Inverse Correlation between Promoter Strength and Excision Activity in Class 1 Integrons

Thomas Jové,Sandra Da Re,Didier Mazel,Marie-Cécile Ploy,François Denis,Josep Casadesús

doi:10.1371/journal.pgen.1000793

Abstract

Class 1 integrons are widespread genetic elements that allow bacteria to capture and express gene cassettes that are usually promoterless. These integrons play a major role in the dissemination of antibiotic resistance among Gram-negative bacteria. They typically consist of a gene (intI) encoding an integrase (that catalyzes the gene cassette movement by site-specific recombination), a recombination site (attI1), and a promoter (Pc) responsible for the expression of inserted gene cassettes. The Pc promoter can occasionally be combined with a second promoter designated P2, and several Pc variants with different strengths have been described, although their relative distribution is not known. The Pc promoter in class 1 integrons is located within the intI1 coding sequence. The Pc polymorphism affects the amino acid sequence of IntI1 and the effect of this feature on the integrase recombination activity has not previously been investigated. We therefore conducted an extensive in silico study of class 1 integron sequences in order to assess the distribution of Pc variants. We also measured these promoters' strength by means of transcriptional reporter gene fusion experiments and estimated the excision and integration activities of the different IntI1 variants. We found that there are currently 13 Pc variants, leading to 10 IntI1 variants, that have a highly uneven distribution. There are five main Pc-P2 combinations, corresponding to five promoter strengths, and three main integrases displaying similar integration activity but very different excision efficiency. Promoter strength correlates with integrase excision activity: the weaker the promoter, the stronger the integrase. The tight relationship between the aptitude of class 1 integrons to recombine cassettes and express gene cassettes may be a key to understanding the short-term evolution of integrons. Dissemination of integron-driven drug resistance is therefore more complex than previously thought.

Highlights

Integrons are natural genetic elements that can acquire, exchange and express genes within gene cassettes
Multi-resistant integrons (MRI) play a major role in the dissemination of antibiotic resistance among Gram-negative bacteria, through horizontal gene transfer [8]
We analyzed the sequences of 321 distinct class 1 integrons containing the complete sequences of both gene cassette arrays and Pc-P2 promoters

Summary

Introduction

Integrons are natural genetic elements that can acquire, exchange and express genes within gene cassettes. The integron platform is composed of a gene, intI, that encodes a site-specific recombinase, IntI, a recombination site, attI, and a functional promoter, Pc, divergent to the integrase gene [1] (Figure 1). Gene cassettes are small mobile units composed of one coding sequence and a recombination site, attC. Integrons exchange gene cassettes through integrase-catalyzed site-specific recombination between attI and attC sites, resulting in the insertion of the gene cassette at the attI site, or between two attC sites, leading to the excision of the gene cassette(s) from the gene cassette array [2,3,4,5,6]. Five classes of MRI have been described on the basis of the integrase coding sequence, class 1 being the most prevalent [8]

Methods

Results

Conclusion