Convergent Transcription & fine-tuned Regulation of Genomic Imprinting – CORGI

During development, the transcriptome is profoundly reshaped by regulatory DNA sequences, transcription factors, epigenetic modifications, and the spatial reorganisation of the genome. In non-mammalian species, antisense transcription is an important regulatory mechanism of gene expression. However, the role of antisense transcription during mammalian development is underappreciated. How antisense transcription interacts with other mechanisms that reshape the transcriptome and whether it also influences genes that are distant from the convergence zone is also elusive.
The objective of CORGI (Convergent Transcription & fine-tuned Regulation of Genomic Imprinting) is to determine the influence of antisense transcription on the expression of a pair of convergent genes but also on genes distant from the convergence zone during neurodevelopment in mice.
Imprinted genes (IGs) play a major role in neurodevelopment. IGs are expressed according to the parental origin, and sometimes in an organ-specific manner. Convergent transcription is thought to be a mechanism involved in the brain-specific expression of certain IGs, including Mest/Copg2, a pair of convergent IGs whose deregulation is involved in neurodevelopmental disorders. Our experimental model is the cortex generated from hybrid embryonic stem cells (ESCs). This model recapitulates the in vivo epigenetic control of IG expression during neurodevelopment (including for Mest/Copg2).
Our unpublished data on Mest/Copg2 obtained by a broad panel of approaches (CRISPRi to modulate transcription, RRBS -allelic DNA methylation-, oriented RNA-seq -parental origin dependent expression-, Cut & Run-seq -allelic histone and protein factor markings-, and allelic 4C-seq -DNA-DNA interactome of the Mest ICR) support the hypothesis that convergent transcription regulates expression of IGs at the Mest locus by:
1) a proximal effect on convergent Mest and Copg2 transcripts (via collision of PolII RNAs transcribing them into antisense directions?)
2) a remote effect on other IGs at the Mest locus (conceivably by promoting an allele-specific 3D configuration, masking regulatory sequences in cis, or via 3D collision).
To further test our hypothesis, we will use brain organoids, genetic engineering tools based on CRISPR and high-resolution techniques (TT-seq, mNET-seq, RNA-seq, Cut & Run and 4C-seq) on a hybrid background to obtain allele-specific information. Our project relies on the complementary expertise of partner 1 (brain organoid model from hybrid ESCs and CRISPR-based tools to modulate transcription) and partner 2 (chromatin and 3D-based studies and associated bioinformatics analyses). Most of the tools have been already generated and we routinely use most of the techniques to modulate convergent transcription and estimate its effect on gene expression. After having extensively studied Mest/Copg2, we will extend our approach to other duos of convergent genes. Finally, we will produce atlas of convergent transcription events and RNA PolII occupancy during neural differentiation. This atlas, made publicly available, will be useful for unravelling the 'biology' of convergent transcriptional events during mammalian brain development.
CORGI will have a synergistic effect on the fields of convergent/antisense transcription and genomic imprinting. It should generate original insights into the influence of convergent transcription in the local and remote regulation of imprinted genes, providing 1) a framework to further elucidate convergent transcription role in cell fate acquisition in mammals, 2) a new means of understanding the events leading to aberrant regulation of imprinted genes in neurodevelopmental disorders.