Genetic variation in the odorant receptor OR2J3 is associated with the ability to detect the "grassy" smelling odor, cis-3-hexen-1-ol

The ability to detect many odors varies among individuals; however, the contribution of genotype to this variation has been assessed for relatively few compounds. We have identified a genetic basis for the ability to detect the flavor compound cis-3-hexen-1-ol. This compound is typically described as "green grassy" or the smell of "cut grass," with variation in the ability to detect it linked to single nucleotide polymorphisms (SNPs) in a region on human chromosome 6 containing 25 odorant receptor genes. We have sequenced the coding regions of all 25 receptors across an ethnically mixed population of 52 individuals and identified 147 sequence variants. We tested these for association with cis-3-hexen-1-ol detection thresholds and found 3 strongly associated SNPs, including one found in a functional odorant receptor (rs28757581 in OR2J3). In vitro assays of 13 odorant receptors from the region identified 3 receptors that could respond to cis-3-hexen-1-ol, including OR2J3. This gene contained 5 predicted haplotypes across the 52 individuals. We tested all 5 haplotypes in vitro and several amino acid substitutions on their own, such as rs28757581 (T113A). Two amino acid substitutions, T113A and R226Q, impaired the ability of OR2J3 to respond to cis-3-hexen-1-ol, and together these two substitutions effectively abolished the response to the compound. The haplotype of OR2J3 containing both T113A and R226Q explains 26.4% of the variation in cis-3-hexen-1-ol detection in our study cohort. Further research is required to examine whether OR2J3 haplotypes explain variation in perceived flavor experience and the consumption of foods containing cis-3-hexen-1-ol.     

Cloning and characterization of an odorant receptor in five Pacific salmon

The olfactory system of fish is extremely important as it is able to recognize and distinguish a vast of odorous molecules involved in wide ranges of behaviors including reproduction, homing, kin recognition, feeding and predator avoidance; all of which are paramount for their survival. We cloned and characterized one type olfactory receptors (ORs) from five congeneric salmonids: lacustrine sockeye salmon (Oncorhynchus nerka), pink salmon (O. gorbuscha), chum salmon (O. keta), masu salmon (O. masou) and rainbow trout (O. mykiss). Lacustrine sockeye salmon olfactory receptor 1 (LSSOR1) showed high sequence homology to the OR subfamily, and was expressed only in the olfactory epithelium (as indicated by PCR amplified genomic DNA and cDNA). OR genes from the five salmonids examined all showed strong homology (96-99%) to each other. Hypervariable regions, believed to be ligand-binding pockets, showed homologous completely matched amino acid sequences except for one amino acid in pink salmon olfactory receptor 1 (PSOR1), revealing that these ORs may be well conserved among salmon species. These results suggest that the isolated 5 salmonid ORs might play an important role in salmon life cycles.     

Axon guidance of mouse olfactory sensory neurons by odorant receptors and the beta2 adrenergic receptor

Odorant receptors (ORs) provide the core determinant of identity for axons of olfactory sensory neurons (OSNs) to coalesce into glomeruli in the olfactory bulb. Here, using gene targeting in mice, we examine how the OR protein determines axonal identity. An OR::GFP fusion protein is present in axons, consistent with a direct function of ORs in axon guidance. When the OR coding region is deleted, we observe OSNs that coexpress other ORs that function in odorant reception and axonal identity. It remains unclear if such coexpression is normally prevented by negative feedback on OR gene choice. A drastic reduction in OR protein level produces axonal coalescence into novel, remote glomeruli. By contrast, chimeric ORs and ORs with minor mutations perturb axon outgrowth. Strikingly, the beta2 adrenergic receptor can substitute for an OR in glomerular formation when expressed from an OR locus. Thus, ORs have not evolved a unique function in axon guidance.     

