Mechanisms controlling human microglia gene expression

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Abstract Microglia are tissue macrophages that reside in the central nervous system (CNS) and perform unique and critical auxiliary functions important to CNS development, homeostasis, immunity and repair. These roles, along with the progressive appreciation that microglia can contribute to neurological disease processes, provides a compelling case to more clearly understand the mechanisms that regulate their development and functions. Major unanswered questions include determining the combination of signals within the brain that trigger the differentiation of erythromyeloid progenitor (EMP) cells to become mature microglia and how alterations in these signals specify distinct microglia phenotypes in health and disease. Studies performed under the support of this grant for the past four years provide the foundations for addressing these questions. Four Specific Aims are proposed. Specific Aim 1 is to define expression and chromatin Quantitative Trait Loci and collaborative transcription factors in human microglia. These studies will generate a valuable resource for the neuroscience community and inform studies in Aims 2, 3 and 4. Specific Aim 2 is to define cis regulatory elements that mediate brain environment-dependent regulation of microglia gene expression, focusing on the microglia-specific lineage determining factor SALL1. Importantly, our experimental plan will exploit the recent ability to achieve an in vivo human microglia phenotype within the mouse brain as the context for analysis of the function of environment- dependent enhancers. Specific Aim 3 is to test the hypothesis that brain environment-dependent genes can be activated in iPSC-derived microglia in vitro by conditional expression of environment-dependent transcription factors. Forced expression of these factors in human iPSC-derived microglia in vitro will provide insights into their molecular functions and may enable development of improved in vitro microglia model systems. Specific Aim 4 is to perform in vivo ASO-mediated loss of function experiments to identify transcriptional mediators of brain environmental factors. This aim is based on advances in anti-sense oligonucleotide (ASO) chemistry that now make it possible to use ASOs to significantly alter gene expression in microglia and other cell types of the brain in vivo. In concert, the proposed studies are intended to qualitatively advance understanding of mechanisms that establish the brain environment dependent program of human microglia gene expression.