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The conserved WhiA protein influences the fatty acid composition of the Bacillus subtilis cell membrane

ABSTRACT: The conserved WhiA protein family is present in most Gram-positive bacteria and plays a role in cell division. WhiA contains a DNA-binding motive and has been identified as a transcription factor in the actinomycetes. In Bacillus subtilis, a deletion of whiA influences cell division and chromosome segregation. However, WhiA does not seem to function as a transcription factor in this organism, and it is still unclear how WhiA influences these processes. The B. subtilis whiA gene is part of an operon and flanked by two metabolic genes, including yvcK required for growth under gluconeogenic conditions, and the gene encoding the catabolite repressor Crh. Therefore, B. subtilis WhiA might play a role in carbon metabolism which could indirectly affect cell division and chromosome maintenance. To study this, we followed a metabolomics approach and measured exo-metabolome flux during growth on different carbon sources, using nuclear magnetic resonance spectroscopy. The carbon utilization of wild type and whiA mutant cells was similar, however, the whiA mutant exhibited depleted pools of branched-chain fatty acid precursors and higher levels of acetate and 2-oxoglutarate. Transcriptome data could not link these effects to gene regulatory differences, but the reduction in branched-chain fatty acid precursors influenced the fatty acid composition of the cell membrane. In conclusion, WhiA does not participate in carbon catabolite regulation, but affects fatty acid composition of the membrane by an as yet unknown mechanism. We speculate that the effect on the membrane might be related to the pleiotropic phenotype of the B. subtilis whiA mutant.

ORGANISM(S): Bacillus subtilis

PROVIDER: GSE121479 | GEO | 2019/10/18

REPOSITORIES: GEO

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Project description:Fatty acid synthesis is closely linked to nutrient availability and cellular energetic status. The committed step in fatty acid synthesis is the acetyl CoA carboxylase. Eukaryotes have two genes encoding acetyl CoA carboxylases, one encoding a cytosolic enzyme and another coding for a mitochondrial enzyme. They catalyze the synthesis of malonyl CoA in the cytosol and the mitochondria, respectively. While cytosolic malonyl CoA is the precursor for fatty acid synthesis, mitochondrial malonyl CoA controls the transfer of fatty acyl group into the mitochondria by inhibiting carnitine/palmitoyl transferase activity and thus, regulates Î²-oxidation. In Saccharomyces cerevisiae, Î²-oxidation is restricted to the peroxisomes, raising the question of the function of the mitochondrial isoform (HFA1). In this study, we replaced the cytosolic Acc1 with Hfa1 expressed in the cytosol by removing the mitochondrial leader peptide, under control of the HFA1 promoter. We studied fatty acid synthesis and transcription profiles in this strain during starvation for carbon or nitrogen, using glucose or ethanol as the carbon source. Under all the conditions studied, the key sensor of energetic status, Snf1, was activated, indicating active inhibition of fatty acid synthesis. The pool size of fatty acids was smaller when Acc1 was replaced with truncated Hfa1 for fatty acid synthesis. Yet, the transcription profiles were similar in both the cases. These results point towards the conclusion that Hfa1 is either catalytically less efficient or it is more sensitive to inhibition by Snf1. Gene expression from a strain of Saccharomyces cerevisiae where cytosolic fatty acid synthesis occurs by mitochondrial acetyl CoA carboxylase (without its mitochondrial leader peptide) is compared with that in a reference strain while growing in chemostats on carbon or nitrogen starvation using glucose or ethanol as the carbon source. There are two strains (reference or mutant), two carbon sources (glucose or ethanol) and two limitations (carbon or nitrogen), resulting in 8 comparisons. Each array was performed in duplicate, resulting in 16 CEL files. Growth was limited by either carbon or nitrogen. When carbon was the limited nutrient, we tested growth on either glucose or ethanol (both using ammonium sulfate as the nitrogen source). When ammonium sulfate was limiting, we used either glucose or ethanol as the carbon source.

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The conserved WhiA protein influences the fatty acid composition of the Bacillus subtilis cell membrane

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