Role of Cell Cycle-regulated Expression in the Localized Incorporation of Cell Wall Proteins in Yeast

Abstract

The yeast cell wall is an essential organelle that protects the cell from mechanical damage and antimicrobial peptides, participates in cell recognition and adhesion, and is important for the generation and maintenance of normal cell shape. We studied the localization of three covalently bound cell wall proteins in Saccharomyces cerevisiae. Tip1p was found only in mother cells, whereas Cwp2p was incorporated in small-to-medium-sized buds. When the promoter regions of TIP1 and CWP2 (responsible for transcription in early G 1 and S/G 2 phases, respectively) were exchanged, the localization patterns of Tip1p and Cwp2p were reversed, indicating that the localization of cell wall proteins can be completely determined by the timing of transcription during the cell cycle. The third protein, Cwp1p, was incorporated into the birth scar, where it remained for several generations. However, we could not detect any role of Cwp1p in strengthening the birth scar wall or any functional interaction with the proteins that mark the birth scar pole as a potential future budding site. Promoter-exchange experiments showed that expression in S/G 2 phase is necessary but not sufficient for the normal localization of Cwp1p. Studies of mutants in which septum formation is perturbed indicate that the normal asymmetric localization of Cwp1p also depends on the normal timing of septum formation, composition of the septum, or both. INTRODUCTIONThe establishment and maintenance of asymmetry are of vital importance for the growth, differentiation, and morphogenesis of cells and organisms. For example, the yeast Saccharomyces cerevisiae grows asymmetrically, producing a bud that becomes the daughter cell (reviewed by Lew and Reed, 1995; Pruyne and Bretscher 2000a,b). In the G 1 phase of the cell cycle, a bud site is selected in a mating type-dependent manner. After the cell cycle-commitment point START, proteins required for bud initiation are recruited to this site, and a bud is formed in a process that depends largely on the delivery of vesicles containing new cell surface material by the polarized actin cytoskeleton. Growth occurs exclusively in the bud, first preferentially at the tip and later isotropically, until the time of cytokinesis. The actin cytoskeleton and cell surface growth are then redirected toward the mother-bud neck, where a chitinous primary septum is formed, followed by the deposition of secondary septa on both sides of the primary septum. Cells then separate by partial digestion of the primary septum, and a new cell cycle is initiated.The cell wall of yeast is essential for growth and is involved in the establishment and maintenance of asymmetry, as illustrated by the loss of both buds and polarized localization of cytoskeletal proteins when the cell wall is removed (Reck-Peterson et al., 1999). The cell wall is comprised of glucans, mannoproteins, and a small amount of chitin (reviewed by Orlean, 1997;Smits et al., 2001;Klis et al., 2006). The mannoproteins can be divided into four classes: 1) noncovalently linked prote...

Architecture and Biosynthesis of the Saccharomyces cerevisiae Cell Wall

“…Localization of these proteins correlates with their expression. Tip1 is expressed in G 1 and found in mother cells only, whereas Cwp1, Cwp2, and Tir1 are expressed during the S-to-G 2 transition, Cwp2 being found in small-to-medium-sized buds Smits et al 2006). Localization of Cwp2 and Tip1 is determined by the timing of their expression in the cell cycle (Smits et al 2006;File S9).…”

“…Tip1 is expressed in G 1 and found in mother cells only, whereas Cwp1, Cwp2, and Tir1 are expressed during the S-to-G 2 transition, Cwp2 being found in small-to-medium-sized buds Smits et al 2006). Localization of Cwp2 and Tip1 is determined by the timing of their expression in the cell cycle (Smits et al 2006;File S9). Tip1 and Tip2 are also heatand cold-shock-inducible, and Tir1 and Tir4 are induced by cold shock (Kowalski et al 1995;Abramova et al 2001).…”

“…A swap of the promoters of CWP2 and TIP1 caused these genes' products to exchange their cellular location, indicating that the localization of Cwp2 and Tip1, and perhaps that of other CWPs, is influenced by the timing of their expression in the cell cycle (Smits et al 2006). Cwp1, however, is localized to the birth scar in a manner that depends on normal septum formation, but, because neither Tip1 nor Cwp2 is targeted to the birth scar when expressed behind CWP1's promoter, additional CWP1 sequences are required for Cwp1 localization (Smits et al 2006).…”

Mitotic Exit and Separation of Mother and Daughter Cells

“…Similarly, Cwp1 is also trafficked to the site depending on when it is produced (Smits et al 2006). These proteins help cross-link the outer layers of the chitinous primary septum to the adjacent glucanrich secondary septum, ensuring that the multilayered structure does not delaminate.…”

Deregulation of DSE1 Gene Expression Results in Aberrant Budding within the Birth Scar and Cell Wall Integrity Pathway Activation in Saccharomyces cerevisiae

“…The elucidation of Dse1 links to potential interacting proteins and to other proteins localized to similar areas of daughter cells, e.g., Cwp1 protein that is specifically targeted to the birth scar area (25), is an important task to be undertaken in the near future. It may help to uncover the precise function of the Dse1 protein and its role in cell morphogenesis.…”