Genome-wide identification, expression analysis of taars (trace amine-associated receptors) and response to cadaverine and putrescine in Mandarin fish (Siniperca chuatsi)

BackgroundAccumulating evidences suggest that trace amine-associated receptor 1 (TAAR1) is a new promising target for psychiatric disorders since TAAR1 was discovered in 2001[1, 2]. Stressful life events are a key environmental risk factor to develop major depressive disorder (MDD) [3]. Studies in humans and animals have demonstrated that depression and stress induced pathologies are implicated in the dentate gyrus (DG), one of the key regions for HPA axis glucocorticoid feedback [4]. TAAR1 exhibits widespread expression in the hippocampus and the data gained in transcriptomic data that indicate an important role of TAARs in the regulation of adult neurogenesis [5]. However, the role of TAAR1 in dentate gyrus (DG) mediating the negative outcomes induced by chronic stress remains unknown.Aims & ObjectivesIn this study, we aimed to investigate the role of TAAR1 involved in stress-induced structural and functional impairments of the DG.MethodsFirstly, chronic social defeat stress model (21 days) in mice was established. Via behavioral measurement and molecular biology techniques, hippocampus-related behaviors, adult neurogenesis, the expression levels of TAAR1 induced by chronic social defeat stress were investigated. Secondly, we explored whether selective knockout TAAR1 in the dentate gyrus (DG) can mimic the chronic stress induced impaired adult neurogenesis and cognitive function deficits. A Cre recombinase-responsive adeno-associated virus (AAV) was bilaterally stereotactically microinjected into the DG of TAAR1 CKO mice. Lastly, we investigated whether and how activating TAAR1 during stress exposure could block the negative outcomes of chronic stress exposure. RO5263397, a TAAR1 agonist, prior to each stress exposure were systematically administered.ResultsMale C57BL/6N mice subjected to chronic social defeat stress exhibited significant reduction in the TAAR1 mRNA level in the DG, impaired social approach behavior, social memory and spatial object recognition memory, and impaired proliferation and maturation of adult-born dentate granule cells.Furthermore, selective knockout TAAR1 in the DG can mimic the chronic stress induced hippocampus- dependent cognitive function deficits and damage of neurogenesis. Lastly, Systemic TAAR1 agonist RO5263397 administration during chronic stress exposure prevented stress-induced deficits of hippocampus-dependent cognitive ability. RO5263397 reversed the inhibitory effects on adult neurogenesis induced by chronic stress and alleviated chronic stress-evoked morphological changes of dental granule cells, including dendrite retraction, shrinkage of apical branches and spine loss.Discussion & ConclusionTaken together, our findings indicate TAAR1 is an important molecular mediating chronic stress-induced deficit of hippocampal morphology and function. The TAAR1 agonist RO5263397 has the potential role towards the treatment of patients with MDD-related cognitive impairments.ReferencesAlnefeesi, Y., et al., Trace amine-associated receptor 1 (TAAR1): Potential application in mood disorders: A systematic review. Neurosci Biobehav Rev, 2021. 131: p. 192-210.Halff, E.F., et al., Trace amine- associated receptor 1 (TAAR1) agonism as a new treatment strategy for schizophrenia and related disorders. Trends Neurosci, 2023. 46(1): p. 60-74.Kendler, K.S., L.M. Karkowski, and C.A. Prescott, Causal relationship between stressful life events and the onset of major depression. Am J Psychiatry, 1999. 156(6): p. 837-41.Gould, E. and P. Tanapat, Stress and hippocampal neurogenesis. Biol Psychiatry, 1999. 46(11): p. 1472-9.Katolikova, N.V., et al., Expression of Trace Amine-Associated Receptors in the Murine and Human Hippocampus Based on Public Transcriptomic Data. Cells, 2022. 11(11).

