Processing of the small GTP-binding protein SpYPT1p in Schizosaccharomyces pombe and in mammalian COS cells.
SpYPT1p belongs to a family of ras-like GTP-binding proteins which is believed to be involved in the regulation of intracellular vesicular trafficking. We have analyzed the processing of this protein in its natural environment, the fission yeast Schizosaccharomyces pombe, and when expressed in transfected mammalian COS cells. In COS cells SpYPT1p exists in two forms: a cytosolic 24 kDa protein which represents the unprocessed precursor form and a prenylated 23.5 kDa protein which is equally distributed between membranes and cytosol. In contrast, in S.pombe we have been unable to detect any prenylation of the protein, despite the presence of a potential C-terminal CysCys prenylation site. In addition a 23.5 kDa form was localized exclusively in the cytosol and a third membrane-bound 23 kDa form was also detected. Pulse-chase experiments revealed that in S.pombe SpYPT1p is co-translationally or immediately after translation converted to the 23.5 kDa form which is then rapidly processed to the 23 kDa membrane bound form. We have been unable to detect any significant soluble pool of the protein.
National Institute for Medical Research, The Ridgeway, Mill Hill, London, U.K.
T
T
Giannakouros
C M
CM
Newman
J
J
Armstrong
A I
AI
Magee
D000818:Animals; D002460:Cell Line; D003001:Cloning, Molecular; D005656:Fungal Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D007700:Kinetics; D008715:Methionine / Q000378:metabolism; D008798:Mevalonic Acid / Q000378:metabolism; D014176:Protein Biosynthesis; D012156:Reticulocytes / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D012568:Schizosaccharomyces / Q000235:genetics / Q000378:metabolism*; D013462:Sulfur Radioisotopes; D014157:Transcription Factors / Q000096:biosynthesis* / Q000235:genetics; D014158:Transcription, Genetic; D014162:Transfection; D014316:Tritium
D005656:Fungal Proteins; C075582:SPT3 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D013462:Sulfur Radioisotopes; D014157:Transcription Factors; D014316:Tritium; D008715:Methionine; D008798:Mevalonic Acid