Stoichiometry of the Sm proteins in yeast spliceosomal snRNPs supports the heptamer ring model of the core domain.

Seven Sm proteins (B/B', D1, D2, D3, E, F and G proteins) containing a common sequence motif form a globular core domain within the U1, U2, U5 and U4/U6 spliceosomal snRNPs. Based on the crystal structure of two Sm protein dimers we have previously proposed a model of the snRNP core domain consisting of a ring of seven Sm proteins. This model postulates that there is only a single copy of each Sm protein in the core domain. In order to test this model we have determined the stoichiometry of the Sm proteins in yeast spliceosomal snRNPs. We have constructed seven different yeast strains each of which produces one of the Sm proteins tagged with a calmodulin-binding peptide (CBP). Further, each of these strains was transformed with one of seven different plasmids coding for one of the seven Sm proteins tagged with protein A. When one Sm protein is expressed as a CBP-tagged protein from the chromosome and a second protein was produced with a protein A-tag from the plasmid, the protein A-tag was detected strongly in the fraction bound to calmodulin beads, demonstrating that two different tagged Sm proteins can be assembled into functional snRNPs. In contrast when the CBP and protein A-tagged forms of the same Sm protein were co-expressed, no protein A-tag was detectable in the fraction bound to calmodulin. These results indicate that there is only a single copy of each Sm protein in the spliceosomal snRNP core domain and therefore strongly support the heptamer ring model of the spliceosomal snRNP core domain. Copyright 2001 Academic Press.