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PMID: 10611239 Published · ppublish English Journal Article Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S.

A nuclear 3'-5' exonuclease involved in mRNA degradation interacts with Poly(A) polymerase and the hnRNA protein Npl3p.

Molecular and cellular biology ·Vol. 20 ·No. 2 ·2000-01-00 ·Pages 604-16

Burkard KT, Butler JS

Abstract

Inactivation of poly(A) polymerase (encoded by PAP1) in Saccharomyces cerevisiae cells carrying the temperature-sensitive, lethal pap1-1 mutation results in reduced levels of poly(A)(+) mRNAs. Genetic selection for suppressors of pap1-1 yielded two recessive, cold-sensitive alleles of the gene RRP6. These suppressors, rrp6-1 and rrp6-2, as well as a deletion of RRP6, allow growth of pap1-1 strains at high temperature and partially restore the levels of poly(A)(+) mRNA in a manner distinct from the cytoplasmic mRNA turnover pathway and without slowing a rate-limiting step in mRNA decay. Subcellular localization of an Rrp6p-green fluorescent protein fusion shows that the enzyme residues in the nucleus. Phylogenetic analysis and the nature of the rrp6-1 mutation suggest the existence of a highly conserved 3'-5' exonuclease core domain within Rrp6p. As predicted, recombinant Rrp6p catalyzes the hydrolysis of a synthetic radiolabeled RNA in a manner consistent with a 3'-5' exonucleolytic mechanism. Genetic and biochemical experiments indicate that Rrp6p interacts with poly(A) polymerase and with Npl3p, a poly(A)(+) mRNA binding protein implicated in pre-mRNA processing and mRNA nuclear export. These findings suggest that Rrp6p may interact with the mRNA polyadenylation system and thereby play a role in a nuclear pathway for the degradation of aberrantly processed precursor mRNAs.

MeSH Terms
Amino Acid Sequence Catalytic Domain Cell Nucleus/enzymology,genetics Exoribonucleases/genetics,metabolism Exosome Multienzyme Ribonuclease Complex Fungal Proteins/chemistry,genetics,metabolism Genes, Fungal/genetics,physiology Half-Life Molecular Sequence Data Mutation/genetics Nuclear Proteins/metabolism Pancreatitis-Associated Proteins Polynucleotide Adenylyltransferase/genetics,metabolism Protein Binding RNA Processing, Post-Transcriptional/genetics RNA Stability/genetics RNA, Fungal/genetics,metabolism RNA, Heterogeneous Nuclear/genetics,metabolism RNA, Messenger/genetics,metabolism RNA-Binding Proteins Recombinant Fusion Proteins/metabolism Saccharomyces cerevisiae/cytology,enzymology,genetics,metabolism Saccharomyces cerevisiae Proteins Sequence Alignment Suppression, Genetic/genetics Temperature
Chemicals
Fungal Proteins NPL3 protein, S cerevisiae Nuclear Proteins Pancreatitis-Associated Proteins REG3A protein, human RNA, Fungal RNA, Heterogeneous Nuclear RNA, Messenger RNA-Binding Proteins Recombinant Fusion Proteins Saccharomyces cerevisiae Proteins Polynucleotide Adenylyltransferase Exoribonucleases Exosome Multienzyme Ribonuclease Complex RRP6 protein, S cerevisiae
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Burkard K T
Department of Microbiology, University of Rochester School of Medicine and Dentistry, Rochester, New York 14618, USA.
Butler J S
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/MCB.20.2.604-616.2000
Published
2000-01-00
Pages
604-16
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC85144
Subset
IM
Grants
NIGMS NIH HHS · R01 GM059898 · United States
NIGMS NIH HHS · GM 59898 · United States
External Links
PubMed source DOI: 10.1128/MCB.20.2.604-616.2000 PMC Full Text View journal details
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