The molecular basis of odor recognition

Lancet, Doron; Pace, Umberto

doi:10.1016/0968-0004(87)90032-6

Cited by 77 publications

(34 citation statements)

References 26 publications

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“…Lipids were purchased from Avanti Polar Lipids (Birmingham, AL). After bilayer formation, ciliary membrane vesicles, preequilibrated in 0.5 M sucrose, were added to the cis side of the bilayer to a final protein concentration of [1][2][3][4][5] ,ug/ml. The vesicles were fused to the bilayer by first adding 15 by nanomolar concentrations of odorants (Fig.…”

Section: Methodsmentioning

confidence: 99%

“…It has been postulated that signal transduction at the olfactory membrane may be mediated by cAMP and that this second messenger would activate ion channels either directly or via a protein kinase cascade (2,4). The electroolfactogram, a summated potential recorded from the surface of the olfactory epithelium after application of odorant, is modified by cAMP (5,6).…”

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confidence: 99%

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Activation by odorants of a multistate cation channel from olfactory cilia.

Labarca

Simon

Anholt

1988

Proc. Natl. Acad. Sci. U.S.A.

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Single-channel records were obtained after fusion of ciliary membranes from the olfactory epithelium of Rana catesbeiana to planar lipid bilayers, and odorantactivated cation-selective channels were identified. In addition, a 190-pS potassium-selective channel and a 40-pS cationselective channel were found in a 0.2 M salt-containing buffer. Odorant-sensitive channels were directly and reversibly activated by nanomolar concentrations of the bell pepper odorant 3-isobutyl-2-methoxypyrazine and the citrus odorant 3,7,-dimethyl-2,6-octadienenitrile. These channels display burst kinetics, multiple conductance levels between 35 and 420 pS, and open times in the millisecond range. With increasing concentrations of odorant, the probability of populating the higher conductance levels increases. These results show that direct activation of channels by odorants may mediate excitation of the olfactory receptor cell.Initial transduction events in olfaction occur at the membranes of chemosensory cilia, which protrude from the dendritic tips of olfactory receptor neurons (1-3). The binding of an odorant to the ciliary membrane must result directly or indirectly in the activation of ion channels, which in turn culminates in the generation of action potentials.It has been postulated that signal transduction at the olfactory membrane may be mediated by cAMP and that this second messenger would activate ion channels either directly or via a protein kinase cascade (2, 4). The electroolfactogram, a summated potential recorded from the surface of the olfactory epithelium after application of odorant, is modified by cAMP (5, 6). Olfactory cilia contain a high activity of adenylate cyclase (7). In addition, Nakamura and Gold (8) recently patch-clamped olfactory cilia and observed a conductance that is activated by micromolar concentrations of both cAMP and cGMP. Moreover, the olfactory adenylate cyclase activity can be stimulated by micromolar concentrations of some, but not all, odorants via a regulatory GTP-binding protein (7,9,10 We fused ciliary membrane vesicles from the olfactory epithelium of the bullfrog, Rana catesbeiana, to planar lipid bilayers and observed three distinct cation-selective channels. Only one of these, a multistate channel with millisecond open times, is reversibly activated by nanomolar concentrations of two structurally unrelated odorants. Our observations indicate that direct activation of ion channels by odorants may underlie the initial event in olfactory transduction. MATERIALS AND METHODSBullfrogs (R. catesbeiana) were obtained from Amphibians of North America (Nashville, TN) and were maintained in a well-ventilated facility in a tank with circulating water. Olfactory cilia were prepared as described (14) and resuspended in a small volume of Ringer's solution containing 2 mM EGTA. The protein concentration of the suspension was measured by the method of Lowry et al. (15) with bovine serum albumin as standard.The bilayer system used was modeled after the description of the apparatus by Alvarez (1...

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Section: Methodsmentioning

confidence: 99%

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confidence: 99%

Activation by odorants of a multistate cation channel from olfactory cilia.

Labarca

Simon

Anholt

1988

Proc. Natl. Acad. Sci. U.S.A.

