Aflatoxin conducive and non-conducive growth conditions reveal new gene associations with aflatoxin production

Research on aflatoxin (AF) production has traditionally focused on defining the AF biosynthetic pathway with the goal of identifying potential targets for intervention. To understand the effect of nitrogen source, carbon source, temperature, and pH on the regulation of AF biosynthesis, a targeted cDNA microarray consisting of genes associated with AF production over time was employed. Expression profiles for genes involved in AF biosynthesis grouped into five clades. A putative regulon was identified consisting of 20 genes that were induced in the conducive nitrogen and pH treatments and the non-conducive carbon and temperature treatments, as well as four other putative regulons corresponding to each of the four variables studied. Seventeen genes exhibited consistent induction/repression profiles across all the experiments. One of these genes was consistently downregulated with AF production. Overexpression of this gene resulted in repression of AF biosynthesis. The cellular function of this gene is currently unresolved.     

Comparative genomic analysis of dha regulon and related genes for anaerobic glycerol metabolism in bacteria

The dihydroxyacetone (dha) regulon of bacteria encodes genes for the anaerobic metabolism of glycerol. In this work, genomic data are used to analyze and compare the dha regulon and related genes in different organisms in silico with respect to gene organization, sequence similarity, and possible functions. Database searches showed that among the organisms, the genomes of which have been sequenced so far, only two, i.e., Klebsiella pneumoniae MGH 78578 and Clostridium perfringens contain a complete dha regulon bearing all known enzymes. The components and their organization in the dha regulon of these two organisms differ considerably from each other and also from the previously partially sequenced dha regulons in Citrobacter freundii, Clostridium pasteurianum, and Clostridium butyricum. Unlike all of the other organisms, genes for the oxidative and reductive pathways of anaerobic glycerol metabolism in C. perfringens are located in two separate organization units on the chromosome. Comparisons of deduced protein sequences of genes with similar functions showed that the dha regulon components in K. pneumoniae and C. freundii have high similarities (80-95%) but lower similarities to those of the Clostridium species (30-80%). Interestingly, the protein sequence similarities among the dha genes of the Clostridium species are in many cases even lower than those between the Clostridium species and K. pneumoniae or C. freundii, suggesting two different types of dha regulon in the Clostridium species studied. The in silico reconstruction and comparison of dha regulons revealed several new genes in the microorganisms studied. In particular, a novel dha kinase that is phosphoenolpyruvate-dependent is identified and experimentally confirmed for K. pneumoniae in addition to the known ATP-dependent dha kinase. This finding gives new insights into the regulation of glycerol metabolism in K. pneumoniae and explains some hitherto not well understood experimental observations.     

Genome-wide transcriptional analysis of the phosphate starvation stimulon of Bacillus subtilis

Bacillus subtilis responds to phosphate starvation stress by inducing the PhoP and SigB regulons. While the PhoP regulon provides a specific response to phosphate starvation stress, maximizing the acquisition of phosphate (P(i)) from the environment and reducing the cellular requirement for this essential nutrient, the SigB regulon provides nonspecific resistance to stress by protecting essential cellular components, such as DNA and membranes. We have characterized the phosphate starvation stress response of B. subtilis at a genome-wide level using DNA macroarrays. A combination of outlier and cluster analyses identified putative new members of the PhoP regulon, namely, yfkN (2',3' cyclic nucleotide 2'-phosphodiesterase), yurI (RNase), yjdB (unknown), and vpr (extracellular serine protease). YurI is thought to be responsible for the nonspecific degradation of RNA, while the activity of YfkN on various nucleotide phosphates suggests that it could act on substrates liberated by YurI, which produces 3' or 5' phosphoribonucleotides. The putative new PhoP regulon members are either known or predicted to be secreted and are likely to be important for the recovery of inorganic phosphate from a variety of organic sources of phosphate in the environment.     

The phosphate-starvation response in Vibrio cholerae O1 and phoB mutant under proteomic analysis: disclosing functions involved in adaptation, survival and virulence

A proteomic analysis of a wild-type and of a phoB mutant showed that Vibrio cholerae expresses genes of two major regulons in response to phosphate starvation. The Pho regulon, expressed by the wild-type, allowed the cells to adapt to the new environment. Induction of the general stress regulon was mainly observed in the phoB mutant as a strategy to resist stress and survive. Some functions of the adaptative and survival responses play roles in the pathogenicity of the bacteria. Among the members of the Pho regulon, we found a porin described as an important factor for the intestinal colonisation. Other functions not obviously related to phosphate metabolism, expressed preferentially by the wild-type cells, have also been implicated in virulence. These findings might explain the lack of virulence of the phoB mutant. The Pho regulon picture of V. cholerae, however, will not be complete until minor members and membrane proteins are identified. Among the phosphate-starvation induced genes we have found 13 hypothetical ones and for some of them functions have been assigned. The majority of the genes identified here have not been described before, thus they could be used to expand the proteomic reference map of V. cholerae El Tor.     

Genetic variability of the emm-related genes of the large vir regulon of group A streptococci: potential intra- and intergenomic recombination events

One of the most prevalent genetic lineages of group A streptococci (GAS) harbors a genomic locus termed the large vir regulon, which contains an emm gene encoding the antiphagocytic M protein, and structurally related fcrA and enn (emm-related) genes encoding immunoglobulin-binding proteins. In the present study more than 100 large vir regulons from 42 different GAS serotypes were analyzed by PCR and partial DNA sequencing. On comparing these data to published sequences, sites of mutational and putative recombinational events were identified and ordered with respect to their intra/intergenic or intra/intergenomic nature. The emm-related genes were found to display small intragenic deletions or insertions, were completely deleted from, or newly inserted into the genome, or were fused to adjacent genes. Intergenomic exchanges of complete emm-related genes, or segments thereof, between different vir regulons were detected. Most of these processes seem to involve short flanking direct repeats. Occasionally, the structural changes could be correlated with changes in the functions of the encoded proteins.     

Genetic variability of the emm-related genes of the large vir regulon of group A streptococci: potential intra- and intergenomic recombination events

One of the most prevalent genetic lineages of group A streptococci (GAS) harbors a genomic locus termed the large vir regulon, which contains an emm gene encoding the antiphagocytic M protein, and structurally related fcrA and enn (emm-related) genes encoding immunoglobulin-binding proteins. In the present study more than 100 large vir regulons from 42 different GAS serotypes were analyzed by PCR and partial DNA sequencing. On comparing these data to published sequences, sites of mutational and putative recombinational events were identified and ordered with respect to their intra/intergenic or intra/intergenomic nature. The emm-related genes were found to display small intragenic deletions or insertions, were completely deleted from, or newly inserted into the genome, or were fused to adjacent genes. Intergenomic exchanges of complete emm-related genes, or segments thereof, between different vir regulons were detected. Most of these processes seem to involve short flanking direct repeats. Occasionally, the structural changes could be correlated with changes in the functions of the encoded proteins.