RESUMO
Male development in mammals depends on the activity of the two SOX gene: Sry and Sox9, in the embryonic testis. As deletion of Enhancer 13 (Enh13) of the Sox9 gene results in XY male-to-female sex reversal, we explored the critical elements necessary for its function and hence, for testis and male development. Here, we demonstrate that while microdeletions of individual transcription factor binding sites (TFBS) in Enh13 lead to normal testicular development, combined microdeletions of just two SRY/SOX binding motifs can alone fully abolish Enh13 activity leading to XY male-to-female sex reversal. This suggests that for proper male development to occur, these few nucleotides of non-coding DNA must be intact. Interestingly, we show that depending on the nature of these TFBS mutations, dramatically different phenotypic outcomes can occur, providing a molecular explanation for the distinct clinical outcomes observed in patients harboring different variants in the same enhancer.
Assuntos
Elementos Facilitadores Genéticos , Processos de Determinação Sexual , Animais , Feminino , Humanos , Masculino , Camundongos , Sítios de Ligação/genética , Elementos Facilitadores Genéticos/genética , Deleção de Sequência , Processos de Determinação Sexual/genética , Proteína da Região Y Determinante do Sexo/genética , Proteína da Região Y Determinante do Sexo/metabolismo , Fatores de Transcrição SOX9/genética , Fatores de Transcrição SOX9/metabolismo , Testículo/crescimento & desenvolvimento , FenótipoRESUMO
TET2/3 play a well-known role in epigenetic regulation and mouse development. However, their function in cellular differentiation and tissue homeostasis remains poorly understood. Here we show that ablation of TET2/3 in intestinal epithelial cells results in a murine phenotype characterized by a severe homeostasis imbalance in the small intestine. Tet2/3-deleted mice show a pronounced loss of mature Paneth cells as well as fewer Tuft and more Enteroendocrine cells. Further results show major changes in DNA methylation at putative enhancers, which are associated with cell fate-determining transcription factors and functional effector genes. Notably, pharmacological inhibition of DNA methylation partially rescues the methylation and cellular defects. TET2/3 loss also alters the microbiome, predisposing the intestine to inflammation under homeostatic conditions and acute inflammation-induced death. Together, our results uncover previously unrecognized critical roles for DNA demethylation, possibly occurring subsequently to chromatin opening during intestinal development, culminating in the establishment of normal intestinal crypts.
Assuntos
Dioxigenases , Epigênese Genética , Animais , Camundongos , Diferenciação Celular/genética , Dioxigenases/metabolismo , Metilação de DNA/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Homeostase , Inflamação/metabolismo , Intestino Delgado/metabolismoRESUMO
Sex determination is the process by which an initial bipotential gonad adopts either a testicular or ovarian cell fate. The inability to properly complete this process leads to a group of developmental disorders classified as disorders of sex development (DSD). To date, dozens of genes were shown to play roles in mammalian sex determination, and mutations in these genes can cause DSD in humans or gonadal sex reversal/dysfunction in mice. However, exome sequencing currently provides genetic diagnosis for only less than half of DSD patients. This points towards a major role for the non-coding genome during sex determination. In this review, we highlight recent advances in our understanding of non-coding, cis-acting gene regulatory elements and discuss how they may control transcriptional programmes that underpin sex determination in the context of the 3-dimensional folding of chromatin. As a paradigm, we focus on the Sox9 gene, a prominent pro-male factor and one of the most extensively studied genes in gonadal cell fate determination.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Gônadas , Processos de Determinação Sexual , Animais , Transtornos do Desenvolvimento Sexual/genética , Feminino , Gônadas/crescimento & desenvolvimento , Humanos , Masculino , Mamíferos/genética , Camundongos , Ovário , Sequências Reguladoras de Ácido Nucleico , Fatores de Transcrição SOX9/genética , Processos de Determinação Sexual/genética , TestículoRESUMO
Although much research has been done on the diversity of the gut microbiome, little is known about how it influences intestinal homeostasis under normal and pathogenic conditions. Epigenetic mechanisms have recently been suggested to operate at the interface between the microbiota and the intestinal epithelium. We performed whole-genome bisulfite sequencing on conventionally raised and germ-free mice, and discovered that exposure to commensal microbiota induced localized DNA methylation changes at regulatory elements, which are TET2/3-dependent. This culminated in the activation of a set of 'early sentinel' response genes to maintain intestinal homeostasis. Furthermore, we demonstrated that exposure to the microbiota in dextran sodium sulfate-induced acute inflammation results in profound DNA methylation and chromatin accessibility changes at regulatory elements, leading to alterations in gene expression programs enriched in colitis- and colon-cancer-associated functions. Finally, by employing genetic interventions, we show that microbiota-induced epigenetic programming is necessary for proper intestinal homeostasis in vivo.
