Study on the development and differentiation of endoplasmic reticulum of yeast from the view point of lipid synthesis.

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The aim of this research is to elucidate the mechanism of development and differentiation of endoplasmic reticulum (ER) of Saccharomyces cerevisiae, especially in relation to the organization of phospholipid synthetic enzymes that reside mainly in ER.Results are as follows,(1) We succeeded in development of a gentle subcellular fractionation method that employs D-sorbitol density gradient centrifugation.(2) We prepared antibodies specific to N-terminaus of phosphatidylserine synthase (PSS) and specific to phosphorylethanolamine cytidylyltransferase of S.cervisiae.(3) A series of fusion genes coding for N-terminal regions of PSS and Escherichia coli beta-galactosidase were constructed. In addition to them, a deletion PSS that lacks N-terminal 50 amino acid residues was constructed. From their localization analysis, both the hydrophilic N-terminal region and a hydrophobic region from 115th to 129th were necessary for the proper ER localization of PSS.(4) A pss ect mutant contained low amaount of phosphatidylethanolamine (PE). This double mutant was temperature-sensitive in growth and had altered ER structures, suggesting the necessity of PE in proper ER morphology.