MAMMALIAN CELL SURFACE GENETICS AND BIOCHEMISTRY

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We will continue to study the structure, functions, and dynamics of the surface membrane components of eukaryotic cells. We have used bacterial and plant toxins as probes to investigate the pathways for the entry and delivery of these toxins into the cytosol of Chinese hamster ovary (CHO) cells. We have employed these toxins to select somatic cell mutants defective in the intoxication process. Studies in the next five years will be directed towards (1) studies on the fatty acylation of membrane glycoproteins in CHO and yeast mutants altered in N- glycosylation; (2) isolation and characterization of CHO mutants resistant to ionophores monensin and nigericin; (3) isolation and characterization of Vero cell mutants defective in diphtheria toxin receptor; and (4) isolation and characterization of CHO mutants defective in the uptake, transport and release of toxins into the cytosol, including the conditionally lethal mutants. Techniques to be employed include molecular and somatic cell genetics and biochemical analysis of the structures, functions and dynamics of membrane components involved in the receptor- mediated endocytosis of toxins. Results from these studies should provide a better understanding of the intracellular protein transport process and the role of post-translational modification of membrane glycoproteins in cellular physiology.