Execution of cell fate choice by cooperative gene activities

点击翻译按钮获取中文摘要

Wider research context:Our work on the exit of mouse embryonic stem cells from a naïve to a formative pluripotent identity showed that this model cell-fate decision is controlled by multiple cooperative and partially redundant processes. This redundancy is highlighted by the fact that no single factor mutation, but only dual signalling inhibition or genetic codeletion of multiple factors is sufficient to lock in naïve identity. Hence, genetic approaches focusing on single genes, will certainly miss important redundantly wired functions that control cell fate.Objectives:The central goal of this project is to provide fundamental insights into the processes under redundant genetic control that are crucial for proper developmental progression by completion of two central Aims:Aim 1: To identify redundant and cooperative gene activities during the exit from naïve pluripotency using computational and novel combinatorial screening approaches.Aim 2: To investigate the molecular mechanisms of how cooperative cellular functions enable and execute the exit from naïve pluripotency.Approach:We have developed an innovative high-throughput compatible 3D cell-aggregate screen to identify pairs of interacting genes/processes that act in a cooperative manner. This setup solves the problem of compartmentalizing complex mutation profiles and phenotype, while allowing high throughput. Molecular mechanisms will be revealed through application of genetic, biochemical and network-analysis approaches. These methods will integrate functional genetics data with high-dimensional molecular profiling of knockouts and differentiation time-course information.Level of innovation:Conceptually, the combined activity of two cooperating and partially redundant genes that individually result in only weak phenotypes, but if co-depleted cause a complete differentiation block, is functionally more important than the individual activity of a single gene that exhibits a strong exit-delay upon KO. Hence, we have much to discover and only by starting to dissect complex genetic interactions, we will truly understand cell fate decisions. We propose that our project can reveal entirely unknown regulatory control of differentiation processes which are not accessible due to redundancy in single-factor based genetic approaches. We envision this project to set a standard for performing combinatorial genetics.