Genetic variation at minisatellite loci D1S7, D4S139, D5S110 and D17S79 among three population groups of eastern India

Genetic variation at four minisatellite loci D1S7, D4S139, D5S110 and D17S79 in three predominant population groups of eastern India, namely Brahmin, Kayastha and Garo, are reported in this study. The Brahmin and Kayastha are of Indo-Caucasoid origin while the Garo community represents the Indo-Mongoloid ethnic group. The methodology employed comprised generation of HaeIII-restricted fragments of isolated DNA, Southern blotting, and hybridization using chemiluminescent probes MS1, pH30, LH1 and V1 for the four loci. All four loci were highly polymorphic in the population groups. Heterozygosity values for the four loci ranged between 0.68 and 0.95. Neither departure from Hardy-Weinberg expectations nor evidence of any association across alleles among the selected loci was observed. The gene differentiation value among the loci is moderate (G_ST = 0.027). A neighbour-joining tree constructed on the basis of the generated data shows very low genetic distance between the Brahmin and Kayastha communities in relation to the Garo. Our results based on genetic distance analysis are consistent with results of earlier studies based on serological markers and linguistic as well as morphological affiliations of these populations and their Indo-Caucasoid and Indo-Mongoloid origin. The minisatellite loci studied here were found to be not only useful in showing significant genetic variation between the populations but also to be suitable for human identity testing among eastern Indian populations.     

Allelic variation of the dopamine receptor D4 gene polymorphic region in gibbons

We examined the tandem repeat sequence of the dopamine receptor D4 (DRD4) gene in 73 individuals derived from 8 species of gibbons (genusHylobates) in an attempt to assess the variability of this gene in gibbon species.H. syndactylus (subgenusSymphalangus) andH. concolor (subgenusNomascus), which were inferred to have diverged at an early time within the family Hylobatidae, shared only long repeat (7–8) alleles. On the other hand, DRD4 was highly polymorphic in gibbons of the subgenusHylobates, with 4-, 5-, 6-, 7-, and 8-repeat alleles being recognized. In this subgenus, 4- and 5-repeat alleles were found in the species distributed mainly in the southern islands such as Sumatra, Java, and Borneo but not in the species inhabiting the Asian continent. Sequence analysis indicated that the repeat structure of the gibbon DRD4 gene was quite complex but most of the 48-bp units could be classified into several groups across the species based on sequence similarities. However, the sequence of the 7-repeat allele ofH. muelleri was unique, since the repeat units had low similarities to other units of gibbons.     

Molecular and functional characterization of the odorant receptor2 (OR2) in the tiger mosquito Aedes albopictus

In mosquitoes, the olfactory system plays a crucial role in many types of behavior, including nectar feeding, host preference selection and oviposition. Aedes albopictus, known also as the tiger mosquito, is an anthropophilic species, which in the last few years, due to its strong ecological plasticity, has spread throughout the world. Although long considered only a secondary vector of viruses, the potential of its vector capacity may constitute a threat to public health. Based on the idea that an improved understanding of the olfactory system of mosquitoes may assist in the development of control methods that interfere with their behavior, we have undertaken a study aimed at characterizing the A. albopictus Odorant Receptors. Here we report the identification, cloning and functional characterization of the AalOR2 ortholog, that represents the first candidate member of the odorant receptor (OR) family of proteins from A. albopictus. AalOR2 is expressed in the larval heads and antennae of adults. Our data indicate that A. albopictus OR2 (AalOR2) shares a high degree of identity with other mosquito OR2 orthologs characterized to date, confirming that OR2 is one of the most conserved mosquito ORs. Our data indicate that AalOR2 is narrowly tuned to indole, and inhibited by (-)-menthone. In agreement with this results, these two compounds elicit two opposite effects on the olfactory-based behavior of A. albopictus larvae, as determined through a larval behavioral assay. In summary, this work has led to the cloning and de-orphaning of the first Odorant Receptor in the tiger mosquito A. albopictus. In future control strategies this receptor may be used as a potential molecular target.     

