Reference: Schwer B, et al. (2013) Structure-function analysis of the 5' end of yeast U1 snRNA highlights genetic interactions with the Msl5*Mud2 branchpoint-binding complex and other spliceosome assembly factors. Nucleic Acids Res 41(15):7485-500

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Abstract

Yeast pre-mRNA splicing initiates via formation of a complex comprising U1 snRNP bound at the 5' splice site (5'SS) and the Msl5?Mud2 heterodimer engaged at the branchpoint (BP). Here, we present a mutational analysis of the U1 snRNA, which shows that although enlarging the 5' leader between the TMG cap and the (3)ACUUAC(8) motif that anneals to the 5'SS is tolerated, there are tight constraints on the downstream spacer between (3)ACUUAC(8) and helix 1 of the U1 fold. We exploit U1 alleles with 5' extensions, variations in the (3)ACUUAC(8) motif, downstream mutations and a longer helix 1 to discover new intra-snRNP synergies with U1 subunits Nam8 and Mud1 and the trimethylguanosine (TMG) cap. We describe novel mutations in U1 snRNA that bypass the essentiality of the DEAD-box protein Prp28. Structure-guided mutagenesis of Msl5 distinguished four essential amino acids that contact the BP sequence from nine other BP-binding residues that are inessential. We report new synthetic genetic interactions of the U1 snRNP with Msl5 and Mud2 and with the nuclear cap-binding subunit Cbc2. Our results fortify the idea that spliceosome assembly can occur via distinct genetically buffered microscopic pathways involving cross-intron-bridging interactions of the U1 snRNP?5'SS complex with the Mud2?Msl5?BP complex.

Reference Type
Journal Article
Authors
Schwer B, Chang J, Shuman S
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