Trace amine-associated receptor 1 (TAAR1) is a G-protein coupled receptor (GPCR) found in monoaminergic neurons of the central nervous system. Amphetamine-like psychostimulants activate TAAR1, which couples to Gαs and Gα13 downstream signaling cascades, and the latter regulates the RhoA-mediated internalization of neurotransmitter transporters. TAAR1 stimulation is crucial to methamphetamine (MA) mechanism of action, and a common variant of TAAR1 is associated with increased MA craving in MA users. Although the impact of TAAR1 on behavior has been characterized, the molecular mechanisms underlying TAAR1 pharmacology are incompletely understood. TAAR1 is a unique GPCR as it resides inside the cell rather than at the cell surface, though the precise location of TAAR1 within the cell is unknown. Understanding the intracellular localization, orientation, and trafficking of TAAR1 will further inform the molecular mechanisms of TAAR1 signaling. However, available antibodies and fluorescent fusion proteins for imaging TAAR1 are inadequate for such studies. Therefore, we have developed a novel method for imaging human TAAR1 (hTAAR1) through both light and electron microscopy using the miniVIPER peptide labeling system for multiscale microscopy. The hTAAR1-miniVIPER system has been applied to in vitro systems and has been used in conjunction with light microscopy to explore subcellular localization of the recombinant receptor in Chinese hamster ovary cells. Further investigation with electron microscopy will be carried out to determine the orientation of hTAAR1 on internal membranes and will indicate how TAAR1 mobilizes spatially distinct signaling cascades within the cell. In order to study the receptor in native tissue, AAV constructs will be employed to express the hTAAR1-miniVIPER construct in the dopamine neurons of mice. Tissue from these animals will be imaged through correlative light and electron microscopy to determine the subcellular localization and orientation of TAAR1. Animals expressing hTAAR1-miniR will be subjected to acute and chronic drug treatments to determine how TAAR1 agonism affects receptor localization and trafficking. Together these studies will expand our current understanding of molecular mechanisms of TAAR1 signaling and will inform future work concerning the role TAAR1 plays in the modulation of neural circuity relevant to addiction and methamphetamine use disorder.

Increased dopamine transmission and adult neurogenesis in trace amine-associated receptor 5 (TAAR5) knockout mice

Triple-negative breast cancer (TNBC) is a highly aggressive cancer subtype with poor patient survival due to high rates of metastasis and a lack of targeted treatments. Recently, a positive correlation between Trace Amine-Associated Receptor 1 (TAAR1) expression and breast cancer (BC) survival was reported, suggesting a possible protective role of the receptor in this disease state. Further, the TAAR1 agonist T1AM has been shown to induce inhibition of cancer cell proliferation and growth, but T1AM-TAAR1 mechanisms are not yet understood in this context. This dissertation was designed to systematically assess TAAR1 signaling effects on a range of malignancy-related cellular functions in BC. A panel of molecularly and phenotypically diverse human BC cell lines was assessed to characterize TAAR1 expression and validate use in functional studies. My studies identified two cell lines exhibiting distinct patterns of TAAR1 localization: MDA-MB-231 and MCF-7 cells. These cells preferentially express TAAR1 on the cell surface and intracellularly, respectively. This dichotomy was chosen to assess whether availability of cell surface T1AM binding sites translated into functional differences in cancer cell functions. My studies revealed that congruent with TAAR1 expression patterns, MDA-MB-231 cells were more sensitive to T1AM-induced cAMP accumulation, cell-killing, and inhibition of the metastatic phenotype than MCF-7 cells. TAAR1 activation with T1AM significantly reduced the migratory capacity of TNBC cells in-vitro and drug preincubation enhanced these effects, suggesting there may be modulation of downstream gene targets to modulate metastatic capacity. Agonist induced morphological changes to MDA-MB-231 cells resembled those undergone during transition from metastatic mesenchymal to epithelial phenotypes and may explain the protective effect reported in the literature. MCF-7 cells were largely devoid of significant responses, supporting the hypothesis that TAAR1 activation and its subcellular localization is indeed driving these anti-cancer effects. Taken together, the data generated in this dissertation provide evidence for TAAR1 modulation of cellular functions critical to metastatic progression and identify a novel receptor target for TNBC.--Author's abstract