Associations between dopamine D4 receptor gene variation with both infidelity and sexual promiscuity

Background

Human sexual behavior is highly variable both within and between populations. While sex-related characteristics and sexual behavior are central to evolutionary theory (sexual selection), little is known about the genetic bases of individual variation in sexual behavior. The variable number tandem repeats (VNTR) polymorphism in exon III of the human dopamine D4 receptor gene (DRD4) has been correlated with an array of behavioral phenotypes and may be predicatively responsible for variation in motivating some sexual behaviors, particularly promiscuity and infidelity.

Methodology/Principal Findings

We administered an anonymous survey on personal history of sexual behavior and intimate relationships to 181 young adults. We also collected buccal wash samples and genotyped the DRD4 VNTR. Here we show that individuals with at least one 7-repeat allele (7R+) report a greater categorical rate of promiscuous sexual behavior (i.e., having ever had a “one-night stand”) and report a more than 50% increase in instances of sexual infidelity.

Conclusions/Significance

DRD4 VNTR genotype varies considerably within and among populations and has been subject to relatively recent, local selective pressures. Individual differences in sexual behavior are likely partially mediated by individual genetic variation in genes coding for motivation and reward in the brain. Conceptualizing these findings in terms of r/K selection theory suggests a mechanism for selective pressure for and against the 7R+ genotype that may explain the considerable global allelic variation for this polymorphism.     

Genetic variation in the vitamin D receptor gene and vitamin D serum levels in Egyptian women with polycystic ovary syndrome

Obesity, insulin resistance, and hyperandrogenism are considered crucial parameters of polycystic ovary syndrome (PCOS) which might be related to vitamin D metabolism. The aim of this study was to investigate the associations between polymorphisms (TaqI and ApaI) in the vitamin D receptor gene (VDR) and PCOS among Egyptian women. We aimed also to elucidate the impact of these polymorphisms on vitamin D level, hormonal and metabolic parameters of PCOS. One hundred and fifty Egyptian women with PCOS and 150 unrelated controls were enrolled in this study. Polymorphisms of VDR Taq-I T/C (rs731236) and Apa-I A/C (rs7975232) gene were genotyped using polymerase chain reaction restriction fragment length polymorphism (PCR–RFLP). Serum 25 hydroxy vitamin D [25(OH) D] levels were measured by high-performance liquid chromatography. PCOS women had significantly lower levels of 25(OH) D compared to healthy women. Our results revealed that Taq-I CC genotype and C allele were associated with increased risk of PCOS, while the Apa-I polymorphism was not. Haplotype Taq-I C/ Apa-I C was associated with a higher PCOS risk more than controls. Moreover, there was a significant decrease of 25(OH) D levels in carriers of haplotype Taq-I C/ Apa-I C (with variant alleles) compared to the non-carriers. Results showed also that there was an obesity- VDR Taq-I genotypes interactions. These results suggested that, VDR Taq-I gene polymorphism is associated with increased risk of PCOS in Egyptian women.     

Identification and functional characterization of an odorant receptor in pea aphid,Acyrthosiphon pisum

The sensitive olfactory system is necessary for survival of insects.Odorant receptors (ORs)are located on the dendrites of olfactory receptor neurons and play a critical role in odor detection.Insect ORs are functionally analyzed via heterologous expression in a Xenopus oocyte system using a two-electrode voltage-clamp (TEVC)electrophysiological recording.Here,we have identified a novel OR in the pea aphid,Acyrthosiphon pisum,then we cloned and named it ApisOR4.We analyzed the ApisOR4 tissue expression patterns and found expression only in antennae tissues.Further functional analysis using TEVC revealed that ApisOR4 is broadly tuned to eight volatiles,which elicit electrophysiological response in pea aphid antennae.This study provides an initial functional analysis of aphid ORs and identifies candidate volatiles to be used in developing new strategies for aphid control.     

