Identification of genetic innovations in vertebrates

• Main Author: Tuberosa, Joël
• Other Authors: Rodriguez, Ivan
• Format: Doctoral or Postdoctoral Thesis
• Language: English
• Published: Université de Genève 2020
• Subjects: info:eu-repo/classification/ddc/590; Rodriguez; toothed whales
• Online Access: https://archive-ouverte.unige.ch/unige:156655; https://doi.org/10.13097/archive-ouverte/unige:156655

The evolution of life forms is marked by the emergence of new genes and the transformation of old ones. Thus, genomes possess a reservoir of sequences escaping selective pressures and of fortuitous transcriptional activity, with a potential to encode entirely new functions. In parallel, recycling of genetic material with preexisting function takes place, a process likely more common than the production of genuine original genes. This reconditioning is at the origin of incremental novelties, but also at the origin of spectacular innovations, such as the specialization of oxygen transport or the emergence of chromatic vision. During my thesis, I explored different instances of genetic innovations in neurons. My studies focused, on the one hand, on the evolution of mammalian olfactory chemoreceptors, which constitute a remarkable model to study evolution by gene duplication and acquisition of new expression patterns. Indeed, the gene families encoding for these receptors are characterized by an exceptional rate of gene duplication, diversification and death. On the other hand, my research was initiated by the discovery of an entirely new gene that emerged during vertebrate evolution. The genes encoding for odorant receptors (ORs), one of the olfactory receptor superfamilies, constitute the largest gene family in the mammalian genome. With hundreds of representatives, ORs represent a highly diversified panoply of chemoreceptors that may attend non-olfactory functions when expressed outside of the olfactory system. Ectopic transcription of OR genes was indeed reported multiple times, but the adaptive value of this putative pleiotropy remains unclear. In the first part of this work, we used a comparative genomic approach with the aim to identify ectopic ORs with a non-olfactory role and of evolutionary importance in mammals. For this, we took advantage of the toothed whales evolution, a group which has lost its olfactory system 35 millions of years ago. We found three intact OR coding sequences that were present in the genome of all mammalian species, including in cetaceans, pointing to a putative non-olfactory function of these genes. Supporting our results, several studies reported the transcription of two of these particular genes in non-olfactory tissues, with evidences suggesting a role in host-microbiota interactions. The role played by the third candidate remains a mystery. The Rodriguez laboratory has previously shown that during the evolution of rodents, the vomeronasal organ, which is dedicated to the perception of pheromones, has acquired a family of chemoreceptors, the formyl peptide receptors (FPRs), originally expressed by immune cells(Dietschi et al., 2017). In the second part of this work, we investigated the history of genomic accidents that led to this innovation, by combining synteny analyses and phylogenetic comparisons. In this study, we identified two independent events of exon shuffling, occurring millions of years apart, and leading to two instances of vomeronasal FPR acquisition. During each of these events, the duplicated coding exon of an immune FPR landed in front of a vomeronasal receptor gene promoter, hijacking its regulatory pattern. Finally, we show that under immune stress, one of these neofunctionalized vomeronasal FPRs can also be expressed in immune cells via the production of an intergenic transcript using an immune FPR promoter. Our findings demonstrate how the hijack of promoters by exon shuffling can lead to the acquisition of novel transcriptional regulations and phenotypical innovations, and how this novel tissue specificity can revert to its ancient pattern. Another family of vomeronasal organ chemoreceptors, present in most mammalian species, is encoded by the type 1 vomeronasal receptor (V1R) genes. V1R genes, like ORs, are clustered in the mouse genome. We hypothesized that this particular organization reflects a dependence to local regulatory elements, preventing the conservation of genes duplicated outside of clusters. In the third part of this thesis, we evaluated the potential evolutionary constraints that may have prevented the dispersion of functional V1R genes during mammalian evolution. To that end, we mapped the intact coding sequences and pseudogenes of all V1Rs in the genomes of representative mammalian species. We found that V1R genes – and the rodent vomeronasal FPR genes – are consistently organized in clusters, despite an exceptionally high rate of gene duplication. A few interspersed V1R genes were also found, but most of them were pseudogenes. To evaluate the role of putative local regulatory elements in the transcription of a functional vomeronasal receptor gene, we studied the expression of a vomeronasal FPR transgene when placed outside of its endogenous cluster and of other vomeronasal clusters. Using this model of long-range duplication, we observed a punctate and tissue-specific expression of the transgene in immature neurons, but that was not sustained. Hence, tissue specificity does not require the proximity of a vomeronasal gene cluster, whereas transcriptional stability seems to depend on the latter. Taken together, our data support the existence of local regulatory elements that have restrained chemoreceptor gene dispersion during evolution. In the last part of this work, we studied the emergence and the role of a previously unknown vertebrate gene, that we discovered. In mice, this gene, that we termed ghost, was found to be almost exclusively transcribed in the brain, in the claustrum and in the thalamic reticular nucleus (TRN). These two brain regions that are involved in attention allocation and sleep regulation, have homologous structures in all amniotes, if not all vertebrates. In analyzing more than 300 vertebrate genomes, we found that ghost originated in an ancestor of bony vertebrates, approximately 450 millions years ago. Since then, it has remained highly conserved and almost always present in a single copy in genomes. The product encoded by ghost has no homology with any other known proteins. It is a single-pass transmembrane protein that we localized in the perinuclear region of neurons. We generated ghost knockout mice, that grow and breed normally, but show specific behaviors that point towards an alteration of cognitive functions and of sleep regulation. Our findings retrace the emergence and conservation of a peculiar gene that has certainly provided the bony vertebrate with a novel function that they could not afford to lose. Our data point towards a neuronal function in mammals, which constitutes a step in understanding the evolutionary history of ghost as well as that of the claustrum and the TRN.