Danger perception and stress response through an olfactory sensor for the bacterial metabolite hydrogen sulfide

•Type B cells are ultrasensitive H2S detectors that integrate distinct danger inputs•Cnga2 and Trpc2 operate in separate subcellular compartments of the same cell•Cnga2 but not is required for detection by type cells•H2S drives stress hormone release, self-grooming, place avoidance The olfactory system serves a critical function as to trigger defense responses essential survival. cellular molecular mechanisms drive such defenses mammals incompletely understood. Here, we have discovered an sensor highly poisonous bacterial metabolite hydrogen sulfide (H2S) mice. An atypical class sensory neurons main epithelium, cells, activated both low O2. These two stimuli trigger, respectively, Cnga2- Trpc2-signaling pathways, which compartments, cilia dendritic knob. This activation defensive responses: elevation ACTH, stress-related self-grooming behavior, conditioned avoidance. Our findings identify previously unknown signaling paradigm mammalian olfaction define chemosensory inputs from external environment with appropriate outputs. detector early warning remain poorly efficient way danger-associated chemicals generated harmful microbes, resulting social distancing (Curtis, 2014Curtis V.A. Infection-avoidance behaviour humans other animals.Trends Immunol. 2014; 35: 457-464Abstract Full Text PDF PubMed Scopus (131) Google Scholar; Rivas et al., 2014Rivas F.V. Chervonsky A.V. Medzhitov R. ART immunology.Trends 451Abstract (11) Scholar). Mice avoid sick conspecifics through chemostimuli vomeronasal (VSNs) (Boillat 2015Boillat M. Challet L. Rossier D. Kan C. Carleton A. Rodriguez I. mediates conspecific avoidance.Curr. Biol. 2015; 25: 251-255Abstract (70) Scholar), pathogen-associated, bacteria-derived molecules G protein-coupled chemoreceptors these (Bufe 2019Bufe B. Teuchert Y. Schmid Pyrski Pérez-Gómez Eisenbeis J. Timm T. Ishii Lochnit G. Bischoff al.Bacterial MgrB peptide activates chemoreceptor Fpr3 mouse accessory behaviour.Nat. Commun. 2019; 10: 4889Crossref (15) Similar behaviors triggered predator-derived odors involve multiple subsystems, including epithelium (MOE) (Dewan 2013Dewan Pacifico Zhan Rinberg Bozza Non-redundant coding aversive odours pathway.Nature. 2013; 497: 486-489Crossref (134) Ferrero 2011Ferrero D.M. Lemon J.K. Fluegge Pashkovski S.L. Korzan W.J. Datta S.R. Spehr Fendt Liberles S.D. Detection carnivore odor prey.Proc. Natl. Acad. Sci. U S 2011; 108: 11235-11240Crossref (245) Kobayakawa 2007Kobayakawa K. Matsumoto H. Oka Imai Ikawa Okabe Ikeda Itohara S. Kikusui al.Innate versus learned odour processing bulb.Nature. 2007; 450: 503-508Crossref (461) Li Liberles, 2015Li Q. Aversion attraction olfaction.Curr. R120-R129Abstract (103) 2015Pérez-Gómez Bleymehl Stein Birnbaumer Munger Leinders-Zufall Zufall F. Chamero P. Innate predator aversion driven parallel subsystems converge ventromedial hypothalamus.Curr. 1340-1346Abstract (95) We hypothesized has evolved dedicated life-threatening environment. One chemical (H2S), toxic molecule poses severe threat all organisms use oxygen (O2) primary source metabolic energy (Beauchamp 1984Beauchamp Jr., R.O. Bus J.S. Popp J.A. Boreiko C.J. Andjelkovich D.A. A review literature on toxicity.Crit. Rev. Toxicol. 1984; 13: 25-97Crossref (774) Steventon, 2007Steventon G.B. Hydrogen sulphide.in: Waring R.H. Steventon Mitchell S.C. Molecules Death. Imperial College Press, London, UK2007: 131-147Crossref (2) one principal compounds involved natural sulfur cycle played major role during life’s evolution (Olson Straub, 2016Olson K.R. Straub K.D. metabolism signaling.Physiology (Bethesda). 2016; 31: 60-72Crossref (139) Pal 2018Pal V.K. Bandyopadhyay Singh physiology pathogenesis bacteria viruses.IUBMB Life. 2018; 70: 393-410Crossref (39) usually produced breakdown organic material under anaerobic conditions therefore typically coincides O2 depletion. existence specialized could provide selective advantage, especially rodents display burrowing behavior (Deacon, 2006Deacon R.M. Burrowing rodents: sensitive method detecting behavioral dysfunction.Nat. Protoc. 