RESUMEN
Active DNA demethylation via ten-eleven translocation (TET) family enzymes is essential for epigenetic reprogramming in cell state transitions. TET enzymes catalyze up to three successive oxidations of 5-methylcytosine (5mC), generating 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), or 5-carboxycytosine (5caC). Although these bases are known to contribute to distinct demethylation pathways, the lack of tools to uncouple these sequential oxidative events has constrained our mechanistic understanding of the role of TETs in chromatin reprogramming. Here, we describe the first application of biochemically engineered TET mutants that unlink 5mC oxidation steps, examining their effects on somatic cell reprogramming. We show that only TET enzymes proficient for oxidation to 5fC/5caC can rescue the reprogramming potential of Tet2-deficient mouse embryonic fibroblasts. This effect correlated with rapid DNA demethylation at reprogramming enhancers and increased chromatin accessibility later in reprogramming. These experiments demonstrate that DNA demethylation through 5fC/5caC has roles distinct from 5hmC in somatic reprogramming to pluripotency.
Asunto(s)
5-Metilcitosina/metabolismo , Reprogramación Celular , Proteínas de Unión al ADN/metabolismo , Embrión de Mamíferos/metabolismo , Elementos de Facilitación Genéticos , Epigénesis Genética , Fibroblastos/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Animales , Proteínas de Unión al ADN/genética , Dioxigenasas , Embrión de Mamíferos/citología , Fibroblastos/citología , Células HEK293 , Humanos , Ratones , Ratones Noqueados , Mutación , Células 3T3 NIH , Proteínas Proto-Oncogénicas/genéticaRESUMEN
We report the genome sequences of 14 cluster K mycobacteriophages isolated using Mycobacterium smegmatis mc²155 as host. Four are closely related to subcluster K1 phages, and 10 are members of subcluster K6. The phage genomes span considerable sequence diversity, including multiple types of integrases and integration sites.