Loss of catabolite repression function of HPr, the phosphocarrier protein of the bacterial phosphotransferase system, affects expression of the cry4A toxin gene in Bacillus thuringiensis subsp. israelensis.
HPr, the phosphocarrier protein of the bacterial phosphotransferase system, mediates catabolite repression of a number of operons in gram-positive bacteria. In order to participate in the regulatory process, HPr is activated by phosphorylation of a conserved serine-46 residue. To study the potential role of HPr in the regulation of Cry4A protoxin synthesis in Bacillus thuringiensis subsp. israelensis, we produced a catabolite repression-negative mutant by replacing the wild-type copy of the ptsH gene with a mutated copy in which the conserved serine residue of HPr was replaced with an alanine. HPr isolated from the mutant strain was not phosphorylated at Ser-45 by HPr kinase, but phosphorylation at His-14 was found to occur normally. The enzyme I and HPr kinase activities of the mutant were not affected. Analysis of the B. thuringiensis subsp. israelensis mutant harboring ptsH-S45A in the chromosome showed that cry4A expression was derepressed from the inhibitory effect of glucose. The mutant strain produced both cry4A and sigma(35) gene transcripts 4 h ahead of the parent strain, but there was no effect on sigma(28) synthesis. In wild-type B. thuringiensis subsp. israelensis cells, cry4A mRNA was observed from 12 h onwards, while in the mutant it appeared at 8 h and was produced for a longer period. The total amount of cry4A transcripts produced by the mutant was higher than by the parent strain. There was a 60 to 70% reduction in the sporulation efficiency of the mutant B. thuringiensis subsp. israelensis strain compared to the wild-type strain.
Khan
Sharik R
SR
Centre For Biotechnology, Jawaharlal Nehru University. International Centre For Genetic Engineering and Biotechnology, New Delhi, India.
Banerjee-Bhatnagar
Nirupama
N
D001413:Bacillus thuringiensis / Q000235:genetics / Q000254:growth & development / Q000502:physiology*; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001427:Bacterial Toxins; D004731:Endotoxins / Q000235:genetics / Q000378:metabolism*; D004794:Enzyme Repression; D015964:Gene Expression Regulation, Bacterial; D005947:Glucose / Q000378:metabolism; D006460:Hemolysin Proteins; D007700:Kinetics; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D010770:Phosphotransferases / Q000235:genetics / Q000378:metabolism; D017354:Point Mutation; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism; D013171:Spores, Bacterial / Q000502:physiology; D014158:Transcription, Genetic
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004731:Endotoxins; C064971:FliA protein, Bacteria; D006460:Hemolysin Proteins; D012808:Sigma Factor; C030956:insecticidal crystal protein, Bacillus Thuringiensis; D010770:Phosphotransferases; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System; C030182:phosphocarrier protein HPr; D005947:Glucose