Functional validation of a cis-regulatory element associated with coagulation disturbances and severe COVID-19

Although there is an accelerated advance in the knowledge of the COVID-19 pathogenesis, the causes that lead patients to respond in different ways during the evolution of this disease are poorly known. Enhancers are cis-regulatory elements which are the main determinants of the specificity of gene expression in different cell types. In addition, enhancers are considered important regions hotspots to genetic predisposition to diseases. Based on public data, mainly from: Genome-wide association studies (GWAS), High-throughput chromosome conformation capture (Hi-C), Chromatin Immunoprecipitation with sequencing (ChIP-seq), and regions DNAse I hypersensitive sites, we collected evidence which indicate that a genetic variant associated with severe COVID-19 is located within a putative enhancer of a gene previously associated with coagulopathies. Therefore, in this project we propose to carry out functional assays (ex: capture of chromatin conformation - 3C, in vitro enhancer activity through luciferase reporter, ChIP-qPCR, ATAC-seq, deletion of the enhancer region using CRISPR / Cas9 technology, and in vivo enhancer activity through the generation of transgenic mouse embryos) to evaluate if the enhancer predicted by in silico analyzes, participate of the transcriptional regulation of the gene associated with coagulopathies cited above. The execution of this project may bring new knowledge about the genetic and epigenetic role in the normal coagulation process, as well as coagulopathies and COVID-19 pathogenesis. (AU)