RESUMO
The naturally occurring polyamines putrescine, spermidine or spermine are ubiquitous in all cells. Although polyamines have prominent regulatory roles in cell division and growth, precise molecular and cellular functions are not well-established in vivo. In this work we have performed microarray experiments with a spermidine synthase, spermine oxidase mutant (Deltaspe3 Deltafms1) strain to investigate the responsiveness of yeast genes to supplementation with spermidine or spermine. Expression analysis identified genes responsive to the addition of either excess spermidine (10(-5) M) or spermine (10(-5) M) compared to a control culture containing 10(-8) M spermidine. 247 genes were upregulated > two-fold and 11 genes were upregulated >10-fold after spermidine addition. Functional categorization of the genes showed induction of transport-related genes and genes involved in methionine, arginine, lysine, NAD and biotin biosynthesis. 268 genes were downregulated more than two-fold, and six genes were downregulated > eight-fold after spermidine addition. A majority of the downregulated genes are involved in nucleic acid metabolism and various stress responses. In contrast, only a few genes (18) were significantly responsive to spermine. Thus, results from global gene expression profiling demonstrate a more major role for spermidine in modulating gene expression in yeast than spermine.
Assuntos
Deleção de Genes , Regulação Fúngica da Expressão Gênica , Saccharomyces cerevisiae/enzimologia , Saccharomyces cerevisiae/fisiologia , Espermidina Sintase/genética , Espermidina/metabolismo , Espermina/metabolismo , Regulação para Baixo , Genes Fúngicos , Análise de Sequência com Séries de Oligonucleotídeos , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Regulação para CimaRESUMO
We investigated the role of LMNA in adipose tissue by developing a novel mouse model of lipodystrophy. Transgenic mice were generated that express the LMNA mutation that causes familial partial lipodystrophy of the Dunnigan type (FPLD2). The phenotype observed in FPLD-transgenic mice resembles many of the features of human FPLD2, including lack of fat accumulation, insulin resistance, and enlarged, fatty liver. Similar to the human disease, FPLD-transgenic mice appear to develop normally, but after several weeks they are unable to accumulate fat to the same extent as their wild-type littermates. One poorly understood aspect of lipodystrophies is the mechanism of fat loss. To this end, we have examined the effects of the FPLD2 mutation on fat cell function. Contrary to the current literature, which suggests FPLD2 results in a loss of fat, we found that the key mechanism contributing to the lack of fat accumulation involves not a loss, but an apparent inability of the adipose tissue to renew itself. Specifically, preadipocytes are unable to differentiate into mature and fully functional adipocytes. These findings provide insights not only for the treatment of lipodystrophies, but also for the study of adipogenesis, obesity, and insulin resistance.
Assuntos
Tecido Adiposo/metabolismo , Lamina Tipo A/genética , Lamina Tipo A/metabolismo , Lipodistrofia Parcial Familiar/genética , Lipodistrofia Parcial Familiar/metabolismo , Mutação , Adipócitos/metabolismo , Adipócitos/patologia , Tecido Adiposo/patologia , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Marrom/patologia , Animais , Sequência de Bases , Diferenciação Celular , Primers do DNA/genética , Modelos Animais de Doenças , Fígado Gorduroso/genética , Fígado Gorduroso/metabolismo , Fígado Gorduroso/patologia , Humanos , Resistência à Insulina/genética , Resistência à Insulina/fisiologia , Lipodistrofia Parcial Familiar/patologia , Lipólise/genética , Lipólise/fisiologia , Masculino , Camundongos , Camundongos Transgênicos , Fenótipo , Termogênese/genética , Termogênese/fisiologiaRESUMO
The Wilms' tumor suppressor gene (WT1) encodes a zinc finger transcription factor that is vital during development of several organs including metanephric kidneys. Despite the critical regulatory role of WT1, the pathways and mechanisms by which WT1 orchestrates development remain elusive. To identify WT1 target genes, we performed a genome-wide expression profiling analysis in cells expressing inducible WT1. We identified a number of direct WT1 target genes, including the epidermal growth factor (EGF)-family ligands epiregulin and HB-EGF, the chemokine CX3CL1, and the transcription factors SLUG and JUNB. The target genes were validated using quantitative reverse transcriptase-polymerase chain reaction, small interfering RNA knockdowns, chromatin immunoprecipitation, and luciferase reporter analyses. Immunohistochemistry of fetal kidneys confirmed that a number of the WT1 target genes had overlapping expression patterns with the highly restricted spatiotemporal expression of WT1. Finally, using an in vitro embryonic kidney culture assay, we found that the addition of recombinant epiregulin, amphiregulin, CX3CL1, and interleukin-11 significantly enhanced ureteric bud branching morphogenesis. Our genome-wide screen implicates WT1 in the transcriptional regulation of the EGF-family of growth factors as well as the CX3CL1 chemokine during nephrogenesis.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Rim/embriologia , Organogênese/fisiologia , Fatores de Transcrição/metabolismo , Proteínas WT1/metabolismo , Anfirregulina , Animais , Linhagem Celular Tumoral , Quimiocina CXCL1 , Quimiocinas CXC/biossíntese , Quimiocinas CXC/genética , Quimiocinas CXC/farmacologia , Família de Proteínas EGF , Fator de Crescimento Epidérmico/biossíntese , Fator de Crescimento Epidérmico/genética , Fator de Crescimento Epidérmico/farmacologia , Epirregulina , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Genoma/fisiologia , Glicoproteínas/farmacologia , Fator de Crescimento Semelhante a EGF de Ligação à Heparina , Humanos , Imuno-Histoquímica , Peptídeos e Proteínas de Sinalização Intercelular/farmacologia , Interleucina-11/biossíntese , Interleucina-11/genética , Interleucina-11/farmacologia , Rim/citologia , Organogênese/efeitos dos fármacos , Gravidez , Proteínas Proto-Oncogênicas c-jun/biossíntese , Proteínas Proto-Oncogênicas c-jun/genética , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Transcrição da Família Snail , Fatores de Transcrição/biossíntese , Fatores de Transcrição/genética , Proteínas WT1/genética , Dedos de Zinco/genéticaRESUMO
People with 59-200 CGG.CCG-repeats in the 5' UTR of one of their FMR1 genes are at risk for Fragile X tremor and ataxia syndrome. Females are also at risk for premature ovarian failure. These symptoms are thought to be due to the presence of the repeats at the DNA and/or RNA level. We show here that long transcribed but untranslated CGG-repeat tracts are toxic to human cells and alter the expression of a wide variety of different genes including caspase-8, CYFIP, Neurotensin and UBE3A.
