Dataset Information

Olfactory receptor pseudo-pseudogenes.

ABSTRACT: Pseudogenes are generally considered to be non-functional DNA sequences that arise through nonsense or frame-shift mutations of protein-coding genes. Although certain pseudogene-derived RNAs have regulatory roles, and some pseudogene fragments are translated, no clear functions for pseudogene-derived proteins are known. Olfactory receptor families contain many pseudogenes, which reflect low selection pressures on loci no longer relevant to the fitness of a species. Here we report the characterization of a pseudogene in the chemosensory variant ionotropic glutamate receptor repertoire of Drosophila sechellia, an insect endemic to the Seychelles that feeds almost exclusively on the ripe fruit of Morinda citrifolia. This locus, D. sechellia Ir75a, bears a premature termination codon (PTC) that appears to be fixed in the population. However, D. sechellia Ir75a encodes a functional receptor, owing to efficient translational read-through of the PTC. Read-through is detected only in neurons and is independent of the type of termination codon, but depends on the sequence downstream of the PTC. Furthermore, although the intact Drosophila melanogaster Ir75a orthologue detects acetic acid-a chemical cue important for locating fermenting food found only at trace levels in Morinda fruit-D. sechellia Ir75a has evolved distinct odour-tuning properties through amino-acid changes in its ligand-binding domain. We identify functional PTC-containing loci within different olfactory receptor repertoires and species, suggesting that such 'pseudo-pseudogenes' could represent a widespread phenomenon.

SUBMITTER: Prieto-Godino LL

PROVIDER: S-EPMC5164928 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

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Olfactory receptor pseudo-pseudogenes.

Prieto-Godino Lucia L LL Rytz Raphael R Bargeton Benoîte B Abuin Liliane L Arguello J Roman JR Peraro Matteo Dal MD Benton Richard R

Nature 20161024 7627

Pseudogenes are generally considered to be non-functional DNA sequences that arise through nonsense or frame-shift mutations of protein-coding genes. Although certain pseudogene-derived RNAs have regulatory roles, and some pseudogene fragments are translated, no clear functions for pseudogene-derived proteins are known. Olfactory receptor families contain many pseudogenes, which reflect low selection pressures on loci no longer relevant to the fitness of a species. Here we report the characteriz ...[more]

PMID: 27776356

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Project description:BackgroundOlfactory receptors (ORs), the largest mammalian gene superfamily (900-1400 genes), has >50% pseudogenes in humans. While most of these inactive genes are identified via coding frame (nonsense) disruptions, seemingly intact genes may also be inactive due to other deleterious (missense) mutations. An ultimate assessment of the actual size of the functional human OR repertoire thus requires an accurate distinction between genes and pseudogenes.ResultsTo characterize inactive ORs with intact open reading frame, we have developed a probabilistic Classifier for Olfactory Receptor Pseudogenes (CORP). This algorithm is based on deviations from a functionally crucial consensus, constituting sixty highly conserved positions identified by a comparison of two evolutionarily-constrained OR repertoires (mouse and dog) with a small pseudogene fraction. We used a logistic regression analysis to assign appropriate coefficients to the conserved position and thus achieving maximal separation between active and inactive ORs. Consequently, the algorithms identified only 5% of the mouse functional ORs as pseudogenes, setting an upper limit of 0.05 to the false positive detection. Finally we used this algorithm to classify the 384 purportedly intact human OR genes. Of these, 135 were predicted as likely encoding non-functional proteins, and 38 were segregating between active and inactive forms due to missense polymorphisms.ConclusionWe demonstrated that the CORP algorithm is capable to distinguish between functional and non-functional OR genes with high precision even when the encoded protein would differ by a single amino acid. Using the CORP algorithm, we predict that approximately 70% of human OR genes are likely non-functional pseudogenes, a much higher number than hitherto suspected. The method we present may be employed for better annotation of inactive members in other gene families as well. CORP algorithm is available at: http://bioportal.weizmann.ac.il/HORDE/CORP/

Project description:Olfactory sensory neurons choose to express a single odorant receptor (OR) from a large gene repertoire and extend axons to reproducible, OR-specific locations within the olfactory bulb. This developmental process produces a topographically organized map of odorant experience in the brain. The axon guidance mechanisms that generate this pattern of connectivity, as well as those that coordinate OR choice and axonal guidance receptor expression, are incompletely understood. We applied the powerful approach of single-cell RNA-seq on newly born olfactory sensory neurons (OSNs) in young zebrafish larvae to address these issues. Expression profiles were generated for 56 individual Olfactory Marker Protein (OMP) positive sensory neurons by single-cell (SC) RNA-seq. We show that just as in mouse OSNs, mature zebrafish OSNs typically express a single predominant OR transcript. Our previous work suggests that OSN targeting is related to the OR clade from which a sensory neuron chooses to express its odorant receptor. We categorized each of the mature cells based on the clade of their predominantly expressed OR. Transcripts expressed at higher levels in each of three clade-related categories were identified using Penalized Linear Discriminant Analysis (PLDA). A genome-wide approach was used to identify membrane-associated proteins that are most likely to have guidance-related activity. We found that OSNs that choose to express an OR from a particular clade also express specific subsets of potential axon guidance genes and transcription factors. We validated our identification of candidate axon guidance genes for one clade of OSNs using bulk RNA-seq from a subset of transgene-labeled neurons that project to a single protoglomerulus. The differential expression patterns of selected candidate guidance genes were confirmed using fluorescent in situ hybridization. Most importantly, we observed axonal mistargeting in knockouts of three candidate axonal guidance genes identified in this analysis: nrp1a, nrp1b, and robo2. In each case, targeting errors were detected in the subset of axons that normally express these transcripts at high levels, and not in the axons that express them at low levels. Our findings demonstrate that specific, functional, axonal guidance related genes are expressed in subsets of OSNs that that can be categorized by their patterns of OR expression.

Project description:Olfactory perception is mediated by a large array of olfactory receptor genes. The human genome contains 851 olfactory receptor gene loci. More than 50% of the loci are annotated as nonfunctional due to frame-disrupting mutations. Furthermore haplotypic missense alleles can be nonfunctional resulting from substitution of key amino acids governing protein folding or interactions with signal transduction components. Beyond their role in odor recognition, functional olfactory receptors are also required for a proper targeting of olfactory neuron axons to their corresponding glomeruli in the olfactory bulb. Therefore, we anticipate that profiling of olfactory receptor gene expression in whole human olfactory mucosa and analysis in the human population of their expression should provide an opportunity to select the frequently expressed and potentially functional olfactory receptors in view of a systematic deorphanization. To address this issue, we designed a TaqMan Low Density Array (Applied Biosystems), containing probes for 356 predicted human olfactory receptor loci to investigate their expression in whole human olfactory mucosa tissues from 26 individuals (13 women, 13 men; aged from 39 to 81 years, with an average of 67±11 years for women and 63±12 years for men). Total RNA isolation, DNase treatment, RNA integrity evaluation and reverse transcription were performed for these 26 samples. Then 384 targeted genes (including endogenous control genes and reference genes specifically expressed in olfactory epithelium for normalization purpose) were analyzed using the same real-time reverse transcription PCR platform. On average, the expression of 273 human olfactory receptor genes was observed in the 26 selected whole human olfactory mucosa analyzed, of which 90 were expressed in all 26 individuals. Most of the olfactory receptors deorphanized to date on the basis of sensitivity to known odorant molecules, which are described in the literature, were found in the expressed olfactory receptors gene set.

Dataset Information

Olfactory receptor pseudo-pseudogenes.

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Olfactory receptor pseudo-pseudogenes.

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