High-throughput Analysis of Mammalian Olfactory Receptors: Measurement of Receptor Activation via Luciferase Activity

Trimmer, Casey; Snyder, Lindsey L.; Mainland, Joel D.

doi:10.3791/51640

Cited by 14 publications

(20 citation statements)

References 39 publications

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“…Luciferase Assay. The Promega Dual-Glo Luciferase Assay System was used to measure in vitro OR activity as described previously (39,40). Data were analyzed with GraphPad Prism 6.…”

Section: Methodsmentioning

confidence: 99%

Genetic variation across the human olfactory receptor repertoire alters odor perception

Trimmer

Keller

Murphy

et al. 2019

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

155

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Humans use a family of more than 400 olfactory receptors (ORs) to detect odors, but there is currently no model that can predict olfactory perception from receptor activity patterns. Genetic variation in human ORs is abundant and alters receptor function, allowing us to examine the relationship between receptor function and perception. We sequenced the OR repertoire in 332 individuals and examined how genetic variation affected 276 olfactory phenotypes, including the perceived intensity and pleasantness of 68 odorants at two concentrations, detection thresholds of three odorants, and general olfactory acuity. Genetic variation in a single OR was frequently associated with changes in odorant perception, and we validated 10 cases in which in vitro OR function correlated with in vivo odorant perception using a functional assay. In 8 of these 10 cases, reduced receptor function was associated with reduced intensity perception. In addition, we used participant genotypes to quantify genetic ancestry and found that, in combination with single OR genotype, age, and gender, we can explain between 10% and 20% of the perceptual variation in 15 olfactory phenotypes, highlighting the importance of single OR genotype, ancestry, and demographic factors in the variation of olfactory perception. olfaction | genetic variation | human genome | odor intensity | ancestry U nderstanding how the olfactory system detects odorants and translates their features into perceptual information is one of the fundamental questions in olfaction. Although early color vision researchers were unable to directly observe receptor responses, perceptual deficits caused by genetic variation (i.e., color blindness) helped show that color vision is mediated by three receptors responding to different wavelengths of light (1, 2). Guillot (3), and then Amoore (4), extended this idea to olfaction and proposed that cataloging specific anosmias (the inability to perceive a particular odorant) may provide similar clues linking genes and perception. Early applications of this idea failed, presumably because olfaction relies on hundreds of receptors, and without direct observation of their responses, psychophysical tests could not untangle the fundamental rules of odor coding. However, with the advent of next-generation genome sequencing to profile olfactory receptor (OR) genes and cell-based assays to identify ligands for ORs, receptor variation can now be matched to individuals and receptor responses can be directly observed.Humans have approximately 400 OR genes that are intact in at least part of the population, but individuals have different repertoires of pseudogenes, copy number variations, and single nucleotide polymorphisms (SNPs) that can alter receptor responses (5-7). While nonfunctional genes are rare in the genome (on average approximately 100 heterozygous and 20 homozygous pseudogenes in an individual), they are significantly enriched in OR genes (8). This provides a useful set of "natural knockouts" to examine the role of a single OR in olfactory pe...

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“…Luciferase Assay. The Promega Dual-Glo Luciferase Assay System was used to measure in vitro OR activity as described previously (39,40). Data were analyzed with GraphPad Prism 6.…”

Section: Methodsmentioning

confidence: 99%

Genetic variation across the human olfactory receptor repertoire alters odor perception

Trimmer

Keller

Murphy

et al. 2019

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

155

View full text Add to dashboard Cite

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“…The OSN express olfactory receptors, which recognize specific odours. Humans express up to 400 different olfactory receptors, while there are even up to 1200 different receptors in rodents [ 31 ]. The one-neuron-one-receptor rule indicates that one OSN is exclusively expressing one olfactory receptor [ 31 , 32 ].…”

Section: The Nasal Epitheliamentioning

confidence: 99%

“…Humans express up to 400 different olfactory receptors, while there are even up to 1200 different receptors in rodents [ 31 ]. The one-neuron-one-receptor rule indicates that one OSN is exclusively expressing one olfactory receptor [ 31 , 32 ]. The axons of OSN which express the same olfactory receptor converge to form neuronal bundles projecting to the olfactory bulb.…”

