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.     

Azo dye decolorization with a mutant Escherichia coli strain

A recombinant Escherichia coli strain (E. coli NO3) containing genomic DNA fragments from azo-reducing wild-type Pseudomonas luteola strain decolorized a reactive azo dye (C.I. Reactive Red 22) at approx. 17 mg dye h^–1 g cell. The ability to decolorize the azo dye probably did not originate from the plasmid DNA. Acclimation in azo-dye-containing media gave a nearly 10% increase in the decolorization rate of E. coli NO3. Growth with 1.25 g glucose l^–1 completely stopped the decolorization activity. When the decolorization metabolites from E. coli NO3 were analyzed by HPLC and MS, the results suggested that decolorization of the azo dye may be due to cleavage of the azo bond.     

Cooperative interactions of protein kinase C and cAMP-dependent protein kinase systems in human promyelocytic leukemia HL60 cells

Interactions of protein kinase C (PKC) and cAMP-dependent protein kinase (PKA) systems were investigated in HL60 cells. It was found that the differentiating effects of 12-O-tetradecanoylphorbol-13-acetate (TPA) were potentiated by dibutyryl cAMP (dbcAMP) or prostaglandin E2 (PGE2). In addition, dbcAMP or PGE2 inhibited TPA-induced binding of PKC to plasma membrane, leading to decreased protein phosphorylation, and promoted subsequent redistribution of enzyme to the nuclear membrane region. The findings are consistent with the hypothesis that PKC and PKA systems regulate cooperatively the phenotypical differentiation of leukemic cells.