Abstract
Chromosome 2 of Vibrio cholerae carries a chromosomal superintegron, composed of an integrase, a cassette integration site (attI) and an array of mostly promoterless gene cassettes. We determined the precise location of the promoter, Pc, which drives the transcription of the first cassettes of the V. cholerae superintegron. We found that cassette mRNA starts 65 bp upstream of the attI site, so that the inversely oriented promoters Pc and Pint (integrase promoter) partly overlap, allowing for their potential co-regulation. Pint was previously shown to be induced during the SOS response and is further controlled by the catabolite repression cAMP-CRP complex. We found that cassette expression from Pc was also controlled by the cAMP-CRP complex, but is not part of the SOS regulon. Pint and Pc promoters were both found to be induced in rich medium, at high temperature, high salinity and at the end of exponential growth phase, although at very different levels and independently of sigma factor RpoS. All these results show that expression from the integrase and cassette promoters can take place at the same time, thus leading to coordinated excisions and integrations within the superintegron and potentially coupling cassette shuffling to immediate selective advantage.
Highlights
Vibrio cholerae, a marine bacterium, is the causative agent of cholera in humans, a severe diarrheal disease often fatal by dehydration
The gene cassettes are usually composed of a single coding sequence associated with a recombination site, attC, which is the target of IntI-mediated site-specific recombination with a cassette integration site (attI)
Determining the position of the transcriptional start site is crucial to study the expression of the cassette array in the V. cholerae superintegron
Summary
A marine bacterium, is the causative agent of cholera in humans, a severe diarrheal disease often fatal by dehydration. All characterized integrons are composed of a stable platform, which contains the functional elements required for the system operation, associated with a variable array of individual gene cassettes which may encode accessory functions [5]. The integron platform is defined by an integrase encoding gene, intI, a primary recombination site, attI, and a strong promoter, Pc, located upstream of attI, either in the non coding sequence upstream of intI or in intI itself (Figure 1). The gene cassettes are usually composed of a single coding sequence associated with a recombination site, attC, which is the target of IntI-mediated site-specific recombination with attI. In V. cholerae, the SI gathers hundreds of genes cassettes (179 in N16961 strain [6]), carrying the highly conserved V. cholerae specific attC sites, termed VCR (Vibrio cholerae repeats), in a single cassette array. It was suggested that cassette arrays carry important functions that could mediate adaptation and interactions with the environment [5,12]
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