Genetic interactions between KAR7/SEC71, KAR8/JEM1, KAR5, and KAR2 during nuclear fusion in Saccharomyces cerevisiae

During mating of Saccharomyces cerevisiae, two nuclei fuse to produce a sin gle diploid nucleus. Two genes, KAR7 and KAR8, were previously identified b y mutations that cause defects in nuclear membrane fusion. KAR7 is allelic to SEC71, a gene involved in protein translocation into the endoplasmic ret iculum. Two other translocation mutants, sec63-1 and sec72 Delta, also exhi bited moderate karyogamy defects. Membranes from kar7/sec71 Delta and sec72 Delta, but not sec63-1, exhibited reduced membrane fusion in vitro, but on ly at elevated temperatures. Genetic interactions between kar7 and kar5 mut ations were suggestive of protein-protein interactions. Moreover, in sec71 mutants, Kar5p was absent from the SPB and was not detected by Western blot or immunoprecipitation of pulse-labeled protein. KAR8 is allelic to JEMI, encoding an endoplasmic reticulum resident DnaJ protein required for nuclea r fusion. Overexpression of KAR8/JEM1 (but not SEC63) strongly suppressed t he mating defect of kar2-1, suggesting that Kar2p interacts with Kar8/Jem1p for nuclear fusion. Electron microscopy analysis of kar8 mutant zygotes re vealed a nuclear fusion defect different from kar2, kar5, and kar7/sec71 mu tants. Analysis of double mutants suggested that Kar5p acts before Kar8/Jem 1p. We propose the existence of a nuclear envelope fusion chaperone complex in which Kar2p, Kar5p, and Kar8/Jem1p are key components and Sec71p and Se c72p play auxiliary roles.