Affiliation:
1. Department of Cancer Genetics 1 and
2. Department of Biochemistry and Biophysics, School of Medicine and Dentistry, University of Rochester, Rochester, New York 14642 2 ; and
3. Department of Experimental Therapeutics, 3 Roswell Park Cancer Institute, Buffalo, New York 14263;
4. Max-Planck Institute for Medical Research, Molecular Neuroscience, D-69120 Heidelberg, Germany4
Abstract
ABSTRACT
Nonsense-mediated mRNA decay (NMD), also called mRNA surveillance, is an important pathway used by all organisms that have been tested to degrade mRNAs that prematurely terminate translation and, as a consequence, eliminate the production of aberrant proteins that could be potentially harmful. In mammalian cells, NMD appears to involve splicing-dependent alterations to mRNA as well as ribosome-associated components of the translational apparatus. To date, human (h) Upf1 protein (p) (hUpf1p), a group 1 RNA helicase named after its
Saccharomyces cerevisiae
orthologue that functions in both translation termination and NMD, has been the only factor shown to be required for NMD in mammalian cells. Here, we describe human orthologues to
S. cerevisiae
Upf2p and
S. cerevisiae
Upf3p (
Caenorhabditis elegans
SMG-4) based on limited amino acid similarities. The existence of these orthologues provides evidence for a higher degree of evolutionary conservation of NMD than previously appreciated. Interestingly, human orthologues to
S. cerevisiae
Upf3p (
C. elegans
SMG-4) derive from two genes, one of which is X-linked and both of which generate multiple isoforms due to alternative pre-mRNA splicing. We demonstrate using immunoprecipitations of epitope-tagged proteins transiently produced in HeLa cells that hUpf2p interacts with hUpf1p, hUpf3p-X, and hUpf3p, and we define the domains required for the interactions. Furthermore, we find by using indirect immunofluorescence that hUpf1p is detected only in the cytoplasm, hUpf2p is detected primarily in the cytoplasm, and hUpf3p-X localizes primarily to nuclei. The finding that hUpf3p-X is a shuttling protein provides additional indication that NMD has both nuclear and cytoplasmic components.
Publisher
American Society for Microbiology
Subject
Cell Biology,Molecular Biology
Cited by
213 articles.
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