A Comprehensive Biochemical and Genetic Analysis of the Yeast U1 SnRNP Reveals Five Novel Proteins

Overview

Journal RNA

Specialty Molecular Biology

Date 1998 Jun 18

PMID 9630245

Citations 92

Authors

A Gottschalk

J Tang

O Puig

J Salgado

G Neubauer

H V Colot

M Mann

B Seraphin

M Rosbash

R Luhrmann

P Fabrizio

Affiliations

Soon will be listed here.

Abstract

The U1 snRNP is essential for recognition of the pre-mRNA 5'-splice site and the subsequent assembly of the spliceosome. Yeast U1 snRNP is considerably more complex than its metazoan counterpart, which suggests possible differences between yeast and metazoa in early splicing events. We have comprehensively analyzed the composition of yeast U1 snRNPs using a combination of biochemical, mass spectrometric, and genetic methods. We demonstrate the specific association of four novel U1 snRNP proteins, Snu71p, Snu65p, Nam8p, and Snu56p, that have no known metazoan homologues. A fifth protein, Npl3p, is an abundant cellular component that reproducibly co-purifies with the U1 snRNP, but its association is salt-sensitive. Therefore, we are unable to establish conclusively whether it binds specifically to the U1 snRNP. Interestingly, Nam8p and Npl3p were previously assigned functions in (pre-m)RNA-metabolism; however, so far, no association with U1 snRNP has been demonstrated or proposed. We also show that the yeast SmB protein is a U1 snRNP component. Yeast U1 snRNP therefore contains 16 different proteins, including seven snRNP core proteins, three homologues of the metazoan U1 snRNP-specific proteins, and six yeast-specific U1 snRNP proteins. We have simultaneously continued the characterization of additional mutants isolated in a synthetic lethal (MUD) screen for genes that functionally cooperate with U1 snRNA. Consistent with the biochemical results, mud10, mud15, and mud16 are alleles of SNU56, NAM8, and SNU65, respectively. mud10 and mud15 affect the in vivo splicing efficiency of noncanonical introns. Moreover, mud10p strongly affects the in vitro formation of splicing complexes, and extracts from the mud15 strain contain a U1 snRNP that migrates aberrantly on native gels. Finally, we show that Nam8p/Mud15p contributes to the stability of U1 snRNP.

Citing Articles

Mechanisms and regulation of spliceosome-mediated pre-mRNA splicing in Saccharomyces cerevisiae.

Senn K, Hoskins A Wiley Interdiscip Rev RNA. 2024; 15(4):e1866.

PMID: 38972853 PMC: 11585973. DOI: 10.1002/wrna.1866.

Human PRPF39 is an alternative splicing factor recruiting U1 snRNP to weak 5' splice sites.

Espinosa S, De Bortoli F, Li X, Rossi J, Wagley M, Lo H RNA. 2022; .

PMID: 36316087 PMC: 9808567. DOI: 10.1261/rna.079320.122.

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Activation of Prp28 ATPase by phosphorylated Npl3 at a critical step of spliceosome remodeling.

Yeh F, Chang S, Ahmed G, Liu H, Tung L, Yeh C Nat Commun. 2021; 12(1):3082.

PMID: 34035302 PMC: 8149812. DOI: 10.1038/s41467-021-23459-4.

The SR-protein Npl3 is an essential component of the meiotic splicing regulatory network in Saccharomyces cerevisiae.

Sandhu R, Sinha A, Montpetit B Nucleic Acids Res. 2021; 49(5):2552-2568.

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