Induction of a mitosis delay and cell lysis by high-level secretion of mouse alpha-amylase from Saccharomyces cerevisiae.
Some foreign proteins are produced in yeast in a cell cycle-dependent manner, but the cause of the cell cycle dependency is unknown. In this study, we found that Saccharomyces cerevisiae cells secreting high levels of mouse alpha-amylase have elongated buds and are delayed in cell cycle completion in mitosis. The delayed cell mitosis suggests that critical events during exit from mitosis might be disturbed. We found that the activities of PP2A (protein phosphatase 2A) and MPF (maturation-promoting factor) were reduced in alpha-amylase-oversecreting cells and that these cells showed a reduced level of assembly checkpoint protein Cdc55, compared to the accumulation in wild-type cells. MPF inactivation is due to inhibitory phosphorylation on Cdc28, as a cdc28 mutant which lacks an inhibitory phosphorylation site on Cdc28 prevents MPF inactivation and prevents the defective bud morphology induced by overproduction of alpha-amylase. Our data also suggest that high levels of alpha-amylase may downregulate PPH22, leading to cell lysis. In conclusion, overproduction of heterologous alpha-amylase in S. cerevisiae results in a negative regulation of PP2A, which causes mitotic delay and leads to cell lysis.
Wang
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BD
Institute of Molecular Biology, Academia Sinica, Nankang, Taipei 115, Taiwan. bdwang@gate.sinica.edu.tw
Kuo
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TT
D000818:Animals; D017489:CDC28 Protein Kinase, S cerevisiae / Q000235:genetics / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000378:metabolism; D003470:Culture Media; D004271:DNA, Fungal / Q000032:analysis; D015966:Gene Expression Regulation, Fungal; D016200:Maturation-Promoting Factor / Q000378:metabolism; D000090204:Mesothelin; D051379:Mice; D008938:Mitosis / Q000502:physiology*; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D054648:Protein Phosphatase 2; D011994:Recombinant Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000502:physiology* / Q000648:ultrastructure; D029701:Saccharomyces cerevisiae Proteins; D000516:alpha-Amylases / Q000235:genetics / Q000378:metabolism*
C071246:CDC55 protein, S cerevisiae; D018797:Cell Cycle Proteins; D003470:Culture Media; D004271:DNA, Fungal; C000715528:Msln protein, mouse; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D017489:CDC28 Protein Kinase, S cerevisiae; D016200:Maturation-Promoting Factor; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2; D000516:alpha-Amylases; D000090204:Mesothelin