Percorrer por autor "Elgar, Greg"
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- Calcitonin: characterisation and expression in a teleost fish, Fugu rubripesPublication . Clark, M. S.; Bendell, L. A.; Power, Deborah; Warner, S.; Elgar, Greg; Ingleton, P. M.The present report describes the structure and expression of the calcitonin gene in Fugu rubripes. It is composed of 4 exons and 3 introns. Splicing of exons 1, 2 and 3 generates the calcitonin pre-proprotein, while splicing of exons 1, 2 and 4 generates calcitonin gene-related protein (CGRP). Exons 1 and 2 encoding the signal sequence and the N-terminal peptide are common in both the gene products and this gene organisation has been conserved in human, rat, chicken and salmon. The gene environment around calcitonin in Fugu has been poorly conserved when compared with human, apart from a small gene cluster. The calcitonin gene in Fugu has a widespread tissue distribution but it is most highly expressed in the brain. The abundance of gene expression in the ultimobranchial gland and the pituitary indicates that these are important sites of production and that the peptide is probably secreted into the circulation and/or acts as a paracrine or autocrine controlling factor. Whilst the function of calcitonin in fish is still largely unknown, the distribution described here suggests that one of the potential functions may be as a neuropeptide.
- Calcitonin: characterisation and expression in a teleost fish, Fugu rubripesPublication . Clark, Melody; Bendell, L; Power, DM; Warner, S; Elgar, Greg; Ingleton, P. M.The present report describes the structure and expression of the calcitonin gene in Fugu rubripes. It is composed of 4 exons and 3 introns. Splicing of exons 1, 2 and 3 generates the calcitonin pre-proprotein, while splicing of exons 1, 2 and 4 generates calcitonin gene-related protein (CGRP). Exons 1 and 2 encoding the signal sequence and the N-terminal peptide are common in both the gene products and this gene organisation has been conserved in human, rat, chicken and salmon. The gene environment around calcitonin in Fugu has been poorly conserved when compared with human, apart from a small gene cluster. The calcitonin gene in Fugu has a widespread tissue distribution but it is most highly expressed in the brain. The abundance of gene expression in the ultimobranchial gland and the pituitary indicates that these are important sites of production and that the peptide is probably secreted into the circulation and/or acts as a paracrine or autocrine controlling factor. Whilst the function of calcitonin in fish is still largely unknown, the distribution described here suggests that one of the potential functions may be as a neuropeptide.
- Duplicated receptors for VIP and PACAP VPAC 1 R and PAC 1 R in a teleost fish, Fugu rubripesPublication . Cardoso, João CR; Power, Deborah; Elgar, Greg; Clark, M. S.Two principal groups of receptors orthologous with human PAC1R and VPAC1R and were identified and characterised at the genomic level in the teleost fish Fugu rubripes. An additional group orthologous with VPAC2R was also identified and partially characterised. In Fugu, gene duplication of each of the PAC1Rs, VPAC1Rs and VPAC2Rs appears to have occurred. The topology of the tree surrounding the Fugu duplications and other isolated piscine sequences indicates that the duplication events for these six genes clearly preceded the speciation event leading to the Cypriniformes and Tetraodontiformes and is probably teleost-specific. Overall, the combined pattern of gene expression for each pair of duplicated genes mirrored the expression in other vertebrates. However, within each pair of duplicates further specialisation had occurred, with each demonstrating differential tissue distribution profiles suggesting they that may be responsible for the divergent action of the ligands, vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating peptide (PACAP). The Fugu VPAC1R gene regions showed conserved synteny with human chromosome 3p21·3 and also C. elegans chromosome X, indicating that the putative ancestral human chromosome 3 region may be equivalent to chromosome X in Caenorhabditis elegans.
- Genomic characterisation of putative growth hormone releasing hormone (GHRH) receptor genes in the teleost fish Fugu rubripesPublication . Cardoso, João CR; Power, Deborah; Elgar, Greg; Clark, M. S.The genomic organisation of a putative GHRH receptor gene has been characterised in the teleost fish, Fugu rubripes. It comprises 13 exons and is at least 1.2 3 larger than previously described vertebrate GHRH receptors. Sequence conservation with other vertebrate GHRH receptors is highest over the seven transmembrane domains with preservation of 8 conserved cysteines and an N-glycosylation site in the N-terminal region, which are important for receptor activation. Database mining revealed a further putative GHRH receptor in Fugu, with phylogenetic tree topology indicating that this was potentially a teleost-specific duplication event. This is the first time that the genomic organisation of a GHRH receptor gene has been characterised from a nonmammalian vertebrate.
- Genomic structure and expression of parathyroid hormone-related protein gene (PTHrP) in a teleost, Fugu rubripesPublication . Power, Deborah; Ingleton, P. M.; Flanagan, J. A.; Canario, Adelino V. M.; Danks, J. A.; Elgar, Greg; Clark, M. S.In this study we describe the isolation and characterisation of the parathyroid hormone-related protein (PTHrP) gene from the teleost Fugu rubripes. The gene has a relatively simple structure, compared with tetrapod PTHrP genes, composed of three exons and two introns, encompassing 2.25 kb of genomic DNA. The gene encodes a protein of 163 amino acids, with a putative signal peptide of 37 amino acids and a mature peptide of 126 amino acids. The overall homology with known tetrapod PTHrP proteins is low (36%), with a novel sequence inserted between positions 38 and 65, the absence of the conserved pentapeptide (TRSAW) and shortened C-terminal domain. The N-terminus shows greater conservation (62%), suggesting that it may have a hypercalcaemic function similar to that of tetrapod PTHrP. In situ localisation and RT–PCR have demonstrated the presence of PTHrP in a wide range of tissues with varying levels of expression. Sequence scanning of overlapping cosmids has identified three additional genes, TMPO, LDHB and KCNA1, which map to human chromosome 12, with the latter two mapping to 12p12-11.2. PTHrP in human also maps to this chromosome 12 sub-region, thus demonstrating conservation of synteny between human and Fugu.
- Isolation and characterisation of the corticotropin releasing factor receptor 1 (CRFR1) gene in a teleost fish, Fugu rubripesPublication . Cardoso, João CR; Power, Deborah; Elgar, Greg; Clark, M. S.Corticotropin releasing factor receptor (CRF) is a member of the secretin family of the G-protein coupled receptor superfamily. These are characterised by the presence of seven transmembrane domains and six conserved cysteines that are important for receptor conformation and ligand binding. In vertebrates two CRF receptors (CRF1 and CRF2) have been isolated and characterised. In this study the complete structure of the CRF1 receptor was isolated and partially characterised for the first time in a vertebrate using the compact genome of the Japanese pufferfish, Fugu rubripes as a model. The Fugu CRF1 receptor gene is composed of 14 exons is approximately 27 kb in length. A tissue distribution of this receptor in Fugu reveals that it is expressed mainly in liver, gonads, heart and brain, however, expression in the kidney, gut and gills was also detected. In vertebrates this receptor appears to have a different tissue distribution and its presence in the gills may indicate a new role in osmoregulatory processes.
- Persistence of duplicated PAC(1) receptors in the teleost, Sparus auratusPublication . Cardoso, J. C. R.; Vet, Edwin C. J. M. de; Louro, Bruno; Elgar, Greg; Clar, M. S.; Power, DeborahBackground: Duplicated genes are common in vertebrate genomes. Their persistence is assumed to be either a consequence of gain of novel function (neofunctionalisation) or partitioning of the function of the ancestral molecule (sub-functionalisation). Surprisingly few studies have evaluated the extent of such modifications despite the numerous duplicated receptor and ligand genes identified in vertebrate genomes to date. In order to study the importance of function in the maintenance of duplicated genes, sea bream (Sparus auratus) PAC1 receptors, sequence homologues of the mammalian receptor specific for PACAP (Pituitary Adenylate Cyclase-Activating Polypeptide), were studied. These receptors belong to family 2 GPCRs and most of their members are duplicated in teleosts although the reason why both persist in the genome is unknown. Results: Duplicate sea bream PACAP receptor genes (sbPAC1A and sbPAC1B), members of family 2 GPCRs, were isolated and share 77% amino acid sequence identity. RT-PCR with specific primers for each gene revealed that they have a differential tissue distribution which overlaps with the distribution of the single mammalian receptor. Furthermore, in common with mammals, the teleost genes undergo alternative splicing and a PAC1Ahop1 isoform has been characterised. Duplicated orthologous receptors have also been identified in other teleost genomes and their distribution profile suggests that function may be species specific. Functional analysis of the paralogue sbPAC1s in Cos7 cells revealed that they are strongly stimulated in the presence of mammalian PACAP27 and PACAP38 and far less with VIP (Vasoactive Intestinal Peptide). The sbPAC1 receptors are equally stimulated (LOGEC50 values for maximal cAMP production) in the presence of PACAP27 (-8.74 ± 0.29 M and -9.15 ± 0.21 M, respectively for sbPAC1A and sbPAC1B, P > 0.05) and PACAP38 (-8.54 ± 0.18 M and -8.92 ± 0.24 M, respectively for sbPAC1A and sbPAC1B, P > 0.05). Human VIP was found to stimulate sbPAC1A (-7.23 ± 0.20 M) more strongly than sbPAC1B (-6.57 ± 0.14 M, P <0.05) and human secretin (SCT), which has not so far been identified in fish genomes, caused negligible stimulation of both receptors. Conclusion: The existence of functionally divergent duplicate sbPAC1 receptors is in line with previously proposed theories about the origin and maintenance of duplicated genes. Sea bream PAC1 duplicate receptors resemble the typical mammalian PAC1, and PACAP peptides were found to be more effective than VIP in stimulating cAMP production, although sbPAC1A was more responsive for VIP than sbPAC1B. These results together with the highly divergent pattern of tissue distribution suggest that a process involving neofunctionalisation occurred after receptor duplication within the fish lineage and probably accounts for their persistence in the genome. The characterisation of further duplicated receptors and their ligands should provide insights into the evolution and function of novel protein-protein interactions associated with the vertebrate radiation.