RESUMO
Recent genomewide association studies have found multiple genetic variants on chromosome 8q24 that are significantly associated with an increased susceptibility to prostate, colorectal, and breast cancer. These risk loci are located in a "gene desert," a few hundred kilobases telomeric to the Myc gene. To date, the biological mechanism(s) underlying these associations remain unclear. It has been speculated that these 8q24 genetic variant(s) might affect Myc expression by altering its regulation or amplification status. Here, we show that multiple enhancer elements are present within this region and that they can regulate transcription of Myc. We also demonstrate that one such enhancer element physically interacts with the Myc promoter via transcription factor Tcf-4 binding and acts in an allele specific manner to regulate Myc expression.
Assuntos
Cromossomos Humanos Par 8/genética , Elementos Facilitadores Genéticos/genética , Regulação da Expressão Gênica/genética , Predisposição Genética para Doença/genética , Neoplasias/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Sequência de Bases , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Imunoprecipitação da Cromatina , Biologia Computacional , Primers do DNA/genética , Humanos , Luciferases , Dados de Sequência Molecular , Neoplasias/metabolismo , Fator de Transcrição 4 , Fatores de Transcrição/metabolismo , beta Catenina/metabolismoRESUMO
The efficient use of nutrients is important in development and aging. In this study, we asked if the protein repair methyltransferase has a related or additional role in energy metabolism and stress response in the nematode Caenorhabditis elegans. Worms lacking the pcm-1 gene encoding this enzyme exhibit reduced longevity as SDS-isolated dauer larvae and as arrested L1 larvae under starvation stress, while overexpression leads to increased adult longevity. These findings led us to question whether pcm-1 deficient C. elegans may have inappropriate metabolic responses to stress. We assayed dauer and dauer-like larvae for starvation survival and observed a two-fold reduction of median survival time for pcm-1 mutants compared to N2 wild-type worms. Under these conditions, pcm-1 deficient dauer larvae had reduced fat stores, suggesting that PCM-1 may have a role in the initiation of the correct metabolic responses to stress starvation. We show expression of the pcm-1 gene in neurons, body wall and reproductive tissues. Upon heat shock and dauer formation-inducing conditions, we observe additional pcm-1 expression in body wall muscle nuclei and actomyosin filaments and in hypodermal cells. These results suggest that this enzyme may be important in stress response pathways, including proper decision making for energy storage.
Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/enzimologia , Proteínas de Ciclo Celular/metabolismo , Metiltransferases/metabolismo , Envelhecimento/genética , Envelhecimento/metabolismo , Animais , Animais Geneticamente Modificados , Sequência de Bases , Caenorhabditis elegans/genética , Caenorhabditis elegans/crescimento & desenvolvimento , Proteínas de Caenorhabditis elegans/genética , Proteínas de Ciclo Celular/genética , Primers do DNA/genética , DNA de Helmintos/genética , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Regulação Enzimológica da Expressão Gênica , Genes de Helmintos , Resposta ao Choque Térmico , Larva/enzimologia , Larva/genética , Larva/crescimento & desenvolvimento , Metabolismo dos Lipídeos , Longevidade/genética , Longevidade/fisiologia , Masculino , Metiltransferases/genética , Mutação , Fenótipo , Regiões Promotoras Genéticas , Inanição , Estresse Fisiológico , Distribuição TecidualRESUMO
Protein damage that accumulates during aging can be mitigated by a repair methyltransferase, the l-isoaspartyl-O-methyltransferase. In Caenorhabditis elegans, the pcm-1 gene encodes this enzyme. In response to pheromone, we show that pcm-1 mutants form fewer dauer larvae with reduced survival due to loss of the methyltransferase activity. Mutations in daf-2, an insulin/insulin-like growth factor-1-like receptor, and daf-7, a transforming growth factor-beta-like ligand, modulate pcm-1 dauer defects. Additionally, daf-2 and daf-7 mutant dauer larvae live significantly longer than wild type. Although dauer larvae are resistant to many environmental stressors, a proportionately larger decrease in dauer larvae life spans occurred at 25 degrees C compared to 20 degrees C than in adult life span. At 25 degrees C, mutation of the daf-7 or pcm-1 genes does not change adult life span, whereas mutation of the daf-2 gene and overexpression of PCM-1 increases adult life span. Thus, there are both overlapping and distinct mechanisms that specify dauer and adult longevity.
Assuntos
Envelhecimento/metabolismo , Proteínas de Caenorhabditis elegans/fisiologia , Caenorhabditis elegans/fisiologia , Proteínas de Ciclo Celular/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Metiltransferases/fisiologia , Envelhecimento/genética , Animais , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Proteínas de Ciclo Celular/genética , Epistasia Genética , Fatores de Transcrição Forkhead , Regulação da Expressão Gênica no Desenvolvimento/genética , Genes de Helmintos , Larva/fisiologia , Longevidade/genética , Longevidade/fisiologia , Metiltransferases/genética , Mutação , Feromônios , Proteína D-Aspartato-L-Isoaspartato Metiltransferase/genética , Receptor de Insulina/fisiologia , Transdução de Sinais , Fatores de Transcrição/fisiologia , Fator de Crescimento Transformador beta/fisiologiaRESUMO
The protein L-isoaspartyl-O-methyltransferase, coded by the pcm-1 gene in Caenorhabditis elegans, participates in the repair of age-damaged proteins. We tested the ability of pcm-1-deficient nematodes to survive starvation stress as developmentally-arrested L1 larvae. We found that pcm-1 mutant L1 larvae do not survive as well as wild-type L1 larvae when incubated in M9 medium without nutrients. We then tested whether the starved L1 larvae could continue development when allowed access to food in a recovery assay. A loss of recovery ability with age was observed for all larvae, with little or no difference between the pcm-1 mutant and wild-type N2 larvae. Interestingly, when L1 larvae were starved in cholesterol-containing S medium or M9 medium supplemented with cholesterol, the survival rates of both mutant and wild-type animals nearly doubles, with pcm-1 larvae again faring more poorly than N2 larvae. Furthermore, L1 larvae cultured in these cholesterol-containing media show an increase in Sudan Black staining over animals cultured in M9 medium. The longevity defects of pcm-1 mutants previously seen in dauer larvae and here in L1 larvae suggest a defect in the ability of pcm-1 mutants to recycle and reuse old cellular components in pathways such as autophagy. Using an autophagosomal marker, we found evidence suggesting that the pcm-1 mutation may inhibit autophagy during dauer formation, suggesting that the absence of protein repair may also interfere with protein degradation pathways.