Trace amine-associated receptors (TAARs) are a group of G protein-coupled receptors that are expressed in the olfactory epithelium, central nervous system, and periphery. TAAR family generally consists of nine types of receptors (TAAR1-9), which can detect biogenic amines. During the last 5 years, the TAAR5 receptor became one of the most intriguing receptors in this subfamily. Recent studies revealed that TAAR5 is involved not only in sensing socially relevant odors but also in the regulation of dopamine and serotonin transmission, emotional regulation, and adult neurogenesis by providing significant input from the olfactory system to the limbic brain areas. Such results indicate that future antagonistic TAAR5-based therapies may have high pharmacological potential in the field of neuropsychiatric disorders. TAAR5 is known to be expressed in leucocytes as well. To evaluate potential hematological side effects of such future treatments we analyzed several hematological parameters in mice lacking TAAR5. In these mutants, we observed minor but significant changes in the osmotic fragility test of erythrocytes and hematocrit levels. At the same time, analysis of other parameters including complete blood count and reticulocyte levels showed no significant alterations in TAAR5 knockout mice. Thus, TAAR5 gene knockout leads to minor negative changes in the erythropoiesis or eryptosis processes, and further research in that field is needed. The impact of TAAR5 deficiency on other hematological parameters seems minimal. Such negative, albeit minor, effects of TAAR5 deficiency should be taken into account during future TAAR5-based therapy development.

Genomic organization and evolution of olfactory receptors and trace amine-associated receptors in channel catfish, Ictalurus punctatus

Trace amine-associated receptor 1 activation silences GSK3β signaling of TAAR1 and D2R heteromers

TAAR1 in dentate gyrus is involved in chronic stress-induced impairments in hippocampal plasticity and cognitive function

Trace amine-associated receptor 5 (TAAR5) is a G protein-coupled receptor that belongs to the TAARs family (TAAR1-TAAR9). TAAR5 is expressed in the olfactory epithelium and is responsible for sensing 3-methylamine (TMA). However, recent studies showed that TAAR5 is also expressed in the limbic brain regions and is involved in the regulation of emotional behaviour and adult neurogenesis, suggesting that TAAR5 antagonism may represent a novel therapeutic strategy for anxiety and depression. We used the AtomNet® model, the first deep learning neural network for structure-based drug discovery, to identify putative TAAR5 ligands and tested them in an in vitro BRET assay. We found two mTAAR5 antagonists with low to submicromolar activity that are able to inhibit the cAMP production induced by TMA. Moreover, these two compounds also inhibited the mTAAR5 downstream signalling, such as the phosphorylation of CREB and ERK. These two hits exhibit drug-like properties and could be used to further develop more potent TAAR5 ligands with putative anxiolytic and antidepressant activity.

Trace amine‐associated receptor 1 (TAAR1) is a G protein‐coupled receptor (GPCR) in brain regions that are critical to reward and habit formation. TAAR1 is activated by a variety of substituted phenethylamines (e.g. methamphetamine, MA), and is uniquely involved in the mechanism of action of amphetamine‐like psychostimulants. Pharmacological manipulation of TAAR1 alters the spontaneous firing rate of neurons in the ventral tegmental area, and psychiatric disorders characterized by dysregulated dopaminergic neurotransmission, including schizophrenia and substance use disorders, have been associated with altered trace amine levels or TAAR1 polymorphisms.Unlike many GPCRs, TAAR1 is an intracellular receptor; rather than localizing to the plasma membrane, TAAR1 is associated with unidentified membranous compartments within the cell. Previous efforts to identify the intracellular localization of TAAR1 have had limited success due to difficulty with heterologous expression systems and a lack of sufficiently specific antibodies, as well as inherent spatial resolution limits of light‐based microscopic techniques. In order to determine the precise subcellular localization of TAAR1, immunoelectron microscopy was performed in HEK293T cells stably expressing human TAAR1 with a C‐terminal EGFP tag. Preliminary results suggest that TAAR1 localizes to the endoplasmic reticulum and mitochondria. Future experiments using similar techniques will explore TAAR1 localization in primary neurons. Together, these results will further elucidate the molecular mechanisms by which TAAR1 signals within neurons and ultimately alters neurotransmission.

