In this proposal, we will address a fundamentally important problem in glia biology: how do oligodendrocyte precursor cells (OPCs) robustly maintain their numbers? Previous intravital imaging experiments demonstrated that OPCs exert mutual inhibition when they contact each other, but promptly enter cell cycle when neighboring OPCs either differentiate or die, and halt proliferation again when homeostasis is achieved. However, the molecular basis behind OPC homeostasis remains largely unknown. Recently, using a mouse genetic system called Mosaic Analysis of Double Markers (MADM) to model glioma, our lab discovered that OPC is a cell of origin for glioma, and revealed that, instead of passively over-expanding, mutant OPCs actually outcompete WT OPCs and eventually take over the entire brain. Most importantly, when we genetically blocked cell competition of mutant OPCs, glioma can be completely prevented. Putting the observations of OPC homeostasis in health and OPC competition in gliomagenesis together, we realized that these are the two sides of the same coin, and that there is a Yin/Yang mechanism for OPC proliferation that counterbalances each other in health but gets deregulated in cancer. While the Yang network of RTK signaling is well studied, the Yin network is much less understood, let alone their counter-interactions. Here, we hypothesize that phospho-proteomic/proteomic profiling and subsequent candidate validation using highly sensitive OPC competition platforms should enable us to provide mechanistic insights into this Yin/Yang network. To test this hypothesis, we have assembled a team of experts on OPC purification and culture, proteomic profiling with limited materials, and using advanced statistical and unsupervised learning approaches to predict signaling network based on phospho-proteomic/proteomic profiles. As a team, we have successfully performed a pilot experiment that led to a handful of candidate genes. In Aim 1 of this proposal, we will validate the role of these candidate genes in OPC competition. In Aim 2 of this proposal, we will perform further in-depth profiling experiments to gain a comprehensive insight into the signaling network that controls OPC homeostasis and competition. The findings from our project should motivate further functional studies to clearly delineate the entire pathway, deepen our understanding of OPC homeostasis, and shed light on paradigm-shifting therapeutic strategies for glioma based on the concept of OPC competition.