Assembly and Function of the Type IV Secretion System Channel from Integrative and Conjugative Elements – FUN-ICE

The emergence and the spread of antibiotic resistance are natural phenomena of bacterial adaptation to antimicrobial exposure. The last decades, the inappropriate use of antibiotics in human and veterinary medicine has accelerated these processes, leading to the emergence of pathogenic bacteria that are resistant to one or several antibiotics. Conjugative transfer is one of the main mechanisms driving the spread of antibiotic resistance genes among bacteria. Conjugative transfer is carried out by mobile conjugative elements including plasmids and Integrative and Conjugative Elements (ICEs). In Gram-positive bacteria, these mobile genetic elements are highly prevalent in many relevant streptococcal, staphylococcal and enterococcal pathogens responsible for at least half of all hospital-acquired infections. Conjugation involves a close contact between donor and recipient cells and is mediated by a specialized multiprotein complex called the conjugative-type IV secretion system (T4SS). Despite T4SSs have been well described in Gram-negative systems, there is a great lack of knowledge on the biogenesis of T4SSs in Gram-positive bacteria and how they ensure DNA transfer from the donor to recipient cell. The Fun-ICE project ambitions to obtain a comprehensive picture on the mechanistic of assembly and DNA transfer mediated by the T4SS of ICESt3, a prototype from the Tn916 superfamily largely found in streptococci. The ICESt3 genetic element encompasses a conjugation module that encodes for 14 genes (orfA to orfN) involved in DNA transfer. In the first line of the project, we will define and characterize transfer proteins involved in ICESt3 conjugation. In the second line, we will determine the ICESt3 T4SS architecture by structural and biochemical characterization of native or reconstituted T4SS complexes. In the third part of this project we will evaluate the involvement of OrfG in T4SS assembly and DNA translocation by analyzing its structure-function.