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“…Olfactory receptors (ORs) are seven-transmembrane domain (7TM) proteins (Lancet and Pace 1987;Reed 1990;Buck and Axel 1991). Previous work (Lancet and Ben-Arie 1993) suggested their classification into at least eight families within the G proteincoupled receptor (GPCR) superfamily.…”

Section: Introductionmentioning

confidence: 99%

The olfactory receptor gene superfamily: data mining, classification, and nomenclature

et al. 2000

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Abstract. The vertebrate olfactory receptor (OR) subgenome harbors the largest known gene family, which has been expanded by the need to provide recognition capacity for millions of potential odorants. We implemented an automated procedure to identify all OR coding regions from published sequences. This led us to the identification of 831 OR coding regions (including pseudogenes) from 24 vertebrate species. The resulting dataset was subjected to neighbor-joining phylogenetic analysis and classified into 32 distinct families, 14 of which include only genes from tetrapodan species (Class II ORs). We also report here the first identification of OR sequences from a marsupial (koala) and a monotreme (platypus). Analysis of these OR sequences suggests that the ancestral mammal had a small OR repertoire, which expanded independently in all three mammalian subclasses. Classification of "fishlike" (Class I) ORs indicates that some of these ancient ORs were maintained and even expanded in mammals.A nomenclature system for the OR gene superfamily is proposed, based on a divergence evolutionary model. The nomenclature consists of the root symbol 'OR', followed by a family numeral, subfamily letter(s), and a numeral representing the individual gene within the subfamily. For example, OR3A1 is an OR gene of family 3, subfamily A, and OR7E12P is an OR pseudogene of family 7, subfamily E. The symbol is to be preceded by a species indicator. We have assigned the proposed nomenclature symbols for all 330 human OR genes in the database. A WWW tool for automated name assignment is provided.

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“…OR genes encode seven-transmembrane-domain proteins [2,4,5], which are responsible for the recognition and differentiation of millions of odorants. OR genes were first DEFOG: A Practical Scheme for Deciphering…”

Section: Introductionmentioning

confidence: 99%

DEFOG: A Practical Scheme for Deciphering Families of Genes

et al. 2002

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We developed a novel efficient scheme, DEFOG (for "deciphering families of genes"), for determining sequences of numerous genes from a family of interest. The scheme provides a powerful means to obtain a gene family composition in species for which high-throughput genomic sequencing data are not available. DEFOG uses two key procedures. The first is a novel algorithm for designing highly degenerate primers based on a set of known genes from the family of interest. These primers are used in PCR reactions to amplify the members of the gene family. The second combines oligofingerprinting of the cloned PCR products with clustering of the clones based on their fingerprints. By selecting members from each cluster, a low-redundancy clone subset is chosen for sequencing. We applied the scheme to the human olfactory receptor (OR) genes. OR genes constitute the largest gene superfamily in the human genome, as well as in the genomes of other vertebrate species. DEFOG almost tripled the size of the initial repertoire of human ORs in a single experiment, and only 7% of the PCR clones had to be sequenced. Extremely high degeneracies, reaching over a billion combinations of distinct PCR primer pairs, proved to be very effective and yielded only 0.4% nonspecific products. characterized in rat a decade ago [2] and have since been detected in many vertebrate species [reviewed in 6]. So far, about 3000 OR genes and pseudogenes are known in 24 vertebrate species [7][8][9][10][11][12]. They are divided into 32 distinct families based on phylogenetic analysis [8].Roughly 900 OR coding sequences were found in the first draft of the human genome [9,10]. As predicted [7,13], between 53% and 63% of them have frame disruptions and are therefore considered as pseudogenes. OR genes are found on almost all human chromosomes except chromosome 20 and Y [7,9,10,13,14]. Almost 80% of the ORs are organized in clusters of six or more genes [9,10]. This is in good agreement with previous fluorescence in situ hybridization (FISH) experiments and sequencing data [7,13,14]. 296Article doi:10.1006/geno.2002.6830, available online at http://www.idealibrary.com on IDEAL The coding region of OR genes spans approximately 1 kb. This region lacks introns [2,13] and is preceded by a large intron and several short noncoding exons [16][17][18][19][20]. Inside the coding region there are several conserved segments [21] that allow easy amplification of the intronless coding region from genomic DNA by PCR assay. The PCR product is termed the "olfactory receptor sequence tag" (OST) [7].We developed and experimentally tested a practical scheme for deciphering families of genes (DEFOG). The scheme provides a powerful means to obtain a sequence composition of a gene family in species for which high-throughput genomic sequencing data are unavailable. To validate DEFOG, we tested it on the human OR gene superfamily. Starting with a limited number of human ORs, it almost tripled the size of this set in a single experiment. We suggest that DEFOG can be successfully appli...

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The molecular basis of odor recognition

Cited by 77 publications

References 26 publications

Activation by odorants of a multistate cation channel from olfactory cilia.

Activation by odorants of a multistate cation channel from olfactory cilia.

The olfactory receptor gene superfamily: data mining, classification, and nomenclature

DEFOG: A Practical Scheme for Deciphering Families of Genes

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