Genetic Variation Across the Human Olfactory Receptor Repertoire Alters Odor Perception
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Genetic variation across the human olfactory receptor repertoire alters odor perception C. Trimmera,1, A. Kellerb, N. R. Murphya, L. L. Snydera, J. R. Willerc, M. H. Nagaid,e, N. Katsanisc, L. B. Vosshallb,f,g, H. Matsunamid,h, and J. D. Mainlanda,i aMonell Chemical Senses Center, Philadelphia, PA 19104; bLaboratory of Neurogenetics and Behavior, The Rockefeller University, New York, NY 10065; cCenter for Human Disease Modeling, Duke University Medical Center, Durham, NC 27710; dDepartment of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, NC 27710; eDepartment of Biochemistry, University of Sao Paulo, Sao Paulo 03178-200, Brazil; fHoward Hughes Medical Institute, The Rockefeller University, New York, NY 10065; gKavli Neural Systems Institute, The Rockefeller University, New York, NY 10065; hDepartment of Neurobiology and Duke Institute for Brain Sciences, Duke University, Durham, NC 27710; and iDepartment of Neuroscience, University of Pennsylvania School of Medicine, Philadelphia, PA 19104 Edited by John G. Hildebrand, University of Arizona, Tucson, AZ, and approved September 12, 2018 (received for review March 15, 2018) Humans use a family of more than 400 olfactory receptors (ORs) by an odorant has been proposed to encode both intensity and to detect odors, but there is currently no model that can predict pleasantness (9, 10). Alternatively, the large set of ORs may olfactory perception from receptor activity patterns. Genetic varia- redundantly encode odorant representations, such that loss of tion in human ORs is abundant and alters receptor function, allowing function of a single OR only rarely has perceptual consequences. us to examine the relationship between receptor function and Recent work suggests that functional changes in a single receptor perception. We sequenced the OR repertoire in 332 individuals and can have significant perceptual consequences, but data linking examined how genetic variation affected 276 olfactory phenotypes, perceptual and genetic variation exist for only five ORs (11–15). including the perceived intensity and pleasantness of 68 odorants at We extended these studies to the entire OR repertoire, under the two concentrations, detection thresholds of three odorants, and premise that studies of natural perceptual variation will improve our general olfactory acuity. Genetic variation in a single OR was fre- understanding of the normal translation of OR activity to odor per- quently associated with changes in odorant perception, and we ception. Here we identified associations between genetic variation in validated 10 cases in which in vitro OR function correlated with 418 OR genes and 276 olfactory phenotypes, used cell-based func- GENETICS in vivo odorant perception using a functional assay. In 8 of these 10 tional assays to investigate the mechanisms underlying the associa- cases, reduced receptor function was associated with reduced inten- tions, and examined the contributions of single OR genotype, genetic sity perception. In addition, we used participant genotypes to quan- ancestry, age, and gender to variations in olfactory perception. tify genetic ancestry and found that, in combination with single OR Results genotype, age, and gender, we can explain between 10% and 20% of the perceptual variation in 15 olfactory phenotypes, highlighting We carried out high-throughput sequencing of the entire OR sub- the importance of single OR genotype, ancestry, and demographic genome in a cohort of 332 participants who had been previously factors in the variation of olfactory perception. phenotyped for their sense of smell. These data include ratings of the perceived intensity and pleasantness of 68 odorants at two concen- trations (SI Appendix,TableS1), detection thresholds of three olfaction | genetic variation | human genome | odor intensity | ancestry odorants, and overall olfactory acuity (16). Participants rated each stimulus twice, and the median within-subject test-retest correlation nderstanding how the olfactory system detects odorants and was 0.63 for intensity ratings and 0.57 for pleasantness ratings. Utranslates their features into perceptual information is one of the fundamental questions in olfaction. Although early color Significance vision researchers were unable to directly observe receptor re- sponses, perceptual deficits caused by genetic variation (i.e., color A persistent mystery in olfaction is how the combinatorial acti- blindness) helped show that color vision is mediated by three re- vation of a family of 400 olfactory receptors (ORs) encodes odor ceptors responding to different wavelengths of light (1, 2). Guillot perception. We take advantage of the high