Accepted post-doctoral position at Texas A&M IBT after receiving PhD
Now is continuing postdoctoral training at Texas Children’s Hospital, Houston, TX
The susceptibility of most Bacillus anthracis strains to β-lactam antibiotics is intriguing considering that closely-related species, Bacillus cereus and Bacillus thuringiensis, typically produce β-lactamases and the B. anthracis genome harbors two β-lactamase genes, bla1 and bla2. I show that B. anthracis bla expression is affected by two genes, sigP and rsp, predicted to encode an extracytoplasmic function sigma factor and an antisigma factor, respectively. Deletion of the sigP/rsp locus abolished bla expression in a penicillin-resistant clinical isolate and had no effect on bla expression in a prototypical penicillin-susceptible strain. Complementation with sigP/rsp from the penicillin-resistant strain, but not the penicillin-susceptible strain, conferred β-lactamase activity upon both mutants. These results are attributed to a nucleotide deletion near the 5’ end of rsp in the penicillin-resistant strain that is predicted to result a non-functional protein. B. cereus and B. thuringiensis sigP and rsp homologues are required for inducible penicillin resistance in those species. Expression of the B. cereus or B. thuringiensis sigP and rsp genes in a B. anthracis sigP/rsp-null mutant confers resistance to β-lactam antibiotics, suggesting that while B. anthracis contains the genes necessary for sensing β-lactam antibiotics, the B. anthracis sigP/rsp gene products are insufficient for bla induction.
Because alternative sigma factors recognize unique promoter sequence, direct targets can be elucidated by comparing transcriptional profiling results with an in silico search using the sigma factor binding sequence. I identified potential σP -10 and -35 promoter elements upstream from bla1, bla2 and sigP and compared results obtained from searching the B. anthracis genome with the conserved sequences to transcriptional profiling results comparing B. anthracis 32 and an isogenic sigP/rsp-null strain. I determined that while the absence of the sigP gene significantly affects the transcript levels of 16 genes, only bla1, bla2 and sigP are directly regulated by σP. The genomes of B. cereus and B. thuringiensis strains were also analyzed for the potential σP binding elements. The sequence was located upstream from the sigP and bla genes, and previously unidentified genes predicted to encode a penicillin-binding protein (PBP) and a D-alanyl-D-alanine carboxypeptidase, indicating that the σP regulon in these species responds to cell-wall stress caused by β-lactam antibiotics.
β-lactam antibiotics prevent attachment of new peptidoglycan to the cell wall by blocking the active site of PBPs. A B. cereus and B. thuringiensis pbp-encoding gene located near bla1 contains a potential σP recognition sequence upstream from the annotated translational start. Deletion of this gene abolished β-lactam resistance in both strains. Mutations in the active site of the PBP were detrimental to β-lactam resistance in B. cereus, but not B. thuringiensis, indicating that the transpeptidase activity is only important in B. cereus. I also found that transcript levels of the PBP-encoding gene are not significantly affected by the presence of β-lactam antibiotic. Based on these data I hypothesize that the gene product acts a sensor of β-lactam antibiotic.
The molecular basis for β-lactamase gene expression in B. anthracis, B. cereus and B. thuringiensis