Cloning and characterization of GPD2, a second gene encoding sn‐glycerol 3‐phosphate dehydrogenase (NAD⁺) in Saccharomyces cerevisiae, and its comparison with GPD1

Abstract

We have cloned and characterized a homologue of the previously isolated GPD1 gene, encoding sn-glycerol 3-phosphate dehydrogenase (NAD+) in Saccharomyces cerevisiae. This second gene, called GPD2, encodes a protein of 384 amino acids that shares 69% sequence identity with GPD1. Like GPD1 it has an amino-terminal extension of unknown function. GPD2 is located on chromosome VII and cross-hybridizes with GPD1 at chromosome IV as well as with an unknown homologue at chromosome XV. Disruption of the GPD2 gene did not reveal any observable phenotypic effects, whereas overexpression resulted in a slight, but significant, increase of GPD enzyme activity in wild-type cells. Analysis of gene transcription by a CAT-reporter gene fused to the GPD promoters revealed decreased transcriptional activity of the GPD2 promoter in cells grown on nonfermentable as opposed to fermentable carbon sources, and no induction in cells exposed to high osmolarity or heat shock. Similar analysis of GPD1 demonstrated an 8-17-fold higher basal level of transcription compared to GPD2. Furthermore, such analysis revealed that the GPD1 promoter was induced by increased osmolarity essentially independent of the type of stress solute used, the level of GPD1 transcription being increased about sevenfold in cells growing at 1.4 M NaCl.

Global changes in protein synthesis during adaptation of the yeast Saccharomyces cerevisiae to 0.7 M NaCl

“…The great number of synthetic changes also implies that the salt-implemented osmotic dehydration instigated impact on many cellular activities. This assumption is strengthened by the fact that genes involved in diverse cellular functions have been reported to be regulated by the osmotic properties of the medium: HAL1, which is involved in Na ϩ -K ϩ selectivity of the cell (20); ENA1, encoding a P-type ATPase participating in the efflux of sodium ions (15); GPD1, which is involved in glycerol production (1,14,41); and TPS2, which encodes one of the subunits of the trehalose synthase complex (20a). Both the maintenance of a proper intracellular Na ϩ balance and the accumulation of the compatible solute glycerol are key features of the osmoregulatory response in yeast cells (7).…”

“…Salinity-induced expression of heat shock genes may indicate heat shock factor-mediated osmotic signalling; however, this factor is apparently not involved in GPD1 regulation (14,40).…”

“…A key enzyme in this production, at least for the yeast Saccharomyces cerevisiae, is glycerol-3-phosphate dehydrogenase (GPD), and cells have been shown to respond to the osmotic stress by an increased specific activity of this enzyme (6,11). This activity increase reflects an enhanced number of protein molecules in the cell (3), and GPD1 expression is osmotically regulated at the level of DNA transcription (1,2,14,41).…”

Role of 26S proteasome and HRD genes in the degradation of 3-hydroxy-3-methylglutaryl-CoA reductase, an integral endoplasmic reticulum membrane protein.

“…The term GPD has been used on numerous occasions by us and others (e.g., Schena et al, 1991) to indicate the GAPDH promoter as cloned and characterized by Bitter and Egan (1984). However, there are two other yeast genes designated GPD1 and GPD2 that do not encode GAPDH, but rather sn-glycerol-3-phosphate dehydrogenase (NAD+), which are highly regulated enzymes subject to numerous controls (Eriksson et al, 1995). Therefore, we now refer to the strong constitutive promoters used herein and in our previous studies as pGAPDH to distinguish them from the unrelated GPD genes.…”

Yeast: World’s Finest Chef

“…Yet, one must keep in mind that with fermentation is associated culture growth and, biomass composition is more oxidized than glucose, consequently an excess of reducing equivalents may be attained. The way yeast circumvent this problem, under anaerobic conditions, consists on the production of glycerol by reduction of the glycolytic intermediate dihydroxyacetone phosphate to glycerol 3-phosphate catalysed by NAD + -dependent glycerol 3-phosphate dehydrogenase (encoded by the two isogenes GPD1 and GPD2), and its subsequent dephosphorylation due to the action of glycerol 3-phosphatase (encoded by GPP1 and GPP2) [14][15][16].…”