Study of the effect of mutations in the chromatin modifier gene EHMT2 during neurodevelopment

• Grant number: 24/20118-0
• Support Opportunities: Scholarships in Brazil - Master
• Start date: March 01, 2025
• End date: August 31, 2026
• Field of knowledge: Biological Sciences - Genetics - Human and Medical Genetics
• Principal Investigator: Ana Cristina Victorino Krepischi
• Grantee: Beatriz Cristina de Oliveira
• Host Institution: Instituto de Biociências (IB). Universidade de São Paulo (USP). São Paulo , SP, Brazil
• Associated research grant: 23/09879-7 - Genetic and pathophysiological mechanisms in mendelian neurodevelopmental disorders: investigation of the genes AHDC1 and EHMT2, AP.R

Abstract

Changes in various biological pathways may be involved in the etiology of neurodevelopmental disorders (NDD); however, there is an enrichment of genes related to epigenetic regulation. The EHMT1/2 complex primarily acts in the methylation of lysine 9 of histone H3 (H3K9), which is relevant for key processes in neuronal development. Mutations in EHMT1 that alter the binding/function of EHMT2 cause Kleefstra syndrome type 1, a syndromic form of NDD; however, EHMT2 has not yet been conclusively associated with monogenic pathologies.We identified a case of syndromic NDD with overlapping clinical features of Kleefstra syndrome and a homozygous variant at the canonical splicing donor site of EHMT2. In RNAseq analysis of the patient's blood, only aberrant splicing events were detected, and the DNA methylation profile analysis (EpiSign) revealed an epigenetic signature overlapping with that of Kleefstra 1. Given these findings, we consider EHMT2 a strong candidate for a Kleefstra-like condition.The central objective of this project is to evaluate the EHMT2 gene as a candidate for neurodevelopmental alterations. Specific objectives include developing and characterizing induced pluripotent stem cell (iPSC) and neural stem cell (NSC) lines from the homozygous patient and her heterozygous mother for comparison with controls; evaluating morphological aspects and the protein expression pattern of EHMT2 at two different stages; investigating the effect of the splicing mutation on EHMT2 transcripts in NSC using RNASeq compared to controls, as well as altered genes and biological pathways; quantifying the EHMT2 transcripts resulting from the mutation using high-resolution dissociation curve analysis (in blood and during neurodifferentiation); analyzing the impact of EHMT2 expression on cellular processes such as proliferation, cell cycle, and cell death; and evaluating morphology and cellular processes in control lines treated with the EHMT2 inhibitor (BIX-01294). This study will produce data regarding the function of EHMT2 in neurodevelopment and its potential as a candidate for causing NDD.