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Mol Cell. 2001 Jan;7(1):217-26.

Dynamic exchanges of RNA interactions leading to catalytic core formation in the U12-dependent spliceosome.

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Department of Molecular Biophysics, and Biochemistry, Yale University School of Medicine, Howard Hughes Medical Institute, New Haven, CT 06536-0812, USA.


Important general insights into the mechanism of pre-mRNA splicing have emerged from studies of the U12-dependent spliceosome. Here, photochemical cross-linking analyses during U12-dependent spliceosome assembly have surprisingly revealed that an upstream 5' exon region is required for establishing two essential catalytic core interactions, U12/U6atac helix Ib and U6atac/5' splice site contacts, but not for U5/5' exon interactions or partial unwinding of U4atac/U6atac. A novel intermediate, representing an alternative pathway for catalytic core formation, is a ternary snRNA complex containing U4atac/U6atac stem II and U12/U6atac helix Ia that forms even without U6atac replacing U11 at the 5' splice site. A powerful oligonucleotide displacement method suggests that the blocked complexes analyzed to deduce the interdependence of these multiple RNA exchanges are authentic intermediates in U12-dependent spliceosome assembly.

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