High-throughput sequencing of sorted expression libraries reveals inhibitors of bacterial cell division.

ABSTRACT: BACKGROUND:Bacterial filamentation occurs when rod-shaped bacteria grow without dividing. To identify genetically encoded inhibitors of division that promote filamentation, we used cell sorting flow cytometry to enrich filamentous clones from an inducible expression library, and then identified the cloned DNA with high-throughput DNA sequencing. We applied the method to an expression library made from fragmented genomic DNA of uropathogenic E. coli UTI89, which undergoes extensive reversible filamentation in urinary tract infections and might encode additional regulators of division. RESULTS:We identified 55 genomic regions that reproducibly caused filamentation when expressed from the plasmid vector, and then further localized the cause of filamentation in several of these to specific genes or sub-fragments. Many of the identified genomic fragments encode genes that are known to participate in cell division or its regulation, and others may play previously-unknown roles. Some of the prophage genes identified were previously implicated in cell division arrest. A number of the other fragments encoded potential short transcripts or peptides. CONCLUSIONS:The results provided evidence of potential new links between cell division and distinct cellular processes including central carbon metabolism and gene regulation. Candidate regulators of the UTI-associated filamentation response or others were identified amongst the results. In addition, some genomic fragments that caused filamentation may not have evolved to control cell division, but may have applications as artificial inhibitors. Our approach offers the opportunity to carry out in depth surveys of diverse DNA libraries to identify new genes or sequences encoding the capacity to inhibit division and cause filamentation.

SUBMITTER: Mediati DG 

PROVIDER: S-EPMC6206680 | biostudies-literature | 2018 Oct

REPOSITORIES: biostudies-literature