Transcriptional regulation of desaturase genes in Pichia pastoris GS115.

ABSTRACT

Here we investigated the regulation of Pichia pastoris desaturase genes by low temperature and exogenous fatty acids in the late-exponential phase at the transcriptional level. Time-course studies of gene expression showed that mRNA levels of four desaturase genes were rapidly and transiently enhanced by low temperature and suppressed by exogenous oleic acid. Stearic acid showed no obvious repression of mRNA levels of Fad12 and Fad15 and a slight increase in Fad9A and Fad9B mRNA levels. Using a promoter-reporter gene construct, we demonstrated that the pFAD15 promoter activity was induced by low temperature in a time-dependent manner and reduced in a dose- and time-dependent manner by unsaturated fatty acids. Also, there was no absolute correlation between mRNA abundance and production of corresponding fatty acids. Disruption of Spt23 resulted in a decrease in transcript levels of Fad9A and Fad9B, but had little effect on the other desaturase genes. Consistent with these observations, a decrease in the relative amount of oleic acid (OLA) and an increase in the relative content of linoleic acid and ALA with different degrees were clearly observed in the stationary phase cells of ΔSpt23 mutant. Further analysis showed that the effect of low-temperature activation and OLA inhibition on expression of Fad9A and Fad9B seemed to disappear after disruption of the Spt23 gene, which indicated that Spt23p is essential for the expression of two Δ9-desaturase genes internally and probably involved in the regulation of Δ9-desaturase genes transcription in response to external stimuli, and thereby plays a role in the synthesis of OLA.