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Control of Membrane Fusion During Yeast Mating

  • Author(s): Engel, Alex
  • Advisor(s): Walter, Peter K
  • et al.
Abstract

To maintain the integrity and identity of membrane-bound compartments, membrane fusion is promoted by fusases in a non-leaky manner. During yeast mating, two partners of a mating pair must fuse their cell membranes. The multipass membrane protein Prm1p promotes this extracytoplasmic fusion event. Less than half of prm1 x prm1 mating pairs successfully fuse, instead arresting at the step of membrane fusion or undergoing simultaneous cell lysis. prm1 x prm1 mating pair lysis is enhanced when extracellular Ca2+ is removed. As revealed by time-lapse microscopy, mating pair fusion and lysis events initiate with identical kinetics. Furthermore, cytoplasmic continuity is generated concurrent with cell lysis. These results suggest that cell lysis is linked to the mechanism of cell fusion. Prm1p promotes cell fusion and prevents cell lysis as a covalently linked homodimer. Substitution of cys-120 and cys-545 prevents covalent dimerization but does not decrease association of Prm1p monomers. Mutants unable to form the covalent link between dimers are severely compromised for Prm1p activity. PRM1 mutants show a unique genetic interaction with KEX2, a gene encoding a late-Golgi protease. Though prm1 x wt and wt x kex2 (MATa x MATα) mating pairs do not have strong mating defects, prm1 x kex2 mating pairs fuse with less than 50% efficiency. Unfused wt x kex2 mating pairs exhibit membrane enclosed structures embedded in the cell wall separating mating partners ("blebs"). When PRM1 is deleted, some prm1 x prm1kex2 mating pairs extend protrusions into the space of the partner cell that are apparently devoid of cytoplasmic contents ("enormous barren bubbles"). Another mating-induced membrane protein, Fig1p, is required for efficient cell membrane fusion during yeast mating. fig1 x fig1 mating pairs have a mild bilateral fusion defect, and 10% of unfused mating pairs arrest at the step of membrane fusion. PRM1, FIG1, and KEX2 all act during late steps of yeast mating; however, mutants of both KEX2 and FIG1 are pleiotropic and are likely to influence membrane fusion indirectly. In contrast, Prm1p is intimately involved in controlling the fidelity of membrane fusion and may constitute a part of the yeast cell fusion machinery.

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