren J.,University of Helsinki | Niemela E.H.,University of Helsinki | Turunen J.J.,University of Helsinki | Will C.L.,MPI of Biophysical Chemistry | And 3 more authors.
Molecular Cell | Year: 2010

Alternative pre-mRNA splicing is typically regulated by specific protein factors that recognize unique sequence elements in pre-mRNA and affect, directly or indirectly, nearby splice site usage. We show that 5′ splice site sequences (5′ss) of U12-type introns, when repeated in tandem, form a U11 snRNP-binding splicing enhancer, USSE. Binding of U11 to the USSE regulates alternative splicing of U2-type introns by activating an upstream 3′ss. The U12-type 5′ss-like sequences within the USSE have a regulatory role and do not function as splicing donors. USSEs, present both in animal and plant genes encoding the U11/U12 di-snRNP-specific 48K and 65K proteins, create sensitive switches that respond to intracellular levels of functional U11 snRNP and alter the stability of 48K and 65K mRNAs. We conclude that U11 functions not only in 5′ss recognition in constitutive splicing, but also as an activator of U2-dependent alternative splicing and as a regulator of the U12-dependent spliceosome. © 2010 Elsevier Inc. All rights reserved.

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