Molecular similarities in the ligand binding pockets of an odorant receptor and the metabotropic glutamate receptors

The 5.24 odorant receptor is an amino acid sensing receptor that is expressed in the olfactory epithelium of fish. The 5.24 receptor is a G-protein-coupled receptor that shares amino acid sequence identity to mammalian pheromone receptors, the calcium-sensing receptor, the T1R taste receptors, and the metabotropic glutamate receptors (mGluRs). It is most potently activated by the basic amino acids L-lysine and L-arginine. In this study we generated a homology model of the ligand binding domain of the 5.24 receptor based on the crystal structure of mGluR1 and examined the proposed lysine binding pocket using site-directed mutagenesis. Mutants of truncated glycosylated versions of the receptor containing only the extracellular domain were analyzed in a radioligand binding assay, whereas the analogous full-length membrane-bound mutants were studied using a fluorescence-based functional assay. In silico analysis predicted that aspartate 388 interacts with the terminal amino group on the side chain of the docked lysine molecule. This prediction was supported by experimental observations demonstrating that mutation of this residue caused a 26-fold reduction in the affinity for L-lysine but virtually no change in the affinity for the polar amino acid L-glutamine. In addition, mutations in four highly conserved residues (threonine 175, tyrosine 223, and aspartates 195 and 309) predicted to establish interactions with the alpha amino group of the bound lysine ligand greatly reduced or eliminated binding and receptor activation. These results define the essential features of amino acid selectivity within the 5.24 receptor binding pocket and highlight an evolutionarily conserved motif required for ligand recognition in amino acid activated receptors in the G-protein-coupled receptor superfamily.     

Genetic variation at the ApoB 3'HVR, D2S44, and D7S21 loci in the Ewondo Ethnic Group of Cameroon.

A sample of the Ewondo population (a Bantu-speaking group of Southern Cameroon) was analyzed for the polymorphism at three tandem repeated DNA loci (ApoB 3' HVR, D2S44, and D7S21). We observed a greater number of ApoB 3' HVR alleles (17) and a significantly higher estimated heterozygosity (.879 +/- .011) than in previously surveyed populations, with the exception of U.S. Blacks. The higher genetic variability of Ewondo and U.S. Blacks was also shown by the ApoB 3' HVR allele-frequency spectra. A method for measuring population distances, based on cumulative fragment-size distribution, is described. Interpopulation comparisons for ApoB 3' HVR were carried out by this method and were compared with those obtained by a genetic distance measurement. The two sets of results showed a consistent pattern of population differentiation: the Ewondos and the U.S. Blacks clustered together and were well apart from both a Caucasian cluster (Swedes, U.S. Whites, Italians, and Germans) and other well-defined populations (Sikhs of India and Pehuence Indians of Chile). Profile distances were then computed from D2S44 and D7S21 bined data. This analysis indicated a genetic affinity between Ewondos, U.S. Blacks, and Afro-Caribbean Blacks and outlined the genetic diversity between Ewondos, Caucasians, and Asian Indians.     

Melanocortin receptor 4 is induced in nerve-injured motor and sensory neurons of mouse

We previously identified melanocortin receptor 4 (MC4R) in a search for genes associated with hypoglossal nerve regeneration. As melanocortins promote nerve regeneration after axonal injury, we investigated whether MC4R functions as a key receptor for peripheral nerve regeneration. In situ hybridization revealed that MC4R mRNA is induced in mouse hypoglossal motor neurons after axonal injury, whereas mRNAs for MC1R, MC2R, MC3R, and MC5R are not expressed either before or after nerve injury. This result was confirmed by RT-PCR. The level of MC4R mRNA expression increased significantly from day 3 after axotomy, reached a peak on day 5, and decreased to the control level on day 14. Similar induction of MC4R was observed in axotomized mouse dorsal root ganglia (DRGs). MC4R mRNA expression was induced exclusively among the MCR family in the L4-6 DRG after sciatic nerve injury. We further examined whether alpha-melanocortin stimulating hormone (alpha-MSH) promotes neurite elongation via MC4R. In mouse DRG neuron culture, alpha-MSH significantly promoted neurite outgrowth at a concentration of 10(-8) mol/L. This neurite-elongation effect was entirely inhibited by the addition of a selective MC4R blocker, JKC-363. Therefore, it is concluded that alpha-MSH could stimulate neurite elongation via MC4R in DRG neurons. The present results suggest that induction of MC4R is crucial for motor and sensory neurons to regenerate after axonal injury.     

