Unveiling mechanisms mediating gene silencing and escape regulation in X-chromosome inactivation

• Grant number: 23/01147-7
• Support Opportunities: Scholarships abroad - Research Internship - Doctorate
• Start date: June 01, 2023
• End date: May 31, 2024
• Field of knowledge: Health Sciences - Medicine - Medical Clinics
• Principal Investigator: Maria Isabel de Souza Aranha Melaragno
• Grantee: Bianca Pereira Favilla
• Supervisor: Edith Heard
• Host Institution: Escola Paulista de Medicina (EPM). Universidade Federal de São Paulo (UNIFESP). Campus São Paulo. São Paulo , SP, Brazil
• Institution abroad: European Molecular Biology Laboratory, Heidelberg (EMBL), Germany
• Associated to the scholarship: 20/16422-5 - Characterization of the inactivation of autosomal sequences in patients with unbalanced X-autosome translocations, BP.DR

Abstract

X-chromosome inactivation (XCI) is the epigenetic mechanism through which one of the two X chromosomes in females is silenced, allowing for a balanced expression of X-linked genes and dosage compensation between XX and XY individuals. This process is mediated by Xist, a long non-coding RNA that coats the chromosome that will be silenced in cis and recruits several proteins involved in chromosome-wide gene repression. Although XCI is a chromosome-wide phenomenon, a subset of X-linked genes referred to as 'escapees' remain actively expressed from both the active (Xa) and inactive (Xi) X chromosomes in female cells. Despite XCI and Xist-mediated silencing being extensively studied, the exact molecular mechanisms underlying chromosome-wide gene silencing and escape regulation are yet to be elucidated. In this project, we aim to provide novel insights into the mechanisms underlying silencing and escape regulation during XCI by (1) investigating the role of the Polycomb Repressive Complex PRC1 during initiation of X inactivation using allele-specific genomic approaches such as RNA-seq and CUT&RUN in mouse embryonic stem cells (mESC) and (2) validating a putative network of transcription factors and chromatin players that may be involved in escape regulation using single genes knockout strategies and a targeted single-cell-based CRISPRi screen. Such state-of-the-art epigenetic technologies will provide a unique training opportunity with the acquisition of new molecular biology and bioinformatic skills for the analysis and integration of epigenomic and transcriptomic datasets while advancing the understanding of mechanisms behind silencing and escape using multiple techniques. (AU)