Expression of the sigma35 and cry2AB genes involved in Bacillus thuringiensis virulence

Abstract

There are several genes involved in Bacillus thuringiensis sporulation. The regulation and expression of these genes results in an upregulation in Cry protein production, and this is responsible for the death of insect larvae infected by Bacillus thuringiensis. Gene expression was monitored in Bacillus thuringiensis during three developmental phases. DNA macroarrays were constructed for selected genes whose sequences are available in the GenBank database. These genes were hybridized to cDNA sequences from B. thuringiensis var. kurstaki HD-1. cDNA probes were synthesized by reverse transcription from B. thuringiensis RNA templates extracted during the exponential (log) growth, stationary and sporulation phases, and labeled with 33 PadCTP. Two genes were differentially expressed levels during the different developmental phases. One of these genes is related to sigma factor (sigma 35), and the other is a cry gene (cry2Ab). There were differences between the differential levels of expression of various genes and among the expression detected for different combinations of the sigma factor and cry2Ab genes. The maximum difference in expression was observed for the gene encoding sigma 35 factor in the log phase, which was also expressed at a high level during the sporulation phase. The cry2Ab gene was only expressed at a high level in the log phase, but at very low levels in the other phases when compared to the sigma 35. EXPRESSÃO DOS GENES SIGMA 35 E CRY2AB ENVOLVIDOS NA VIRULÊNCIA DE Bacillus thuringiensis RESUMO: Muitos genes estão envolvidos nos mecanismos de esporulação da bactéria Bacillus thuringiensis. A regulação e expressão desses genes resultam em uma produção massiva da proteína Cry, responsável pela morte das larvas de muitos insetos. Neste trabalho monitorou-se a expressão de genes de Bacillus thuringiensis, ao longo de três fases de seu desenvolvimento. Foram construídos macroarrays de DNA dos genes selecionados, cujas seqüências estão disponibilizadas no GenBank. Estes genes foram hibridizados com cDNAs obtidos de B. thuringiensis kurstaki HD-1. As sondas de cDNA foram sintetizadas a partir da transcrição reversa do RNA da bactéria, extraído durante as fases de crescimento logarítmico, estacionária e esporulativa, marcadas com 33 PadCTP. A expressão diferencial encontrada foi significativa para dois genes de B.thuringiensis, um relacionado aos fatores sigma (sigma 35) e outro ao gene cry (cry2Ab). Detectaram-se diferenças entre as médias de expressão do fator sigma e do gene cry2Ab. Os valores máximos de expressão diferencial foram obtidos para o gene codificador do fator sigma 35 na fase log e na fase esporulativa. Na análise de médias observou-se expressão do gene cry2Ab apenas na fase log; no entanto, de forma bem mais baixa quando comparado com a expressão de sigma 35 , nas três fases.

Bacillus thuringiensis berliner cells population growth in some naturally media and the patogenicity against Plutella xylostella Caterpilars

“…Production from the properties of the crystal protein stored in crystals on stem cells has proven especially at the beginning of sporulation (Sedlak et al,, 2000;Guidelli-thuler et al, 2009). Some Cry-genes have been shown to be transcribed from two promoters overlap BTI and BtII by RNA polymerase containing sporulation depends on factor of sigma σE and σK (Sedlak et al, 2000) and mutations in the consensus of σE has been shown to inhibit the transcription of a promoter BTI and BtII (Sedlak et al, 2000).…”

Bacillus thuringiensis Applications in Agriculture

“…The life cycle of Bt can be divided for convenience into phases and these are Phase I: vegetative growth; Phase II: transition to sporulation; Phase III: sporulation; and Phase IV: spore maturation and cell lysis (Hilbert and Piggot 2004;Berbert-Molina et al 2008). The production of the characteristic insecticidal (Cry) proteins deposited in crystals in the mother cell have been shown to mainly start from the onset of sporulation (Sedlak et al 2000;Xia et al 2005;Guidelli-Thuler et al 2009;Pérez-García et al 2010). A number of cry-genes have been shown to be transcribed from two overlapping promoters BtI and BtII by RNA polymerases that contain sporulation dependent sigma factors σ E and σ K (Sedlak et al 2000;Hilbert and Piggot 2004) and a mutation in the consensus region of σ E has been shown to inhibit transcription from BtI and BtII promoters (Sedlak et al 2000).…”