绿盲蝽气味受体基因AlucOR40的克隆及功能研究

【目的】从绿盲蝽Apolygus lucorum触角中克隆气味受体基因Aluc OR40,研究该气味受体基因在绿盲蝽不同组织中的表达水平,然后对该基因的功能进行研究,为理解绿盲蝽的嗅觉识别机制提供理论基础。【方法】在绿盲蝽成虫触角转录组测序与分析的基础上,通过PCR技术克隆得到气味受体基因Aluc OR40的全长序列。用半定量RT-PCR研究该基因在雌雄虫不同组织中的表达水平。然后通过爪蟾卵母细胞体外表达该基因,并结合双电极电压钳检测了该气味受体对60种气味分子包括绿盲蝽性信息素组分和植物挥发物的反应。然后,利用触角电位技术测定羽化后3 d的绿盲蝽成虫对Aluc OR40的气味配体的触角电位反应。【结果】克隆了Aluc OR40(Gen Bank登录号:KU886190)。Aluc OR40编码398个氨基酸,蛋白具有7个跨膜结构域,N末端位于胞内,C端位于细胞外,符合昆虫气味受体的典型特征。半定量RT-PCR的结果显示,Aluc OR40在触角中特异表达,且在雄虫触角中的表达水平高于雌虫。双电极电压钳记录结果显示,Aluc OR40只对反-2-己烯醇一种气味分子具有特异反应。EAG实验结果表明,羽化后3 d的绿盲蝽雌雄虫都对反-2-己烯醇有反应,且雄虫触角的EAG反应显著高于雌虫。【结论】结果提示Aluc OR40参与了绿盲蝽对反-2-己烯醇的识别过程,推测其在绿盲蝽交配过程中雄虫对雌虫的识别中发挥作用。     

Amino acids involved in conformational dynamics and G protein coupling of an odorant receptor: targeting gain-of-function mutation

Thousands of different odorants are recognized and discriminated by odorant receptors (ORs) in the guanine nucleotide-binding protein (G protein)-coupled seven-transmembrane receptor family. Odorant-bound ORs stimulate Gs-type G proteins, Galphaolf, which in turn activates cAMP-mediated signaling pathway in olfactory sensory neurons. To better understand the molecular basis for OR activation and G protein coupling, we analyzed the effects of a series of site-directed mutations of mouse ORs, on function. Mutations of conserved amino acid residues in an intracellular loop or the C-terminus resulted in loss of activity without impairing ligand-binding activity, indicating that these residues are involved in Galphas/olf coupling. Moreover, mutation of the serine in KAFSTC, the OR-specific sequence motif, resulted in a dramatic increase in odorant responsiveness, suggesting that the motif is involved in a conformational change of the receptor that regulates G protein coupling efficiency. Our results provide insights into how ORs switch from an inactive to an active state, as well as where and how activated ORs interact with G proteins.     

Synthesis and estrogen receptor binding affinities of 7-hydroxy-3-(4-hydroxyphenyl)-4H-1-benzopyran-4-ones containing a basic side chain

Two isoflavones containing a sulfur or oxygen hinge with an amine-bearing side chain have been designed and synthesized as potential selective estrogen receptor modulators. The target compounds exhibited low affinities for estrogen receptors (ERs), and binding affinity data indicate that oxygen hinge is more favorable than sulfur for binding. These compounds also displayed selectivity for ERalpha over ERbeta.