Prm3p is a pheromone-induced peripheral nuclear envelope protein required for yeast nuclear fusion.
Nuclear membrane fusion is the last step in the mating pathway of the yeast Saccharomyces cerevisiae. We adapted a bioinformatics approach to identify putative pheromone-induced membrane proteins potentially required for nuclear membrane fusion. One protein, Prm3p, was found to be required for nuclear membrane fusion; disruption of PRM3 caused a strong bilateral defect, in which nuclear congression was completed but fusion did not occur. Prm3p was localized to the nuclear envelope in pheromone-responding cells, with significant colocalization with the spindle pole body in zygotes. A previous report, using a truncated protein, claimed that Prm3p is localized to the inner nuclear envelope. Based on biochemistry, immunoelectron microscopy and live cell microscopy, we find that functional Prm3p is a peripheral membrane protein exposed on the cytoplasmic face of the outer nuclear envelope. In support of this, mutations in a putative nuclear localization sequence had no effect on full-length protein function or localization. In contrast, point mutations and deletions in the highly conserved hydrophobic carboxy-terminal domain disrupted both protein function and localization. Genetic analysis, colocalization, and biochemical experiments indicate that Prm3p interacts directly with Kar5p, suggesting that nuclear membrane fusion is mediated by a protein complex.
Shen
Shu
S
Department of Molecular Biology, Princeton University, Princeton, NJ 08544-1014, USA.
Tobery
Cynthia E
CE
Rose
Mark D
MD
D000595:Amino Acid Sequence; D017124:Conserved Sequence; D005800:Genes, Fungal; D057927:Hydrophobic and Hydrophilic Interactions; D008561:Membrane Fusion / Q000187:drug effects*; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009685:Nuclear Envelope / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D010675:Pheromones / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects; D055550:Protein Stability / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*; D008941:Spindle Apparatus / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure
C109412:KAR5 protein, S cerevisiae; D008565:Membrane Proteins; D009687:Nuclear Proteins; D010675:Pheromones; C542168:Prm3 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins