RESUMO
Mammalian ovarian G-protein-coupled receptor 1 (OGR1) is activated by some metals in addition to extracellular protons and coupling to multiple intracellular signaling pathways. In the present study, we examined whether zebrafish OGR1, zebrafish GPR4, and human GPR4 (zOGR1, zGPR4, and hGPR4, respectively) could sense the metals and activate the intracellular signaling pathways. On one hand, we found that only manganese and cobalt of the tested metals stimulated SRE-promoter activities in zOGR1-overexpressed HEK293T cells. On the other hand, none of the metals tested stimulated the promoter activities in zGPR4- and hGPR4-overexpressed cells. The OGR1 mutant (H4F), which is lost to activation by extracellular protons, did not stimulate metal-induced SRE-promoter activities. These results suggest that zOGR1, but not GPR4, is also a metal-sensing G-protein-coupled receptor in addition to a proton-sensing G-protein-coupled receptor, although not all metals that activate hOGR1 activated zOGR1.
Assuntos
Receptores Acoplados a Proteínas G/genética , Proteínas de Peixe-Zebra/genética , Animais , Cobalto/farmacologia , AMP Cíclico , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Manganês/farmacologia , Regiões Promotoras Genéticas/genética , Prótons , Transdução de Sinais/efeitos dos fármacos , Peixe-Zebra/genéticaRESUMO
BACKGROUND: Forward genetic screens in mice provide an unbiased means to identify genes and other functional genetic elements in the genome. Previously, a large scale ENU mutagenesis screen was conducted to query the functional content of a ~50 Mb region of the mouse genome on proximal Chr 5. The majority of phenotypic mutants recovered were embryonic lethals. RESULTS: We report the high resolution genetic mapping, complementation analyses, and positional cloning of mutations in the target region. The collection of identified alleles include several with known or presumed functions for which no mutant models have been reported (Tbc1d14, Nol14, Tyms, Cad, Fbxl5, Haus3), and mutations in genes we or others previously reported (Tapt1, Rest, Ugdh, Paxip1, Hmx1, Otoe, Nsun7). We also confirmed the causative nature of a homeotic mutation with a targeted allele, mapped a lethal mutation to a large gene desert, and localized a spermiogenesis mutation to a region in which no annotated genes have coding mutations. The mutation in Tbc1d14 provides the first implication of a critical developmental role for RAB-GAP-mediated protein transport in early embryogenesis. CONCLUSION: This collection of alleles contributes to the goal of assigning biological functions to all known genes, as well as identifying novel functional elements that would be missed by reverse genetic approaches.
Assuntos
Mapeamento Cromossômico , Cromossomos/efeitos dos fármacos , Análise Mutacional de DNA , Desenvolvimento Embrionário/genética , Camundongos/genética , Mutação , Animais , Clonagem Molecular , Etilnitrosoureia/toxicidade , Genes Letais , Teste de Complementação Genética , Masculino , Camundongos Endogâmicos C57BL , Deleção de Sequência , Espermatogênese/genéticaRESUMO
L5Jcs1 is a perinatal lethal mutation uncovered in a screen for ENU-induced mutations on mouse chromosome 5. L5Jcs1 homozygotes exhibit posterior-to-anterior transformations of the vertebral column midsection, similar to mice deficient for Hoxc8 and Hoxc9. Positional cloning efforts identified a mutation in a novel, evolutionarily conserved, and ubiquitously expressed gene dubbed Tapt1 (Transmembrane anterior posterior transformation 1). TAPT1 is predicted to contain several transmembrane domains, and part of the gene is orthologous to an unusual alternatively spliced human transcript encoding the cytomegalovirus gH receptor. We speculate that TAPT1 is a downstream effector of HOXC8 that may act by transducing or transmitting extracellular information required for axial skeletal patterning during development.
Assuntos
Osso e Ossos/anormalidades , Proteínas de Homeodomínio/genética , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Mutação/genética , Sequência de Aminoácidos , Animais , Animais Recém-Nascidos , Sequência de Bases , Cromossomos de Mamíferos/genética , Clonagem Molecular , Sequência Conservada , Análise Mutacional de DNA , Embrião de Mamíferos/anormalidades , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Genes Homeobox , Proteínas de Homeodomínio/metabolismo , Humanos , Proteínas de Membrana/química , Camundongos , Dados de Sequência Molecular , Penetrância , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Superfície Celular/metabolismo , Proteínas do Envelope Viral/metabolismoRESUMO
We report here the first genome-wide functional genomic screen for longevity genes. We systematically surveyed Caenorhabditis elegans genes using large-scale RNA interference (RNAi), and found that RNAi inactivation of 89 genes extend C. elegans lifespan. Components of the daf-2/insulin-like signaling pathway are recovered, as well as genes that regulate metabolism, signal transduction, protein turnover, and gene expression. Many of these candidate longevity genes are conserved across animal phylogeny. Genetic interaction analyses with the new longevity genes indicate that some act upstream of the daf-16/FOXO transcription factor or the sir2.1 protein deacetylase, and others function independently of daf-16/FOXO and sir2.1, and might define new pathways to regulate lifespan.
