Growth rate influences MF alpha 1 promoter activity in MAT alpha Saccharomyces cerevisiae.
The signal sequences of the MF alpha 1 prepro alpha-factor gene are frequently used to direct secretion of heterologous proteins from Saccharomyces cerevisiae. They are often employed together with the MF alpha 1 promoter in secretion vectors, such that this promoter directs the transcription of many heterologous gene cassettes in yeast. Most of the existing literature indicates that the MF alpha 1 promoter is constitutive in MAT alpha cells, although some data suggests that it may be more active in respiratory or late logarithmic fermentative cultures. To identify whether there is a growth rate or medium control over MF alpha 1 promoter activity a strain was constructed with an integrated MF alpha 1 promoter-beta-galactosidase (lacZ) reporter gene fusion. Intracellular beta-galactosidase of this strain during batch culture on glucose, raffinose and acetate showed that MF alpha 1 promoter activity was higher during respiratory growth on acetate as compared to more rapid fermentative growth on glucose or raffinose, a result that might indicate this activity being inversely related to growth rate. Chemostat culture confirmed that growth rate does indeed influence MF alpha 1 promoter activity in glucose-grown cells, the activity of this promoter increasing 2- to 2.5-fold as dilution (growth) rates were reduced from maximal values to 0.2 h-1, but then decreasing with the further decreases in dilution rate needed for fully respiratory growth. Thus a promoter generally thought to be constitutive in MAT alpha cells is nevertheless subject to a complex growth rate control.
Kirk
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Department of Biochemistry and Molecular Biology, University College London, UK.
Piper
P W
PW
D001483:Base Sequence; D001709:Biotechnology; D003001:Cloning, Molecular; D003470:Culture Media; D004271:DNA, Fungal / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics*; D005800:Genes, Fungal; D017930:Genes, Reporter; D005947:Glucose / Q000378:metabolism; D007700:Kinetics; D007763:Lac Operon; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011498:Protein Precursors / Q000235:genetics*; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins
D003470:Culture Media; D004271:DNA, Fungal; D005656:Fungal Proteins; C048632:MF(ALPHA)1 protein, S cerevisiae; D011498:Protein Precursors; D029701:Saccharomyces cerevisiae Proteins; D005947:Glucose