BED1, a gene encoding a galactosyltransferase homologue, is required for polarized growth and efficient bud emergence in Saccharomyces cerevisiae.

Abstract

Abstract. The ellipsoidal shape of the yeast Saccharomyces cerevisiae is the result of successive isotropic/apical growth switches that are regulated in a cell cycledependent manner. It is thought that growth polarity is governed by the remodeling of the actin cytoskeleton that is itself under the control of the cell cycle machinery. The cell cycle and the morphogenesis cycle are tightly coupled and it has been recently suggested that a morphogenesis/polarity checkpoint control monitors bud emergence in order to maintain the coupling of these two events (Lew, D. J., and S. I. Reed. 1995. J. Cell BioL 129:739-749). During a screen based on the inability of cells impaired in the budding process to survive when the morphogenesis checkpoint control is abolished, we identified and characterized BED1, a new gene that is required for efficient budding. Cells carrying a disrupted allele of BED1 no longer have the wild-type ellipsoidal shape characteristic of S. cerevisiae, are larger than wild-type cells, are deficient in bud emergence, and depend upon an intact morphogenesis checkpoint control to survive. These cells show defects in polarized growth despite the fact that the actin cytoskeleton appears normal. Our results suggest that Bedl is a type II membrane protein localized in the endoplasmic reticulum. BED1 is significantly homologous to gma12 ÷, a S. pombe gene coding for an et-l,2-galactosyltransferase, suggesting that glycosylation of specific proteins or lipids could be important for signaling in the switch to polarized growth and in bud emergence.T HE ellipsoidal shape of the yeast Saccharomyces cerevisiae reflects cell cycle-regulated polarized growth. At specific times during the cell cycle, cell growth is either isotropic or polarized toward the bud (for review see Lew and Reed, 1995b). A correlation between local deposition of new cell wall components and actin localization has been established (Adams and Pringle, 1984;Kilmartin and Adams, 1984), leading to the proposal that actin directs secretory vesicles to specific regions of the plasma membrane to allow localized cell surface growth during bud initiation and bud growth. During most of the G1 phase, growth is isotropic and cortical actin patches are delocalized throughout the cell. The attainment of a critical cell size and concomitant execution of START lead to the formation of an actin ring at the pre-bud site and the orientation of actin filaments toward this site. Subsequent to START, growth is almost completely restricted to the emerging bud. During bud growth, cortical actin patches are localized to the bud. Initially, bud growth occurs primarily at the distal tip. At some point, though, there is a switch to isotropic growth first in the bud, then also tranAddress all correspondence to S. I. Reed, Department of Molecular Biology, MB7, The Scripps Research Institute, 10666 North Torrey Pines Road, La Jolla, California 92037. Tel.: (619) 554-6188. siently in the mother cell at mitosis. At cytokinesis, the actin cytoskeleton is reorganized and...

Morphogenesis and the Cell Cycle

“…This includes a mutation (bed1 or mnn10) that delays budding by affecting protein glycosylation without overtly perturbing actin organization (Mondesert and Reed 1996;Theesfeld et al 2003). Thus, it seems possible that the checkpoint monitors bud emergence and delays Clb1,2p-Cdc28p activation when there is no bud.…”

The Checkpoint Kinase Hsl1p Is Activated by Elm1p-dependent Phosphorylation

“…The following disruptions and alleles are described in the cited publications: cla4::TRP1 (Benton et al, 1997); gin4-⌬9 (Longtine et al, 1998a); hsl1::URA3 (Ma et al, 1996); mih1::LEU2 (Russell et al, 1989); bed1::URA3 (Mondesert and Reed, 1996); nap1::kan r , HSL7-HA:kan r , and cdc12-6::LEU2 (Longtine et al, 2000); HSL1-13myc::URA3 (McMillan et al, 1999); hsl1 K110R -13myc (Theesfeld et al, 2003); and CDC3-GFP (Caviston et al, 2003). elm1::LEU2 and mih1::TRP1 disruptions were generated by the one-step PCR-based method (Baudin et al, 1993) using pRS305 and pRS304 (Sikorski and Hieter, 1989) as template.…”

Impaired Manganese Metabolism Causes Mitotic Misregulation

“…Moreover, cells lacking Pmr1 show genetic interactions with the cell polarity GTPase, Cdc42 (68). Several reports have shown that mannosyltransferase mutants affected in N-glycosylation are defective in polarized growth and become dependent on an intact mitotic checkpoint for survival, which provides time to produce a bud (56,57). Overriding the morphogenesis checkpoint by Clb2 overexpression in these mutants resembles the multinucleation phenotype of pmr1⌬ swe1⌬ mutants (56,57).…”

“…Several reports have shown that mannosyltransferase mutants affected in N-glycosylation are defective in polarized growth and become dependent on an intact mitotic checkpoint for survival, which provides time to produce a bud (56,57). Overriding the morphogenesis checkpoint by Clb2 overexpression in these mutants resembles the multinucleation phenotype of pmr1⌬ swe1⌬ mutants (56,57). The fact that some components of the cis-Golgi mannosyltransferase complex are Mn 2ϩ -dependent enzymes suggests that the function of these enzymes is impaired if the Golgi is not supplied with Mn 2ϩ (53)(54)(55).…”

“…Other studies have shown that mannosyltransferase mutants affected in protein glycosylation display a pronounced delay in bud formation (56,57). Moreover, SWE1 mRNA levels are elevated in cells that cannot make a bud (58).…”