DEVELOPMENTAL GENETICS 12:281-292 (1991) Cellular Morphogenesis in the Saccharomyces cereuisiae Cell Cycle: Localization of the CDCII Gene Product and the Timing of Events at the Budding Site

The Saccharornyces cerevisiae CDC3, CDClO, CDCl I , and CDC12 genes encode a family of homologous proteins that are not closely related to other known proteins [Haarer BK, Ketcham SR, Ford SK, Ashcroft DJ, and Pringle JR (submitted)]. Temperature-sensitive mutants defective in any of these four genes display essentially identical pleiotropic phenotypes that include abnormal cell-wall deposition and bud growth, an inability to complete cytokinesis, and a failure to form the ring of 10 nm filaments that normally lies directly subjacent to the plasma membrane in the neck region of budding cells. We showed previously that the CDC3 and CDC72 gene products localize to the region of the mother-bud neck and are probably constituents of the ring of 10 nm filaments. We now report the generation of polyclonal antibodies specific for the CDCl1 product (Cdcl l p ) and the use of these antibodies in immunofluorescence experiments with wild-type and mutant cells. The results suggest that Cdcl 1 p is also a constituent of the filament ring, and thus support the hypothesis that the S. cerevisiae 10 nm filaments represent a novel type of eukaryotic cytoskeletal element. Cdcl 1 p and actin both localize to the budding site well in advance of bud emergence and at approximately the same time, and both proteins also remain localized at the old budding site for some time after cytokinesis. Cdcl 1 p also localizes to regions of cell-wall reorganization in mating cells and in cells responding to purified mating pheromone. Surprisingly, most preparations of affinity purified Cdcl 1 p-specific antibodies also stained the nuclear and cytoplasmic microtubules. Although this staining probably reflects the existence of an epitope shared by Cdcl 1 p and some microtubuleassociated protein, the possibility that a fraction of the Cdcl 1 p is associated with the microtubules could not be eliminated.