Phosphorylation mediates the nuclear targeting of the maize Rab17 protein.
The maize abscisic acid-responsive Rab17 protein localizes to the nucleus and cytoplasm in maize cells. In-frame fusion of Rab17 to the reporter protein beta-glucuronidase (GUS) directed GUS to the nucleus and cytoplasm in transgenic Arabidopsis thaliana and in transiently transformed onion cells. Analysis of chimeric constructs identified one region between amino acid positions 66-96, which was necessary for targeting GUS to the nucleus. This region contains a serine cluster followed by a putative consensus site for protein kinase CK2 phosphorylation, and a stretch of basic amino acids resembling the simian virus 40 large T antigen-type nuclear localization signal (NLS). Mutation of two basic amino acids in the putative NLS had a weak effect on nuclear targeting in the onion cell system and did not modify the percentage of nuclear fusion protein in the Arabidopsis cells. The mutation of three amino acids in the consensus site for CK2 recognition resulted in the absence of in vitro phosphorylated forms of Rab17 and in a strong decrease of GUS enzymatic activity in isolated nuclei of transgenic Arabidopsis. These results suggest that phosphorylation of Rab17 by protein kinase CK2 is the relevant step for its nuclear location, either by facilitating binding to specific proteins or as a direct part of the nuclear targeting apparatus.
Departament de Genetica Molecular, Centre d'Investigació i Desenvolupament, (C.S.I.C.), Barcelona, Spain.
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Jensen
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Goday
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Figueras
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Jessop
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Pagès
D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D001693:Biological Transport, Active; D047390:Casein Kinase II; D002467:Cell Nucleus / Q000378:metabolism; D017931:DNA Primers / Q000235:genetics; D017930:Genes, Reporter; D005966:Glucuronidase / Q000235:genetics; D009154:Mutation; D019913:Nuclear Localization Signals; D010766:Phosphorylation; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D030821:Plants, Genetically Modified; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D003313:Zea mays / Q000235:genetics / Q000378:metabolism*
D017931:DNA Primers; D019913:Nuclear Localization Signals; D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; C071710:DHN1 protein, Zea mays; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases; D005966:Glucuronidase