Section: The Nasal Epitheliamentioning

confidence: 99%

Tailoring Formulations for Intranasal Nose-to-Brain Delivery: A Review on Architecture, Physico-Chemical Characteristics and Mucociliary Clearance of the Nasal Olfactory Mucosa

2018

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The blood-brain barrier and the blood-cerebrospinal fluid barrier are major obstacles in central nervous system (CNS) drug delivery, since they block most molecules from entering the brain. Alternative drug delivery routes like intraparenchymal or intrathecal are invasive methods with a remaining risk of infections. In contrast, nose-to-brain delivery is a minimally invasive drug administration pathway, which bypasses the blood-brain barrier as the drug is directed from the nasal cavity to the brain. In particular, the skull base located at the roof of the nasal cavity is in close vicinity to the CNS. This area is covered with olfactory mucosa. To design and tailor suitable formulations for nose-to-brain drug delivery, the architecture, structure and physico-chemical characteristics of the mucosa are important criteria. Hence, here we review the state-of-the-art knowledge about the characteristics of the nasal and, in particular, the olfactory mucosa needed for a rational design of intranasal formulations and dosage forms. Also, the information is suitable for the development of systemic or local intranasal drug delivery as well as for intranasal vaccinations.

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“…9 Fewer than 6% of human ORs have been matched to known ligands. 10,11 The expression and biological function of ectopic ORs in the skin are poorly characterized. Moreover, their role in patients with inflammatory skin diseases, such as AD and psoriasis, are unknown.…”

mentioning

confidence: 99%

Expression and function of the ectopic olfactory receptor OR10G7 in patients with atopic dermatitis

Tham

Dyjack

Kim

et al. 2019

Journal of Allergy and Clinical Immunology

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Background: Ectopic olfactory receptors (ORs) are found in the skin, but their expression and biological function in normal skin and skin form patients with atopic dermatitis (AD) are unknown. Objectives: We sought to characterize the expression of ORs in the skin and assess OR-mediated biological responses of primary human keratinocytes in the presence of odorant ligands. Methods: OR expression was examined by using wholetranscriptome sequencing of skin tape strips collected from patients with AD and healthy control (HC) subjects. OR10G7 and filaggrin 1 (FLG-1) expression was analyzed by using RT-PCR and immunostaining in skin biopsy specimens and primary human keratinocytes from patients with AD and HC subjects. ATP and cyclic AMP production by control and OR10G7 small interfering RNA-transfected keratinocytes in response to odorant stimulation with acetophenone and eugenol was assessed. Results: A total of 381 OR gene transcripts were detected in the skin samples, with the greatest OR expression detected in the skin tape strips corresponding to the upper granular layer of the skin. OR10G7 expression was significantly increased in skin biopsy specimens from patients with AD compared with those from HC subjects (P 5 .01) and inversely correlated with FLG-1 expression (P 5 .009). OR10G7 expression was greatest in undifferentiated keratinocytes from patients with AD and was downregulated with progressive differentiation. Primary human keratinocytes produced ATP, an essential neurotransmitter in sensory pathways, in response to acetophenone and eugenol, odorants previously identified as potential ligands for this receptor. This response was abolished in OR10G7 small interfering RNA-transfected keratinocytes. Conclusions: OR10G7 is expressed at significantly greater levels in undifferentiated keratinocytes from patients with AD compared with HC subjects. OR10G7 is likely involved in transmission of skin-induced chemosensory responses to odorant stimulation, which might modulate differential nociceptive responses in AD skin.

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High-throughput Analysis of Mammalian Olfactory Receptors: Measurement of Receptor Activation via Luciferase Activity

Cited by 14 publications

References 39 publications

Genetic variation across the human olfactory receptor repertoire alters odor perception

Genetic variation across the human olfactory receptor repertoire alters odor perception

Tailoring Formulations for Intranasal Nose-to-Brain Delivery: A Review on Architecture, Physico-Chemical Characteristics and Mucociliary Clearance of the Nasal Olfactory Mucosa

Expression and function of the ectopic olfactory receptor OR10G7 in patients with atopic dermatitis

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