The family of trace amine-associated receptors (TAARs) is distantly related to G protein-coupled biogenic aminergic receptors. TAARs are found in the brain as well as in the olfactory epithelium where they detect biogenic amines. However, the functional relationship of receptors from distinct TAAR subfamilies and in different species is still uncertain. Here, we perform a thorough phylogenetic analysis of 702 TAAR-like (TARL) and TAAR sequences from 48 species. We show that a clade of Tarl genes has greatly expanded in lampreys, whereas the other Tarl clade consists of only one or two orthologs in jawed vertebrates and is lost in amniotes. We also identify two small clades of Taar genes in sharks related to the remaining Taar genes in bony vertebrates, which are divided into four major clades. We further identify ligands for 61 orphan TARLs and TAARs from sea lamprey, shark, ray-finned fishes, and mammals, as well as novel ligands for two 5-hydroxytryptamine receptor 4 orthologs, a serotonin receptor subtype closely related to TAARs. Our results reveal a pattern of functional convergence and segregation: TARLs from sea lamprey and bony vertebrate olfactory TAARs underwent independent expansions to function as chemosensory receptors, whereas TARLs from jawed vertebrates retain ancestral response profiles and may have similar functions to TAAR1 in the brain. Overall, our data provide a comprehensive understanding of the evolution and ligand recognition profiles of TAARs and TARLs.

Members of a large family of odorant receptors found in olfactory sensory neurons (OSNs) in the main olfactory epithelium detect volatile odorants. Liberles and Buck prepared cDNA and then performed quantitative polymerase chain reaction (qPCR) to search mouse OSNs for G protein-coupled receptors (GPCRs) that had not been implicated previously in taste, odor, or pheromone detection. They identified two members of the trace amine-associated receptor (TAAR) family of GPCRs, which have been hypothesized to act in the brain as receptors for biogenic trace amines, such as tyramine and β-phenylethylamine. Further analysis with qPCR and in situ hybridization indicated that eight of nine mouse taar genes were expressed in olfactory epithelium and that, like odorant receptors, different TAARs are found in different OSNs. TAARs did not appear to be coexpressed with olfactory receptors but were coexpressed with Gα olf , the family of heterotrimeric guanine nucleotide-binding protein (G protein) α subunits through which odorant receptors signal. Individual mouse or human TAARs were coexpressed with an adenosine 3′,5′-monophosphate (cAMP)-dependent reporter gene in HEK293 cells that were cultured with potential ligands. Three mouse TAARs responded to volatile amines present in mouse urine: One detected β-phenylethylamine, which is associated with stress, and two others detected substances present in greater abundance in male than female urine. Thus, the authors propose that the TAARs may represent a family of chemosensory receptors that are functionally distinct from odorant receptors and appear to be associated with the detection of social cues. Ngai discusses implications of the research. S. D. Liberles, L. B. Buck, A second class of chemosensory receptors in the olfactory epithelium. Nature 442 , 645-650 (2006). [PubMed] J. Ngai, An extra dimension to olfaction. Nature 442 , 637-638 (2006). [PubMed]

