Studies on the post-transcriptional regulation of genes of glutamate utilization pathway in Pichia pastoris and a role for Retrograde regulation protein 1

Abstract

The methylotrophic yeast, Pichia pastoris can utilize compounds such as glucose, glycerol, methanol, acetate, amino acids, or oleic acid as the sole source of carbon. However, the regulation of synthesis of key enzymes of these metabolic pathways is not well understood. Unlike Saccharomyces cerevisiae, P. pastoris can utilize amino acids not only as a nitrogen source but also a carbon source. The prime focus of this work is to understand the regulatory mechanisms involved in the synthesis of two key enzymes of glutamate utilization pathway, glutamate dehydrogenase (GDH2) and phosphoenolpyruvate carboxykinase (PEPCK) in P. pastoris. We demonstrate that: i) P. pastoris basic, helix-loop-helix, leucine zipper protein known as PpRtg1p functions as a nuclear transcription factor in Saccharomyces cerevisiae and cytosolic post-transcriptional regulator in P. pastoris. ii) PpRtg1p regulates the translation of GDH2 and PEPCK mRNAs when glutamate is the sole source of carbon. iii) GDH2 and PEPCK regulate each other's protein levels via PpRtg1p-independent mechanism when glutamate is the sole source of carbon. iv) Translation of GDH2 and PEPCK mRNAs is induced by metabolites generated during prolonged carbon starvation.