frequency of natural (3), and then Amoore (4), extended this idea to olfaction and OR knockouts in the human genome to tackle a major bottle- proposed that cataloging specific anosmias (the inability to per- neck in the field—namely, how an odor is transduced into per- ceive a particular odorant) may provide similar clues linking genes ceptual characteristics. We demonstrate that loss of function of and perception. Early applications of this idea failed, presumably an individual OR correlates with changes in perceived intensity because olfaction relies on hundreds of receptors, and without and pleasantness. This study demonstrates how natural varia- direct observation of their responses, psychophysical tests could tion can provide important clues to the normal translation of OR not untangle the fundamental rules of odor coding. However, with activation to odor information and places a constraint on the the advent of next-generation genome sequencing to profile ol- amount of redundancy in the olfactory code. factory receptor (OR) genes and cell-based assays to identify li- gands for ORs, receptor variation can now be matched to Author contributions: A.K., L.B.V., H.M., and J.D.M. designed research; C.T., A.K., N.R.M., individuals and receptor responses can be directly observed. L.L.S., J.R.W., M.H.N., N.K., and J.D.M. performed research; C.T. analyzed data; and C.T. Humans have approximately 400 OR genes that are intact in wrote the paper. at least part of the population, but individuals have different Conflict of interest statement: J.D.M is on the scientific advisory board of Aromyx and repertoires of pseudogenes, copy number variations, and single receives compensation for these activities. nucleotide polymorphisms (SNPs) that can alter receptor re- This article is a PNAS Direct Submission. sponses (5–7). While nonfunctional genes are rare in the genome Published under the PNAS license. (on average approximately 100 heterozygous and 20 homozygous Data deposition: The data have been deposited in the database of Genotypes and Phe- pseudogenes in an individual), they are significantly enriched in notypes (dbGaP), www.ncbi.nlm.nih.gov/gap/ (accession no. phs001757.v1.p1). OR genes (8). This provides a useful set of “natural knockouts” 1To whom correspondence should be addressed. Email: [email protected]. to examine the role of a single OR in olfactory perception and This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10. distinguish between different hypotheses of how odor in- 1073/pnas.1804106115/-/DCSupplemental. formation is encoded. For example, the number of ORs activated Published online April 30, 2019. www.pnas.org/cgi/doi/10.1073/pnas.1804106115 PNAS | May 7, 2019 | vol. 116 | no. 19 | 9475–9480 Downloaded by guest on September 25, 2021 High-Throughput Sequencing of the OR Gene Family. We used Illu- difficult to discriminate between the causal mutation and nearby mina short-read DNA sequencing to analyze target regions SNPs in linkage disequilibrium with the causal mutation (18, 19). consisting of 418 ORs and 256 olfactory-related genes (∼800 kb), To illustrate this point, three different ORs in close proximity on obtaining a minimum of 15× coverage for 96% of the targeted chromosome 6 were significantly associated with the perceived bases and identifying 19,535 variants. We validated a subset of intensity of 2-ethylfenchol: OR11A1, OR12D2, and OR10C1 these by comparing them with variants identified from Sanger (Fig. 2A). The most highly associated SNP (Dataset S2), found in sequencing data for 10 ORs. Eight ORs had >95% concordance OR10C1, is in high linkage disequilibrium with SNPs in OR11A1 between the two sequencing methods, while concordance and OR12D2, with >60% correlation in the 1000 Genomes was <85% for OR10G4 and OR10G9. These ORs share >95% Project EUR superpopulation data (20, 21). sequence similarity (17), making it difficult to assign genetic To investigate these associations, we cloned all major haplo- variants to the correct genomic location. Therefore, alternate types in this locus and found that only OR11A1 haplotypes sequencing methods may prove necessary for ORs with high responded in the heterologous assay (Fig. 2B). OR haplotypes sequence similarity. However, high-throughput sequencing can with lower maximum responses in cell culture were associated be expected to accurately identify sequence variants for ORs in with lower perceived intensity ratings (Fig. 2C). These results in- which sequencing reads map with high confidence (90% of ORs dicate that of the three ORs found to significantly associate with have a median mapping confidence >99.9%). the perceived intensity of 2-ethylfenchol in our analysis, OR11A1 was the best candidate for a causal receptor at this locus. Genetic Variation in Single ORs Frequently Associates