SELECTION OF AXIAL GROWTH SITES IN YEAST REQUIRES AXL2P, A NOVEL PLASMA-MEMBRANE GLYCOPROTEIN

Spa2p and Cdc10p both participate in bud site selection and cell morph ogenesis in yeast, and spa2 Delta cdc10-10 cells are inviable. To iden tify additional components important for these processes in yeast, a c olony-sectoring assay was used to isolate high-copy suppressors of the spa2 Delta cdc10-10 lethality. One such gene, AXL2, has been characte rized in detail. axl2 cells are defective in bud site selection in hap loid cells and bud in a bipolar fashion. Genetic analysis indicates th at AXL2 falls into the same epistasis group as BUD3. Axl2p is predicte d to be a type I transmembrane protein. Tunicamycin treatment experime nts, biochemical fractionation and extraction experiments, and protein ase K protection experiments collectively indicate that Axl2p is an in tegral membrane glycoprotein at the plasma membrane. Indirect immunofl uorescence experiments using either Axl2p tagged with three copies of a hemagglutinin epitope or high-copy AXL2 and anti-Axl2p antibodies re veal a unique localization pattern for Axl2p. The protein is present a s a patch at the incipient bud site and in emerging buds, and at the b ud periphery in small-budded cells. In cells containing medium-sized o r large buds, Axl2p is located as a ring at the neck. Thus, Axl2p is a novel membrane protein critical for selecting proper growth sites in yeast. We suggest that Axl2p acts as an anchor in the plasma membrane that helps direct new growth components and/or polarity establishment components to the cortical axial budding site.