Growth rate influences MF alpha 1 promoter activity in MAT alpha Saccharomyces cerevisiae.

Abstract

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.