A novel negative regulatory factor for nematicidal Cry protein gene expression in Bacillus thuringiensis.
A 3-kb HindIII fragment bearing the cry6Aa2 gene and the adjacent and intergenic regions was cloned from Bacillus thuringiensis strain YBT-1518. Two open reading frames (ORFs), namely, orf1 (termed cry6Aa2) and orf2 that were separated by an inverted-repeat sequence were identified. orf1 encoded a 54-kDa protein that exhibited high toxicity to the plant-parasitic nematode Meloidogyne hapla. The orf2 expression product was not detected by SDS-PAGE, but its mRNA was detected by RT-PCR. The orf2 coexpressed with orf1 at a high level in the absence of the inverted-repeat sequence, whereas, the expression level of orf1 was decreased. When orf2 was mutated, the level of orf1 expression was enhanced obviously. In conclusion, the inverted-repeat sequence disturbs orf2 expression, and the orf2 downregulates orf1 expression. This is an example of novel negative regulation in B. thuringiensis and a potential method for enhancing the expression level of cry genes.
Yu
Ziquan
Z
State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University,Wuhan, Hubei, P.R. China.
Bai
Peisheng
P
Bai
Peisheng
P
Ye
Weixing
W
Zhang
Fengjuan
F
Ruan
Lifang
L
Yu
Ziniu
Z
Sun
Ming
M
D000595:Amino Acid Sequence; D000818:Animals; D001413:Bacillus thuringiensis / Q000235:genetics*; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D004731:Endotoxins / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D006460:Hemolysin Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D009876:Operon; D010957:Plasmids; D012091:Repetitive Sequences, Nucleic Acid; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014431:Tylenchoidea / Q000382:microbiology
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis