Phosphorylation by Sky1p promotes Npl3p shuttling and mRNA dissociation.
Mammalian SR proteins are currently thought to function in mRNA export as well as splicing. They contain multiple phosphorylated serine/arginine (RS/SR) dipeptides. Although SR domains can be phosphorylated by many kinases in vitro, the physiologically relevant kinase(s), and the role(s) of these modifications in vivo have remained unclear. Npl3 is a shuttling protein in budding yeast that we showed previously to be a substrate for the mammalian SR protein kinase, SRPK1, as well as the related yeast kinase, Sky1. Here we demonstrate that Sky1p phosphorylates only one of Npl3p's eight SR/RS dipeptides. Mutation of the C-terminal RS to RA, or deletion of SKY1, results in the cytoplasmic accumulation of Npl3p. The redistribution of Npl3p is accompanied by its increased association with poly(A)+ RNA and decreased association with its import receptor, Mtr10p, in vivo. We propose that phosphorylation of Npl3p by the cytoplasmically localized Sky1p is required for efficient release of mRNA upon termination of export.
Department of Biochemistry and Biophysics, University of California, San Francisco 94143-0448, USA.
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Gilbert
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Siebel
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Guthrie
D001120:Arginine / Q000378:metabolism; D001692:Biological Transport; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000378:metabolism; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins / Q000378:metabolism*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D016601:RNA-Binding Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D012694:Serine / Q000378:metabolism; D015003:Yeasts / Q000378:metabolism*
D008164:Luminescent Proteins; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D049452:Green Fluorescent Proteins; D012694:Serine; D001120:Arginine; C490216:SKY1 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases