engleski

SCW and CCW genes inactivation influence cell viability

Saccharomyces cerevisiae cell wall contains more than 20 different mannoproteins. Some of them are attached to wall structural polysaccharides noncovalently (Scw - soluble cell wall proteins) and can be extracted by hot SDS. The second group comprises proteins covalently (Ccw - covalently linked cell wall proteins) bound to glucan and they are usually extracted by glucanases but a group of them (so called Pir-proteins with internal repeats) can also be released from glucan by a treatment with NaOH. The physiolological role of most mannoproteins is still unknown. In order to find out possible cellular roles of these proteins, mutants were constructed by disruption of SCW4, SCW10, SCW11 and SCW8/BGL2 , as well as all four known PIR genes (CCW5/PIR4, CCW6/PIR1, CCW7/PIR2/HSP150 and CCW8/PIR3) and phenotype of different single and multiple mutants is observed at the level of individual cells. Viability of mutants in exponential and stationary growth phase was monitored by methylene blue staining. Both, exponential and stationary phase scw4scw10 and scw4scw10bgl2 showed increased fraction of dead cells in the culture. Disruption of scw11 in scw4scw10 and in scw4scw10bgl2 background suppressed the observed phenotype. These results suggest some synergistic and/or antagonistic behavior of proteins that are involved in building and maintenance of the cell wall. Mutants lacking major covalently bound proteins (ccw mutants) also had increased mortalities. These mutants showed significantly increased mortality in stationary cells suggesting that their physiological role is primarily expressed in that phase of growth. Microscopic investigation of ccw multiple mutants showed an increased fraction of cells with more than one bud and in most cases daughter cells still attached to their mothers stained with methylene blue. All cultivated yeast strains showed lower mortality when cultivated in the presence of sorbitol as osmotic stabilizer, indicating that cells died due to decreased osmotic stability.

cell wall; mannoproteins

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