PhD Student The Hebrew University of Jerusalem Jerusalem, Yerushalayim, Israel
Abstract: Haploinsufficiency describes a phenomenon where only one functioning allele of a gene is insufficient for a normal phenotype of a diploid cell/organism. Although haploinsufficiency underlies several human diseases, the effect of haploinsufficiency on human embryogenesis is largely unknown. Here, we aimed to identify genes affecting the normal growth of human embryonic stem cells (hESCs) when one of their two alleles is lost. To establish a genome-wide loss-of-function screening for heterozygous mutations, we fused normal haploid cells with a library of mutant haploid hESCs. We have identified over 600 genes with a negative effect on hESC growth in a haploinsufficient manner and characterized them as genes depleted from telomeres and X chromosome. Interestingly, a large fraction of these haploinsufficiency genes is associated with the extra-cellular matrix and the plasma membrane. We have revealed an enrichment of genes causing haploinsufficiency disorders within WNT and TGF-b signal transduction pathways. We could thus identify haploinsufficiency-related genes and pathways that show growth retardation in early embryonic cells, suggesting dosage-dependent phenotypes in hESCs. There are several neurodegenerative disorders affected by haploinsufficiency phenomenon, and many of them are related to autism. We differentiated the library of heterozygote mutations into neural progenitor cells (NPCs) and identified about 250 genes essential for their differentiation in a haploinsufficient manner. We were able to identify neuronal-related dosage-sensitive pathways and detected several autism-related genes with a haploinsufficiency effect, suggesting that they can be modelled at early stages of differentiation. By using small molecule activators of haploinsufficient pathways in the heterozygous mutated cells, we were able to see improvements in some of the pathological phenotypes, thus affecting the molecular damages caused by haploinsufficiency in autism-related genes, and creating opportunities for future therapeutic research. Overall, we have constructed a novel model system for studying haploinsufficiency in both hESCs and NPCs, and thus were able to characterize important dosage-dependent genes and pathways involved in their normal growth and pathology.
