S. marcescens subclass C group II intron as a potential intermediate in the formation of an integron gene cassette

CECILIA QUIROGA; DANIELA CENTRÓN; PAUL H. ROY

Cheribourg Hotel, Sherbroke, Quebec, Canada.

Congreso; Eastern Riboclub; 2003

Integrons are genomic elements that code for an integrase, a tyrosine recombinase that allows the insertion and excision of gene cassettes into a specific recombination site or attI. Cassettes usually consist of a promoterless gene accompanied by an attC site, a palindromic sequence of 60-140 bp. The integrase recognizes a secondary structure formed by the heptamer GTTPuPuPuPy and its inverse core site CAAPyPyPyPu. Thus, it integrates and excises cassettes by attI x attC and attC x attC recombination. In plasmid-borne integrons, where cassettes are mostly antibiotic resistance genes, analysis indicates that the structural genes and their attC sites have evolved separately before being assembled. A major unanswered question is how structural genes become associated with attC sites to form cassettes. We have found a group II (GII) intron, Smtr, inserted between the structural gene-attC site junctions in a class 1 integron from Serratia marcescens. Many similar cases have been reported in Salmonella typhymurium, Pseudomonas aeruginosa, Acinetobacter sp., Vibrio cholerae; but also, GII introns have been found associated to either the gene or the recombination site attC in the surroundings of a chromosomal integrase gene in Nitrosomonas europaea and Shewanella oneidensis. Based on these findings, we are attempting to prove that GII introns may capture attC sites (possibly by retrohoming to the left end of attC sites) and structural genes (possibly by the less specific mechanism of ectopic insertion) and recombine to assemble them.  We have proved that the Smtr GII intron homes exactly in TA/ACAA, the junction point between the structural gene and the inverse core site of the attC of phylogenetically different cassettes. We have also studied by site directed mutagenesis the stability of the EBS1 and IBS1 interactions and the role of the poly U tail in the attC site.