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Evidence that phosphorylation of human Upf1 protein varies with intracellular location and is mediated by a wortmannin-sensitive and rapamycin-sensitive PI 3-kinase-related kinase signaling pathway

Published online by Cambridge University Press:  07 February 2001

MAHADEB PAL ,
YASUHITO ISHIGAKI ,
ESZTER NAGY  and
LYNNE E. MAQUAT
MAHADEB PAL
Affiliation:
Department of Cancer Genetics, Roswell Park Cancer Institute, Buffalo, New York 14263, USA Present address: The Cleveland Clinic, Department of Molecular Biology, Lerner Research Institute, 9500 Euclid Avenue, Cleveland, Ohio 44195, USA.
YASUHITO ISHIGAKI
Affiliation:
Department of Cancer Genetics, Roswell Park Cancer Institute, Buffalo, New York 14263, USA Department of Biochemistry and Biophysics, School of Medicine and Dentistry, University of Rochester, Rochester, New York 14642, USA
ESZTER NAGY
Affiliation:
Department of Cancer Genetics, Roswell Park Cancer Institute, Buffalo, New York 14263, USA Present address: Intercell, Inc., Dr Bohr Gasse 1R, A-1030 Vienna, Austria.
LYNNE E. MAQUAT
Affiliation:
Department of Cancer Genetics, Roswell Park Cancer Institute, Buffalo, New York 14263, USA Department of Biochemistry and Biophysics, School of Medicine and Dentistry, University of Rochester, Rochester, New York 14642, USA
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Abstract

Human Upf1 protein (p), a group 1 RNA helicase, has recently been shown to function in nonsense-mediated mRNA decay (NMD) in mammalian cells. Here, we demonstrate that the estimated 3 × 106 copies of hUpf1p per exponentially growing HeLa cell are essentially equally distributed among polysomal, subpolysomal, and ribosome-free fractions. We also demonstrate that hUpf1p binds RNA and is a phosphoprotein harboring phosphoserine and phosphothreonine. hUpf1p is phosphorylated to the highest extent when polysome-associated and to the lowest extent when ribosome free. We find that serum-induced phosphorylation of hUpf1p is inhibited by wortmannin at a concentration that selectively inhibits PI 3-kinase related kinases and, to a lesser extent, by rapamycin. These and other data suggest that phosphorylation is mediated by a wortmannin-sensitive and rapamycin-sensitive PI 3-kinase-related kinase signaling pathway. Comparisons are made of hUpf1p to Upf1p and SMG-2, which are the orthologs to hUpf1p in Saccharomyces cerevisiae and Caenorhabditis elegans, respectively.

Keywords

human Upf1 proteinnonsense-mediated mRNA decayphosphorylationP1 3-kinase-related kinaseRNA helicase
Type
Research Article
Information
RNA , Volume 7 , Issue 1 , January 2001 , pp. 5 - 15
Copyright
© 2001 RNA Society

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