2006; 1: 118-121Crossref (182) Moreover, volatile can signal presence spoiled food or predators (Li 2016Li Ahmed Zhang Pan Matsunami Burger J.L. Block E. Batista V.S. Zhuang Smelling sulfur: copper silver regulate response human odorant receptor OR2T11 low-molecular-weight thiols.J. Am. Chem. Soc. 138: 13281-13288Crossref (43) elevated formation gut been linked pathogen infection disease (Stacy 2021Stacy Andrade-Oliveira V. McCulloch Hild Oh J.H. Perez-Chaparro P.J. Sim C.K. Lim A.I. Link V.M. Enamorado al.Infection trains host microbiota-enhanced resistance pathogens.Cell. 2021; 184: 615-627.e17Abstract (52) Wu 2012Wu Y.C. Wang X.J. Yu Chan F.K. Cheng A.S. Sung J.J. W.K. Cho C.H. lowers proliferation induces protective autophagy colon epithelial cells.PLoS ONE. 2012; 7: e37572Crossref (80) Thus, rise depletion likely encountered animals biological contexts related fundamental principles transduction, each depending ion channel (Brennan Zufall, 2006Brennan P.A. Pheromonal communication vertebrates.Nature. 444: 308-315Crossref (348) Firestein, 2001Firestein How makes sense scents.Nature. 2001; 413: 211-218Crossref (845) Kaupp, 2010Kaupp U.B. Olfactory signalling vertebrates insects: differences commonalities.Nat. Neurosci. 2010; 11: 188-200Crossref (358) canonical (OSNs) MOE rely cyclic nucleotide-gated (CNG) cation channel, second messenger cAMP (Firestein 1991Firestein Darrow Shepherd G.M. Activation current salamander depends protein-mediated system.Neuron. 1991; 6: 825-835Abstract (135) Nakamura Gold, 1987Nakamura Gold G.H. conductance cilia.Nature. 1987; 325: 442-444Crossref (836) 1994Zufall Firestein Cyclic channels transduction neurons.Annu. Biophys. Biomol. Struct. 1994; 23: 577-607Crossref (169) Cnga2 subunit this (Dhallan 1990Dhallan R.S. Yau K.W. Schrader K.A. Reed R.R. Primary structure functional expression nucleotide-activated neurons.Nature. 1990; 347: 184-187Crossref (507) 1995Kaupp Family nucleotide gated channels.Curr. Opin. Neurobiol. 1995; 5: 434-442Crossref (130) Zagotta Siegelbaum, 1996Zagotta W.N. Siegelbaum S.A. Structure channels.Annu. 1996; 19: 235-263Crossref (415) In contrast, VSNs transient potential (TRP) (Leypold 2002Leypold B.G. C.R. Kim M.M. Axel Altered sexual trp2 mutant mice.Proc. 2002; 99: 6376-6381Crossref (444) Liman 1999Liman E.R. Corey D.P. Dulac TRP2: candidate pheromone signaling.Proc. 1999; 96: 5791-5796Crossref (329) Stowers 2002Stowers Holy T.E. Meister Koentges Loss sex discrimination male-male aggression mice deficient TRP2.Science. 295: 1493-1500Crossref (651) phospholipase C (Lucas 2003Lucas Ukhanov diacylglycerol-gated neuron dendrites impaired TRPC2 mice: mechanism transduction.Neuron. 2003; 40: 551-561Abstract (256) It came surprise when identified hitherto unrecognized types co-expressed channels, (Omura Mombaerts, 2014Omura Mombaerts Trpc2-expressing mouse.Cell Rep. 8: 583-595Abstract (53) Scholar, Omura 2015Omura express soluble guanylate cyclase Gucy1b2.Mol. Cell. 65: 114-124Crossref (29) referred cell types, (sGC) Gucy1b2, Type (Bleymehl 2016Bleymehl Moreno-Pérez Macías Bai Z. Johnson environmental epithelium.Neuron. 92: 1196-1203Abstract (33) (Leinders-Zufall 2018Leinders-Zufall Storch U. Mederos Y Schnitzler Frank Konrad D.B. Trauner Gudermann PhoDAGs enable optical control diacylglycerol-sensitive channels.Cell 215-223.e3Abstract (31) 2021Leinders-Zufall Ojha N.K. Koike diacylglycerol photoswitching protocol studying TRPC functions tissue slices.STAR 2: 100527Crossref (1) Given production depletion, reasoned may H2S. demonstrate endowed unique logic enables them detect environment: decrease By using Gucy1b2-driven conditional knockout Cnga2, marker protein (OMP)-driven ciliopathy gene Ift88, constitutive heme oxygenase-2 (Hmox2), show Trpc2-mediated pathways knob, respectively. integration danger-related information already at earliest level discrimination. gene-targeted Gucy1b2-IRES-tauGFP strain (referred Gucy1b2-GFP), Gucy1b2 remains intact its region tauGFP, enabling characterization visualization their axonal projections bulb mapped positions Gucy1b2+ imaging Gucy1b2-GFP homozygous serial two-photon tomography (Ragan 2012Ragan Kadiri L.R. Venkataraju K.U. Bahlmann Sutin Taranda Arganda-Carreras Seung H.S. Osten Serial automated ex vivo brain imaging.Nat. Methods. 