RESUMEN
Accessible chromatin often represents gene regulatory elements, including promoters and enhancers, essential for gene expression. Assay for Transposase Accessible Chromatin sequencing (ATAC-seq) is one of the most popular techniques to investigate chromatin accessibility across the genome. Here we describe, step by step, a series of optimized experimental methods and bioinformatics pipelines for ATAC-seq analysis. As an example, we present an analysis of murine spermatogenic cells: a method to isolate germ cells, a reaction step using Tn5 transposase to insert sequencing adapters into accessible DNA, a library preparation method for high-throughput sequencing, and bioinformatics analysis of sequencing data. Overall, we introduce a framework of ATAC-seq analysis that can be applied to any cell population to identify cell-type-specific gene regulatory elements and their cis-regulatory networks.
Asunto(s)
Secuenciación de Inmunoprecipitación de Cromatina , Cromatina , Animales , Cromatina/genética , ADN/genética , Análisis de Datos , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Ratones , Análisis de Secuencia de ADN/métodos , Transposasas/genética , Transposasas/metabolismoRESUMEN
Genes involved in sexual reproduction diverge rapidly as a result of reproductive fitness. Here, we identify a novel protein domain in the germline-specific Polycomb protein SCML2 that is required for the establishment of unique gene expression programs after the mitosis-to-meiosis transition in spermatogenesis. We term this novel domain, which is comprised of rapidly evolved, DNA-binding repeat units of 28 amino acids, the SCML2 DNA-binding (SDB) repeats. These repeats are acquired in a specific subgroup of the rodent lineage, having been subjected to positive selection in the course of evolution. Mouse SCML2 has two DNA-binding domains: one is the SDB repeats and the other is an RNA-binding region, which is conserved in human SCML2. For the recruitment of SCML2 to target loci, the SDB repeats cooperate with the other functional domains of SCML2 to bind chromatin. The cooperative action of these domains enables SCML2 to sense DNA hypomethylation in an in vivo chromatin environment, thereby enabling SCML2 to bind to hypomethylated chromatin. We propose that the rapid evolution of SCML2 is due to reproductive adaptation, which has promoted species-specific gene expression programs in spermatogenesis.