Dataset Information

Catabolite repression of phosphoenolpyruvate carboxykinase by a zinc finger protein under biotin- and pyruvate carboxylase-deficient conditions in Pichia pastoris

ABSTRACT: We have identified a methanol- and biotin-starvation-inducible zinc finger protein named ROP [repressor of phosphoenolpyruvate carboxykinase (PEPCK)] in the methylotrophic yeast Pichia pastoris. When P.pastoris strain GS115 (wild-type, WT) is cultured in biotin-deficient, glucose ammonium (Bio-) medium, growth is suppressed due to the inhibition of anaplerotic synthesis of oxaloacetate, catalysed by the biotin-dependent enzyme pyruvate carboxylase (PC). Deletion of ROP results in a strain (∆ROP) that can grow under biotin-deficient conditions due to derepression of a biotin- and PC-independent pathway of anaplerotic synthesis of oxaloacetate. Northern analysis as well as microarray expression profiling of RNA isolated from WT and ∆ROP strains cultured in Bio(-) medium indicate that expression of the phosphoenolpyruvate carboxykinase gene (PEPCK) is induced in ∆ROP during biotin- or PC-deficiency even under glucose-abundant conditions. There is an excellent correlation between PEPCK expression and growth of ∆ROP in Bio(-) medium, suggesting that ROP-mediated regulation of PEPCK may have a crucial role in the biotin- and PC-independent growth of the ∆ROP strain. To our knowledge, ROP is the first example, of a zinc finger transcription factor involved in the catabolite repression of PEPCK in yeast cells cultured under biotin- or PC-deficient and glucose-abundant conditions.

ORGANISM(S): Komagataella pastoris

PROVIDER: GSE32948 | GEO | 2011/10/14

SECONDARY ACCESSION(S): PRJNA146463

REPOSITORIES: GEO

ACCESS DATA

Similar Datasets

Project description:DNA affinity chromatography with the promoter region of the Corynebacterium glutamicum pck gene, encoding phosphoenolpyruvate carboxykinase (PEPCk), led to the isolation of four transcriptional regulators, i.e., RamA, GntR1, GntR2 and IolR. Determination of the PEPCk activity of the deletion mutants M-NM-^TramA, M-NM-^TgntR1M-NM-^TgntR2, and M-NM-^TiolR indicated that RamA represses pck during growth on glucose about twofold, whereas GntR1, GntR2, and IolR activate pck expression about twofold, irrespective whether glucose or acetate served as carbon source. The DNA binding sites of the four regulators in the pck promoter region were identified and their positions correlated with the predicted functions as repressor or activators. The iolR gene is located upstream and in divergent orientation to a iol gene cluster, encoding proteins involved in myo-inositol uptake and degradation. Comparative DNA microarray analysis of the M-NM-^TiolR mutant and the parental wild-type revealed strongly elevated (>100-fold) mRNA levels of the iol genes in the mutant, indicating that the primary function of IolR is the repression of the iol genes. IolR binding sites were identified in the promoter regions of iolC, iolT and iolR itself, which presumably is subject to negative autoregulation. A consensus DNA-binding motif was identified (5M-bM-^@M-^Y-KGWCHTRACA-3M-bM-^@M-^Y) which corresponds well to those of other GntR-type regulators of the HutC family. Taken together, our results disclose a complex regulation of the pck gene in C. glutamicum and identify IolR as an efficient repressor of genes involved in myo-inositol catabolism of this organism. To identify genes that are potentially regulated by IolR, the transcriptome profile of the iolR deletion strain was compared to that of the WT using DNA microarray analysis. The strains were grown in CGXII minimal medium with 4% (wt/vol) glucose as sole carbon source and RNA was isolated from cells harvested in the early exponential growth phase (OD600 of about 5). The transcriptome comparison was performed in triplicate starting from independent cultures. The second replicate included a dye-swap.

Dataset Information

Catabolite repression of phosphoenolpyruvate carboxykinase by a zinc finger protein under biotin- and pyruvate carboxylase-deficient conditions in Pichia pastoris

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure