Role of vertebrate GAGA associated factor (vGAF) in early development of zebrafish.

Homeotic genes and their genomic organization show remarkable conservation across bilaterians. Consequently, the regulatory mechanisms, which control hox gene expression, are also highly conserved. The crucial presence of conserved GA rich motifs between Hox genes has been previously observed but what factor binds to these is still unknown. Previously we have reported that the vertebrate homologue of Drosophila Trl-GAF preferentially binds to GA rich regions in Evx2-hoxd13 intergenic region of vertebrate HoxD cluster. In this study, we show that the vertebrate-GAF (v-GAF) binds at known cis-regulatory elements in the HoxD complex of zebrafish and mouse. We further used morpholino based knockdown and CRISPR-cas9 knockout technique to deplete the v-GAF in zebrafish. We checked expression of the HoxD genes and found gain of the HoxD4 gene in GAF knockout embryos. Further, we partially rescued the morphological phenotypes in GAF depleted embryos by providing GAF mRNA. Our results show that GAF binds at intergenic regions of the HoxD complex and is important for maintaining the spatial domains of HoxD4 expression during embryonic development.

Chromatin Insulators and Topological Domains: Adding New Dimensions to 3D Genome Architecture.

The spatial organization of metazoan genomes has a direct influence on fundamental nuclear processes that include transcription, replication, and DNA repair. It is imperative to understand the mechanisms that shape the 3D organization of the eukaryotic genomes. Chromatin insulators have emerged as one of the central components of the genome organization tool-kit across species. Recent advancements in chromatin conformation capture technologies have provided important insights into the architectural role of insulators in genomic structuring. Insulators are involved in 3D genome organization at multiple spatial scales and are important for dynamic reorganization of chromatin structure during reprogramming and differentiation. In this review, we will discuss the classical view and our renewed understanding of insulators as global genome organizers. We will also discuss the plasticity of chromatin structure and its re-organization during pluripotency and differentiation and in situations of cellular stress.

Homeotic gene regulation: a paradigm for epigenetic mechanisms underlying organismal development.

The organization of eukaryotic genome into chromatin within the nucleus eventually dictates the cell type specific expression pattern of genes. This higher order of chromatin organization is established during development and dynamically maintained throughout the life span. Developmental mechanisms are conserved in bilaterians and hence they have body plan in common, which is achieved by regulatory networks controlling cell type specific gene expression. Homeotic genes are conserved in metazoans and are crucial for animal development as they specify cell type identity along the anterior-posterior body axis. Hox genes are the best studied in the context of epigenetic regulation that has led to significant understanding of the organismal development. Epigenome specific regulation is brought about by conserved chromatin modulating factors like PcG/trxG proteins during development and differentiation. Here we discuss the conserved epigenetic mechanisms relevant to homeotic gene regulation in metazoans.