9: 255-258Crossref (406) Zapiec 2015Zapiec Multiplex assessment receptor-specific glomeruli tomography.Proc. 112: E5873-E5882Crossref 2020Zapiec zonal organization choice 2020; 30: 4220-4234.e5Abstract 2016Zapiec Bressel O.C. Khan Walz Neuropilin-1 bulb.eNeuro. 3 (ENEURO.0123-0116.2016)Crossref (12) followed segmentation GFP+ (Figures 1A, 1B , S1A–S1C; Video S1). Three-dimensional (3D) reconstructions revealed located lateral extremes area reminiscent Zolfr1507 zone (Zapiec majority clusters, dorsal ventral, only few between (Figure 1A). superimposing 3D reconstruction MOR28-IRES-gapGFP × M50-IRES-GFP-IRES-taulacZ double-heterozygous mouse, confirmed 1B). distributions Olfr1507+ OSNs differ somewhat within zone, be best appreciated ventral view animation overlaps Olfr6+ OSNs, reside different Zolfr2 https://www.cell.com/cms/asset/88ec2a09-1293-4458-a552-d24719440085/mmc2.mp4Loading ... Download .mp4 (74.26 MB) Help files S1. M50-IRES-GFP mice, Figure 1Animation depicting based dataset PD14. rendered translucent gray, shown green. posterior pans along right toward anterior, clusters cul-de-sac regions. Next, camera returning posterior. areas Olfr1507 (orange) Olfr6 (purple) overlaid x double heterozygous nine known zones (OR) MOE, although they do appear OR genes. To assess key performed immunohistochemistry (IHC) 1C), 1D 1G), 1E 1F), combined IHC GFP visualize cells. For experiments, used cryosections 1C 1D) whole-mount preparations 1E–1G). immunoreactivity (IR) was readily detectable whereas Trpc2- Gucy1b2-IR were present knob Two predictions emerged results: bimodal sensors there least additional stimulus than neurons, occur localized compartment possibly comprising 1H). tested model cellular, systems, levels. activate independently ratiometric Ca2+ fura-2/AM dissociated neural after stimulation stimulate aqueous environment, applied widely H2S-releasing agents donors, sodium hydrosulfide (NaHS) (Na2S) (Abe Kimura, 1996Abe Kimura possible endogenous neuromodulator.J. 16: 1066-1071Crossref Beauchamp Lee 2011Lee Z.W. Zhou Chen C.S. Zhao Tan Moore P.K. Deng L.W. slow-releasing donor, GYY4137, exhibits novel anti-cancer effects vitro vivo.PLoS e21077Crossref (242) 2014Zhao Biggs T.D. Xian releasing agents: chemistry applications.Chem. (Camb.). 50: 11788-11805Crossref (644 787 [82%]) responded 10 s application 0.5 μM NaHS robust elevations fluorescence persist several minutes, nearly non-type did 1I 1J). NaHS-evoked repeated times without obvious decline 1K). Na2S 1L). Nearly (85 97 [88%]) capable reduction extracellular 1L 1M). respond shifting proton (H+) concentration pH 7.7 1K) variety chemostimuli, semiochemical CS2 concentration-response measurements individual 1N 1O). experiments cover relatively wide 10%–90% operating range, spanning more orders magnitude. overall curve flat, Hill coefficient 0.6 ± 0.05 half maximal effective (EC50) 0.63 0.14 (or 0.021 0.005 ppm). threshold extremely low, close nM (0.34 ppb). comparison technical terms sensitivity, gas chromatograph outperform S1D). Hence, operates level, far below established toxicity levels (Steventon, Woodall 2005Woodall Smith R.L. Granville G.C. Proceedings health research risk symposium October 31-November 2, 2000.Inhal. 2005; 17: 593-639Crossref Scholar) S1E). Coupled earlier studies results indicate changes investigated mechanistic basis Removal abolished reversible manner 2A), indicating primarily entry into determine if hemizygous (males) (females) mutation gene, X chromosome (Zheng 2000Zheng Feinstein Peripheral differentially affected subunit.Neuron. 2000; 26: 81-91Abstract (203) knockin leaves intact, Trpc2-IRES-taumCherry (abbreviated Trpc2-mCherry) genotypes Cnga2-KO∗ MUT, asterisk referring intrinsic mCherry (which cells; see Method details STAR Methods) MUT collectively Cnga2-knockout either 8-bromo-cGMP absent KCl normal 2B 2E). similar (Trpc2-KO) genotype Trpc2-KO∗–/– cells). NaHS, 8-bromo-cGMP, normal, 2C 2F). forms Ca2+-permeable CNG cGMP (Biel 1996Biel Zong X. Hofmann diversity, structure, functions.Trends Cardiovasc. Med. 274-280Crossref (20) Dhallan involvement enzymes whether required, Trpc2-mCherry Gucy1b2-KO∗. found remained Gucy1b2-KO∗–/– 2D 2G). Using ODQ (10 μM), targets heme-nitric oxi