Benoit Cousineau

Biography

Lyman Duff Medical Building

Research Interest

Our laboratory, located at the Duff Medical Building (room 617), offers the opportunity to acquire a solid knowledge of the principles and techniques of molecular biology, genetics and bacteriology, in a dynamic and stimulating environment. Through the diversity of our research projects, members of our laboratory widen their knowledge and expertise on a great variety of subjects. The main topic of study of our laboratory is on the 'Mechanisms and applications of group II intron mobility; Evolution of mobile group II introns; Lactococcus lactis live vaccines'. Group II introns are ribozymes. They splice autocatalytically from their pre-mRNA transcript by two consecutive transesterification reactions. This process is identical to the splicing of nuclear introns and functions to ligate the flanking exons and release the intron in the form of a lariat. Some group II introns contain open reading frames (ORFs) and are also mobile retroelements. Group II introns are found in bacteria, archaea and bacterial-derived organelles. Very little sequence similarity is observed amongst group II introns; however, their secondary structure is highly conserved. It consists of six domains (DI-DVI) that radiate from a central hub (Belfort et al., 2001). By means of a wide variety of molecular biology and genetic techniques, we study various functional and evolutionary aspects of group II introns. We use the Lactococcus lactis Ll.LtrB group II intron as a model system. Ll.LtrB was the first bacterial group II intron shown to splice and mobilize in vivo. Ll.LtrB harbors a 599 amino acid ORF called LtrA in domain IV. This multifunctional intron-encoded protein has reverse transcriptase, maturase and endonuclease activities and is essential for Ll.LtrB mobility by retrohoming (Cousineau et al., 1998). Following translation, LtrA binds specifically to the Ll.LtrB intron as a dimer within the pre-mRNA. Through its maturase function, LtrA promotes self-splicing of Ll.LtrB and concurrent ligation of the flanking exons. After Ll.LtrB splicing, the LtrA dimer remains associated with the excised intron lariat as a ribonucleoprotein particle (RNP; intron RNA lariat+two LtrA proteins). Upon RNP recognition of the homing site, mainly through base-pairing interactions, the intron RNA reverse splices into the sense strand of its double-stranded DNA target. Then, the antisense strand is nicked 9 nt downstream from the intron insertion site by the endonuclease domain of LtrA. Next, using the free 3' OH generated by the endonuclease cleavage as a primer, LtrA generates a cDNA copy of the intron RNA by a process termed target-primed reverse transcription (TPRT). The final steps of the Ll.LtrB retrohoming pathway are independent of the RecA-dependent homologous recombination pathway (Cousineau et al., 1998). This intron can also invade sites other than its homing site through a process called retrotransposition (Cousineau et al., 2000; Ichiyanagi et al., 2002).