Recent advances in understanding the 3D structure of chromosomes has led to increased focus on the role of genome architecture in regulating gene transcription. Above the nucleosomal level, distal DNA elements like enhancer can form loop with promoter to drive gene transcription, and many of these interacting elements would then locally form a topologically associating domain (TAD). Such TAD can be lineage specific and is thus believed to play a crucial role during immune cell development and differentiation. However, how such structure is established and maintained is poorly understood.
Gata3 is essential for the proper development of CD4+ T cells, particularly Th2 cells, and is therefore implicated in mediating allergic reaction such as asthma.
Here the genomic architecture of Gata3 in T and B lymphocytes was investigated using chromosome conformation capture coupled with high-throughput sequencing (Hi-C) and CRISPR/Cas9 genome editing technologies.
Hi-C data of T and B lymphocytes on the Gata3 regions revealed a T cell specific TAD where Gata3 promoter interacts with several upstream cis-elements, one of which is consistent with an already characterised Gata3 enhancer. By exploiting the CRISPR/Cas9 gene editing technology, these elements were bi-allelically deleted in a T cell lymphoma cell line EL4 and the expression of Gata3 was found to be heavily disrupted.
A novel non-coding RNA was discovered right at the aforementioned enhancer and the expression profile was found to correlate with Gata3. This might indicate its importance in regulating the Gata3 expression.
This preliminary work provides a framework of how cis-regulatory regions of the genome can be manipulated and this would further allow us to understand the role of the cistrome during immune development and differentiation.