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Functional analysis of the zinc finger modules of the Saccharomyces cerevisiae splicing factor Luc7.
- Source :
-
RNA (New York, N.Y.) [RNA] 2024 Jul 16; Vol. 30 (8), pp. 1058-1069. Date of Electronic Publication: 2024 Jul 16. - Publication Year :
- 2024
-
Abstract
- Identification of splice sites is a critical step in pre-messenger RNA (pre-mRNA) splicing because the definition of the exon/intron boundaries controls what nucleotides are incorporated into mature mRNAs. The intron boundary with the upstream exon is initially identified through interactions with the U1 small nuclear ribonucleoprotein (snRNP). This involves both base-pairing between the U1 snRNA and the pre-mRNA as well as snRNP proteins interacting with the 5' splice site (5'ss)/snRNA duplex. In yeast, this duplex is buttressed by two conserved protein factors, Yhc1 and Luc7. Luc7 has three human paralogs (LUC7L, LUC7L2, and LUC7L3), which play roles in alternative splicing. What domains of these paralogs promote splicing at particular sites is not yet clear. Here, we humanized the zinc finger (ZnF) domains of the yeast Luc7 protein in order to understand their roles in splice site selection using reporter assays, transcriptome analysis, and genetic interactions. Although we were unable to determine a function for the first ZnF domain, humanization of the second ZnF domain to mirror that found in LUC7L or LUC7L2 resulted in altered usage of nonconsensus 5'ss. In contrast, the corresponding ZnF domain of LUC7L3 could not support yeast viability. Further, humanization of Luc7 can suppress mutation of the ATPase Prp28, which is involved in U1 release and exchange for U6 at the 5'ss. Our work reveals a role for the second ZnF of Luc7 in splice site selection and suggests that different ZnF domains may have different ATPase requirements for release by Prp28.<br /> (© 2024 Carrocci et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.)
- Subjects :
- RNA, Small Nuclear genetics
RNA, Small Nuclear metabolism
RNA Splicing Factors metabolism
RNA Splicing Factors genetics
RNA Splice Sites
Humans
RNA-Binding Proteins metabolism
RNA-Binding Proteins genetics
RNA Splicing
RNA Precursors genetics
RNA Precursors metabolism
Alternative Splicing
Ribonucleoprotein, U1 Small Nuclear metabolism
Ribonucleoprotein, U1 Small Nuclear genetics
Introns genetics
Saccharomyces cerevisiae Proteins genetics
Saccharomyces cerevisiae Proteins metabolism
Saccharomyces cerevisiae genetics
Saccharomyces cerevisiae metabolism
Zinc Fingers
Subjects
Details
- Language :
- English
- ISSN :
- 1469-9001
- Volume :
- 30
- Issue :
- 8
- Database :
- MEDLINE
- Journal :
- RNA (New York, N.Y.)
- Publication Type :
- Academic Journal
- Accession number :
- 38719745
- Full Text :
- https://doi.org/10.1261/rna.079956.124