Transposons are mobile genetic elements that drive evolution and adaptation throughout the tree of life. In bacteria, they often transfer antibiotic resistance genes and contribute to the emergence of multidrug-resistant pathogens. In this talk, I will present our recent discoveries on a group of transposons that efficiently propagate antibiotic resistance genes across diverse microbial communities. Using a dedicated computational pipeline, we mapped the most wide-spread transposons in bacterial genomes and characterized their diversity, genetic cargo, and transfer dynamics. Selected elements were experimentally reconstituted to describe their molecular mechanisms and biochemical pathways. We determined numerous high-resolution crystal and cryo-EM structures of the protein-DNA assemblies involved in transposon movement. The structures capture various stages of transposition and reveal an intricate interplay between the core transposition machinery and host-encoded proteins. Protein interactions shape transposon and genomic DNA in unique ways to cut, exchange and re-join strands in a tightly regulated manner. Importantly, our data also explain how specific DNA distortion and cleavage mechanisms enable the transposons to move between diverse genomic sites, expanding gene transfer across many species. These results shed new light onto the molecular strategies of antibiotic resistance carrying transposons and show how they evolved to effectively spread key genetic traits. Our insights also open new opportunities to develop strategies for blocking transposition and thereby help control resistance spreading.

Computer analysis of a conserved domain, BRCT, first described at the carboxyl terminus of the breast cancer protein BRCA1, a p53 binding protein (53BP1), and the yeast cell cycle checkpoint protein RAD9 revealed a large superfamily of domains that occur predominantly in proteins involved in cell cycle checkpoint functions responsive to DNA damage. The BRCT domain consists of approximately 95 amino acid residues and occurs as a tandem repeat at the carboxyl terminus of numerous proteins, but has been observed also as a tandem repeat at the amino terminus or as a single copy. The BRCT superfamily presently includes approximately 40 nonorthologous proteins, namely, BRCA1, 53BP1, and RAD9; a protein family that consists of the fission yeast replication checkpoint protein Rad4, the oncoprotein ECT2, the DNA repair protein XRCC1, and yeast DNA polymerase subunit DPB11; DNA binding enzymes such as terminal deoxynucleotidyltransferases, deoxycytidyl transferase involved in DNA repair, and DNA-ligases III and IV; yeast multifunctional transcription factor RAP1; and several uncharacterized gene products. Another previously described domain that is shared by bacterial NAD-dependent DNA-ligases, the large subunits of eukaryotic replication factor C, and poly(ADP-ribose) polymerases appears to be a distinct version of the BRCT domain. The retinoblastoma protein (a universal tumor suppressor) and related proteins may contain a distant relative of the BRCT domain. Despite the functional diversity of all these proteins, participation in DNA damage-responsive checkpoints appears to be a unifying theme. Thus, the BRCT domain is likely to perform critical, yet uncharacterized, functions in the cell cycle control of organisms from bacteria to humans. The carboxyterminal BRCT domain of BRCA1 corresponds precisely to the recently identified minimal transcription activation domain of this protein, indicating one such function.