title: Nitric oxide regulation of cyclic di-GMP synthesis and hydrolysis in Shewanella woodyi.
abstract: Although several reports have documented nitric oxide (NO) regulation of biofilm formation, the molecular basis of this phenomenon is unknown. In many bacteria, an H-NOX (heme-nitric oxide/oxygen-binding) gene is found near a diguanylate cyclase (DGC) gene. H-NOX domains are conserved hemoproteins that are known NO sensors. It is widely recognized that cyclic di-GMP (c-di-GMP) is a ubiquitous bacterial signaling molecule that regulates the transition between motility and biofilm. Therefore, NO may influence biofilm formation through H-NOX regulation of DGC, thus providing a molecular-level explanation for NO regulation of biofilm formation. This work demonstrates that, indeed, NO-bound H-NOX negatively affects biofilm formation by directly regulating c-di-GMP turnover in Shewanella woodyi strain MS32. Exposure of wild-type S. woodyi to a nanomolar level of NO resulted in the formation of thinner biofilms, and less intracellular c-di-GMP, than in the absence of NO. Also, a mutant strain in the gene encoding SwH-NOX showed a decreased level of biofilm formation (and a decreased amount of intracellular c-di-GMP) with no change observed upon NO addition. Furthermore, using purified proteins, it was demonstrated that SwH-NOX and SwDGC are binding partners. SwDGC is a dual-functioning DGC; it has diguanylate cyclase and phosphodiesterase activities. These data indicate that NO-bound SwH-NOX enhances c-di-GMP degradation, but not synthesis, by SwDGC. These results support the biofilm growth data and indicate that S. woodyi senses nanomolar NO with an H-NOX domain and that SwH-NOX regulates SwDGC activity, resulting in a reduction in c-di-GMP concentration and a decreased level of biofilm growth in the presence of NO. These data provide a detailed molecular mechanism for NO regulation of c-di-GMP signaling and biofilm formation.
authors.affiliation: Department of Chemistry, Stony Brook University, Stony Brook, New York 11794-3400, United States.
authors.forename: Niu
authors.initials: N
authors.lastname: Liu
authors.forename: Yueming
authors.initials: Y
authors.lastname: Xu
authors.forename: Sajjad
authors.initials: S
authors.lastname: Hossain
authors.forename: Nick
authors.initials: N
authors.lastname: Huang
authors.forename: Dan
authors.initials: D
authors.lastname: Coursolle
authors.forename: Jeffrey A
authors.initials: JA
authors.lastname: Gralnick
authors.forename: Elizabeth M
authors.initials: EM
authors.lastname: Boon
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D018441:Biofilms / Q000254:growth & development; D006152:Cyclic GMP / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000378:metabolism; D004269:DNA, Bacterial / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D005798:Genes, Bacterial; D006868:Hydrolysis; D007700:Kinetics; D008954:Models, Biological; D009154:Mutation; D009569:Nitric Oxide / Q000378:metabolism*; D010727:Phosphoric Diester Hydrolases / Q000235:genetics / Q000378:metabolism; D019761:Phosphorus-Oxygen Lyases / Q000235:genetics / Q000378:metabolism; D020592:Shewanella / Q000235:genetics / Q000502:physiology*; D015398:Signal Transduction
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D009569:Nitric Oxide; C062025:bis(3',5')-cyclic diguanylic acid; D010727:Phosphoric Diester Hydrolases; D019761:Phosphorus-Oxygen Lyases; C060720:diguanylate cyclase; D006152:Cyclic GMP