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Cloning and expression of a novel toxin gene from Bacillus thuringiensis subsp. jegathesan encoding a highly mosquitocidal protein.

ABSTRACT: A gene, designated cry11B, encoding a 81,293-Da crystal protein of Bacillus thuringiensis subsp. jegathesan was cloned by using a gene-specific oligonucleotide probe. The sequence of the Cry11B protein, as deduced from the sequence of the cry11B gene, contains large regions of similarity with the Cry11A toxin (previously CryIVD) from B. thuringiensis subsp. israelensis. The Cry11B protein was immunologically related to both Cry11A and Cry4A proteins. The cry11B gene was expressed in a nontoxic strain of B. thuringiensis, in which Cry11B was produced in large amounts during sporulation and accumulated as inclusions. Purified Cry11B inclusions were highly toxic for mosquito larvae of the species Aedes aegypti, Culex pipiens, and Anopheles stephensi. The activity of Cry11B toxin was higher than that of Cry11A and similar to that of the native crystals from B. thuringiensis subsp. jegathesan, which contain at least seven polypeptides.

SUBMITTER: Delecluse A

PROVIDER: S-EPMC167734 | biostudies-other | 1995 Dec

REPOSITORIES: biostudies-other

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Cloning and expression of a novel toxin gene from Bacillus thuringiensis subsp. jegathesan encoding a highly mosquitocidal protein.

Delécluse A A Rosso M L ML Ragni A A

Applied and environmental microbiology 19951201 12

A gene, designated cry11B, encoding a 81,293-Da crystal protein of Bacillus thuringiensis subsp. jegathesan was cloned by using a gene-specific oligonucleotide probe. The sequence of the Cry11B protein, as deduced from the sequence of the cry11B gene, contains large regions of similarity with the Cry11A toxin (previously CryIVD) from B. thuringiensis subsp. israelensis. The Cry11B protein was immunologically related to both Cry11A and Cry4A proteins. The cry11B gene was expressed in a nontoxic s ...[more]

PMID: 8534090

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Project description:pTAND672-2, a 144-kb resident plasmid of Bacillus thuringiensis serovar israelensis strain TAND672, was sequenced and characterized. This extrachromosomal element carries mosquitocidal toxin-, conjugation-, and recombinase-encoding genes, together with a putative arbitrium system, a genetic module recently discovered in temperate phages controlling lysogeny-lysis transition and in mobile genetic elements (MGEs) where its function remains clarified. Using conjugation experiments, pTAND672-2 is shown to be a novel integrative and conjugative element (ICE), which can horizontally transfer from B. thuringiensis serovar israelensis to Lysinibacillus sphaericus, another mosquitocidal bacterium, where it integrates into the chromosome. Its integration and circularization are reversible and involve a single-cross recombination between 33-bp specific sites, attB in the chromosome of L. sphaericus and attP in pTAND672-2. CDS143, coding for the putative tyrosine integrase Int143 distantly related to site-specific tyrosine Xer recombinases and phage integrases, can mediate the integration of pTAND672-2 to attB. The B. thuringiensis mosquito-killing genes carried by pTAND672-2 are efficiently transcribed and expressed in L. sphaericus, displaying a slight increased toxicity in this bacterium against Aedes albopictus larvae. The occurrence of pTAND672-2-like plasmids within the Bacillus cereus group was also explored and indicated that they all share a similar genetic backbone with diverse plasmid sizes, ranging from 58 to 225 kb. Interestingly, among them, the pEFR-4-4 plasmid of Bacillus paranthracis EFR-4 and p5 of B. thuringiensis BT-59 also display conjugative capability; moreover, like pTAND672-2 displays a chimeric structure between the pCH_133-e- and pBtoxis-like plasmids, pBTHD789-3 also appears to be mosaic of two plasmids. IMPORTANCE Horizontal transfer of mobile genetic elements carrying mosquitocidal toxin genes may play a driving role in the diversity of mosquitocidal bacteria. Here, the 144-kb mosquitocidal toxin-encoding plasmid pTAND672-2 is the first verified integrative and conjugative element (ICE) identified in Bacillus thuringiensis serovar israelensis. The key tyrosine integrase Int143, involved in the specific integration, is distantly related to other tyrosine recombinases. The study also reports the occurrence and potential interspecies transmission of pTAND672-2-like plasmids with varied sizes in B. thuringiensis, Bacillus paranthracis, and Bacillus wiedmannii isolates belonging to the Bacillus cereus group. This study is important for further understanding the evolution and ecology of mosquitocidal bacteria, as well as for providing new direction for the genetic engineering of biopesticides in the control of disease-transmitting mosquitoes.

Dataset Information

Cloning and expression of a novel toxin gene from Bacillus thuringiensis subsp. jegathesan encoding a highly mosquitocidal protein.

Publications

Cloning and expression of a novel toxin gene from Bacillus thuringiensis subsp. jegathesan encoding a highly mosquitocidal protein.

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