PMID:21421754

Citation	Rhodes, RG, Pucker, HG and McBride, MJ (2011) Development and use of a gene deletion strategy for Flavobacterium johnsoniae to identify the redundant gliding motility genes remF, remG, remH, and remI. J. Bacteriol. 193:2418-28
Abstract	Cells of Flavobacterium johnsoniae exhibit rapid gliding motility over surfaces. Cell movement is thought to involve motor complexes comprised of Gld proteins that propel the cell surface adhesin SprB. The four distal genes of the sprB operon (sprC, sprD, sprB, and sprF) are required for normal motility and for formation of spreading colonies, but the roles of the remaining three genes (remF, remG, and fjoh_0982) are unclear. A gene deletion strategy was developed to determine whether these genes are involved in gliding. A spontaneous streptomycin-resistant rpsL mutant of F. johnsoniae was isolated. Introduction of wild-type rpsL on a plasmid restored streptomycin sensitivity, demonstrating that wild-type rpsL is dominant to the mutant allele. The gene deletion strategy employed a suicide vector carrying wild-type rpsL and used streptomycin for counterselection. This approach was used to delete the region spanning remF, remG, and fjoh_0982. The mutant cells formed spreading colonies, demonstrating that these genes are not required for normal motility. Analysis of the genome revealed a paralog of remF (remH) and a paralog of remG (remI). Deletion of remH and remI had no effect on motility of wild-type cells, but cells lacking remF and remH, or cells lacking remG and remI, formed nonspreading colonies. The motility defects resulting from the combination of mutations suggest that the paralogous proteins perform redundant functions in motility. The rpsL counterselection strategy allows construction of unmarked mutations to determine the functions of individual motility proteins or to analyze other aspects of F. johnsoniae physiology.
Links	PubMed PMC3133171 Online version:10.1128/JB.00117-11
Keywords	Anti-Bacterial Agents/pharmacology; Bacterial Proteins/genetics; DNA Mutational Analysis/methods; Drug Resistance, Bacterial; Flavobacterium/genetics; Flavobacterium/physiology; Genes, Bacterial; Genetic Vectors; Genetics, Microbial/methods; Locomotion; Plasmids; Recombination, Genetic; Ribosomal Proteins/genetics; Selection, Genetic; Sequence Deletion; Streptomycin/pharmacology
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
FLAJ1:A1E5U5		GO:0071976: cell gliding	ECO:0000315:		P		Fig. 9A and B shows that the wildtype cells glide successfully whereas the sprB deletion mutant glides hardly at all.	complete CACAO 4859
FLAJ1:A1E5U5	involved_in	GO:0071976: cell gliding	ECO:0000315: mutant phenotype evidence used in manual assertion		P		Seeded From UniProt	complete

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