RESUMO
A custom CGH microarray that covers the SOX9 regulatory region. Log2 ratio scatterplot showing individual data points. Blue box highlights copy number gain with 3' breakpoint region magnified.
Assuntos
Duplicação Gênica , Cariótipo , Sequências Reguladoras de Ácido Nucleico , Fatores de Transcrição SOX9/genética , Desenvolvimento Sexual/genética , Doenças Testiculares/diagnóstico , Doenças Testiculares/genética , Adolescente , Biomarcadores , Estudos de Associação Genética , Humanos , Masculino , Especificidade de Órgãos/genética , FenótipoRESUMO
Hoxb8 overexpression immortalises haematopoietic progenitor cells in a growth-factor-dependant manner and co-operates with interleukin-3 (IL-3) to cause acute myeloid leukaemia. To further understand how Hoxb8 contributes to myeloid cell immortalisation, we generated IL-3-dependant myeloid cells expressing Hoxb8 under the control of an inducible promoter. Downregulation of Hoxb8, in the presence of IL-3, caused cell-cycle arrest and apoptosis in the majority of cells. Apoptosis was dependant on Bax and Bak and, in part, on Bim, which was repressed by Hoxb8. Deletion of the miR-17â¼92 seed sequences in the Bim 3'UTR abolished Hoxb8-dependant regulation of Bim reporter constructs. Expression of all six miRNAs from this cluster were elevated when Hoxb8 was overexpressed. The miR-17â¼92 cluster was required for repression of Bim in Hoxb8-immortalised cells and deletion of the miR-17â¼92 cluster substantially inhibited Hoxb8, but not Hoxa9, mediated survival and proliferation. Hoxb8 appears to promote miR-17â¼92 expression through c-Myc, a known transcriptional regulator of the miR-17â¼92 cluster. We have uncovered a previously unrecognised link between Hoxb8 expression and microRNAs that provides a new insight into the oncogenic functions of Hoxb8.
Assuntos
Proteínas de Homeodomínio/genética , MicroRNAs/metabolismo , Regiões 3' não Traduzidas , Animais , Apoptose/genética , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismo , Proteína 11 Semelhante a Bcl-2 , Morte Celular/genética , Diferenciação Celular/genética , Processos de Crescimento Celular/genética , Regulação da Expressão Gênica , Proteínas de Homeodomínio/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , MicroRNAs/genética , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Transfecção , Proteína Killer-Antagonista Homóloga a bcl-2/metabolismo , Proteína X Associada a bcl-2/metabolismoRESUMO
In an effort to identify novel candidate genes involved in testis determination, we previously used suppression subtraction hybridisation PCR on male and female whole embryonic (12.0-12.5 days post coitum) mouse gonads. One gene to emerge from our screen was Redd1. In the current study, we demonstrate by whole-mount in situ hybridisation that Redd1 is differentially expressed in the developing mouse gonad at the time of sex determination, with higher expression in testis than ovary. Furthermore, Redd1 expression was first detected as Sry expression peaks, immediately prior to morphological sex determination, suggesting a potential role for Redd1 during testis development. To determine the functional importance of this gene during testis development, we generated Redd1-deficient mice. Morphologically, Redd1-deficient mice were indistinguishable from control littermates and showed normal fertility. Our results show that Redd1 alone is not required for testis development or fertility in mice. The lack of a male reproductive phenotype in Redd1 mice may be due to functional compensation by the related gene Redd2.
Assuntos
Reprodução/fisiologia , Testículo/embriologia , Fatores de Transcrição/metabolismo , Animais , Biomarcadores/metabolismo , Cruzamentos Genéticos , Embrião de Mamíferos/metabolismo , Desenvolvimento Embrionário/genética , Feminino , Fertilidade , Feto/embriologia , Imunofluorescência , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Camundongos , Fenótipo , Reação em Cadeia da Polimerase , Reprodutibilidade dos Testes , Células de Sertoli/metabolismo , Testículo/citologia , Testículo/crescimento & desenvolvimento , Testículo/metabolismo , Fatores de Transcrição/deficiência , Fatores de Transcrição/genéticaRESUMO
The sex of most mammals is determined by the action of SRY. Its presence initiates testis formation resulting in male differentiation, its absence results in ovary formation and female differentiation. We have used suppression subtraction hybridisation between 12.0-12.5 days postcoitum (dpc) mouse testes and ovaries to identify genes that potentially lie within the Sry pathway. Normalised urogenital ridge libraries comprising 8,352 clones were differentially screened with subtracted probes. A total of 272 candidate cDNAs were tested for qualitative differential expression and localisation by whole mount in situ hybridisation; germ cell-dependent or -independent expression was further resolved using busulfan. Fifty-four genes were identified that showed higher expression in the testis than the ovary. One novel gene may be a candidate for interactions with WT1, based on its localisation to Sertoli cells and map position (16q24.3).
