TY - JOUR
T1 - Cloning and characterization of HUPF1, a human homolog of the Saccharomyces cerevisiae nonsense mRNA-reducing UPF1 protein
AU - Applequist, S E
AU - Selg, M
AU - Raman, C
AU - Jäck, H M
N1 - Funding Information:
We thank Laura Hartwell for help in preparing rabbit antisera, Drs G.Napolitano, C.Blobel, G.Weskamp, P.Tucker, G.Wu and S.Elledge for genomic and cDNA libraries, Drs T.Ellis, R.Pieper and R.Shankar for human cell lines, Dr M.Wabl for discussions and Drs I.Haas and S.Amero for critical reading of the manuscript. This work was supported in part by the National Institutes of Health (R29CA56772-01A1), the Cancer Society of America and the Tobacco Research Council of America.
PY - 1997/2/15
Y1 - 1997/2/15
N2 - Levels of most nonsense mRNAs are normally reduced in prokaryotes and eukaryotes when compared with that of corresponding functional mRNAs. Genes encoding polypeptides that selectively reduce levels of nonsense mRNA have so far only been identified in simple eukaryotes. We have now cloned a human cDNA whose deduced amino acid sequence shows the highest degree of homology to that of UPF1, a bona fide Saccharomyces cerevisiae group I RNA helicase required for accelerated degradation of nonsense mRNA. Based on the total sequence of the shorter yeast UPF1 protein, the overall identity between the human protein and UPF1 is 51%. Besides NTPase and other RNA helicase consensus motifs, UPF1 and its human homolog also share similar putative zinc finger motifs that are absent in other group I RNA helicases. Northern blot analysis with the human cDNA probe revealed two transcripts in several human cell lines. Further, antibodies raised against a synthetic peptide of the human polypeptide detected a single 130 kDa polypeptide on Western blots from human and mouse cells. Finally, immunofluorescence and Western blot analyses revealed that the human and mouse polypeptides, like yeast UPF1, are expressed in the cytoplasm, but not in the nucleus. We have thus identified the first mammalian homolog of yeast UPF1, a protein that regulates levels of nonsense mRNA, and we tentatively name this protein human HUPF1 (for human homolog of UPF1).
AB - Levels of most nonsense mRNAs are normally reduced in prokaryotes and eukaryotes when compared with that of corresponding functional mRNAs. Genes encoding polypeptides that selectively reduce levels of nonsense mRNA have so far only been identified in simple eukaryotes. We have now cloned a human cDNA whose deduced amino acid sequence shows the highest degree of homology to that of UPF1, a bona fide Saccharomyces cerevisiae group I RNA helicase required for accelerated degradation of nonsense mRNA. Based on the total sequence of the shorter yeast UPF1 protein, the overall identity between the human protein and UPF1 is 51%. Besides NTPase and other RNA helicase consensus motifs, UPF1 and its human homolog also share similar putative zinc finger motifs that are absent in other group I RNA helicases. Northern blot analysis with the human cDNA probe revealed two transcripts in several human cell lines. Further, antibodies raised against a synthetic peptide of the human polypeptide detected a single 130 kDa polypeptide on Western blots from human and mouse cells. Finally, immunofluorescence and Western blot analyses revealed that the human and mouse polypeptides, like yeast UPF1, are expressed in the cytoplasm, but not in the nucleus. We have thus identified the first mammalian homolog of yeast UPF1, a protein that regulates levels of nonsense mRNA, and we tentatively name this protein human HUPF1 (for human homolog of UPF1).
KW - Amino Acid Sequence
KW - Animals
KW - B-Lymphocytes
KW - Base Sequence
KW - Chromosomes, Human, Pair 19
KW - Cloning, Molecular
KW - Codon, Nonsense
KW - Cytoplasm/metabolism
KW - DNA, Complementary/analysis
KW - Fungal Proteins/biosynthesis
KW - Gene Library
KW - Humans
KW - Hybridomas
KW - Mice
KW - Molecular Sequence Data
KW - RNA Helicases
KW - RNA Nucleotidyltransferases/biosynthesis
KW - RNA, Messenger/metabolism
KW - Saccharomyces cerevisiae/genetics
KW - Saccharomyces cerevisiae Proteins
KW - Sequence Homology, Amino Acid
KW - Trans-Activators
KW - Tumor Cells, Cultured
UR - http://www.scopus.com/inward/record.url?scp=0030745114&partnerID=8YFLogxK
U2 - 10.1093/nar/25.4.814
DO - 10.1093/nar/25.4.814
M3 - Article
C2 - 9064659
SN - 0305-1048
VL - 25
SP - 814
EP - 821
JO - Nucleic Acids Research
JF - Nucleic Acids Research
IS - 4
ER -