The mammalian nose is a powerful chemosensor, capable of detecting and distinguishing a myriad of chemicals. Sensory neurons in the olfactory epithelium contain two types of chemosensory G protein-coupled receptors (GPCRs): odorant receptors (ORs), which are encoded by the largest gene family in mammals, and trace amine-associated receptors (TAARs), a smaller family of receptors distantly related to biogenic amine receptors. Do TAARs play a specialized role in olfaction distinct from that of ORs? Genes encoding TAARs are found in diverse vertebrates, from fish to mice to humans. Like OR genes, each Taar gene defines a unique population of canonical sensory neurons dispersed in a single zone of the olfactory epithelium. Ligands for mouse TAARs include a number of volatile amines, several of which are natural constituents of mouse urine, a rich source of rodent social cues. One chemical, 2-phenylethylamine, is reported to be enriched in the urine of stressed animals, and two others, trimethylamine and isoamylamine, are enriched in male versus female urine. Furthermore, isoamylamine has been proposed to be a pheromone that induces puberty acceleration in young female mice. These data raise the possibility that some TAARs are pheromone receptors in the nose, a hypothesis consistent with recent data suggesting that the olfactory epithelium contains dedicated pheromone receptors, separate from pheromone receptors in the vomeronasal organ. Future experiments will clarify the roles of TAARs in olfaction.

The trace amine-associated receptors (TAARs) form a specific family of G protein-coupled receptors in vertebrates. TAARs were initially considered neurotransmitter receptors, but recent study showed that mouse TAARs function as chemosensory receptors in the olfactory epithelium. To clarify the evolutionary dynamics of the TAAR gene family in vertebrates, near-complete repertoires of TAAR genes and pseudogenes were identified from the genomic assemblies of 4 teleost fishes (zebrafish, fugu, stickleback, and medaka), western clawed frogs, chickens, 3 mammals (humans, mice, and opossum), and sea lampreys. Database searches revealed that fishes had many putatively functional TAAR genes (13-109 genes), whereas relatively small numbers of TAAR genes (3-22 genes) were identified in tetrapods. Phylogenetic analysis of these genes indicated that the TAAR gene family was subdivided into 5 subfamilies that diverged before the divergence of ray-finned fishes and tetrapods. In tetrapods, virtually all TAAR genes were located in 1 specific region of their genomes as a gene cluster; however, in fishes, TAAR genes were scattered throughout more than 2 genomic locations. This possibly reflects a whole-genome duplication that occurred in the common ancestor of ray-finned fishes. Expression analysis of zebrafish and stickleback TAAR genes revealed that many TAARs in these fishes were expressed in the olfactory organ, suggesting the relatively high importance of TAARs as chemosensory receptors in fishes. A possible evolutionary history of the vertebrate TAAR gene family was inferred from the phylogenetic and comparative genomic analyses.

Trace amine-associated receptor 1 (TAAR1) is a member of TAAR family of G protein-coupled receptors (GPCRs). The members of this class of receptors discovered in 2001 have been found in some tissues ranging from the central nervous system to the olfactory epithelium and in some peripheral organs. The best studied receptor, TAAR1, is activated by a class of compounds named trace amines (TAs) that include compounds such as β-phenylethylamine (PEA), p-tyramine, octopamine, and tryptamine normally present at low levels in the mammalian brain. Although TA levels have been associated with many neuropsychiatric disorders, only the discovery of TAAR1 validated their physiological role. TAAR1 can modulate monoamine neurotransmission and, in particular, dopamine systems. Several studies have demonstrated that TAAR1 knockout (TAAR1-KO) mice display a supersensitive dopaminergic system, while activation of TAAR1 can reduce dopaminergic hyperactivity obtained either with pharmacological tools or present in genetic mouse model. For these reasons, TAAR1 has been proposed as a novel therapeutic target for neuropsychiatric disorders such as schizophrenia, bipolar disorder, and addiction. Moreover, several peripheral functions of TAAR1 have been described recently indicating intriguing novel TAAR1 roles in system physiology. Here we will review brain and peripheral functions mediated by TAAR1 and other TAARs.