PROJECT ABSTRACT We propose to use single cell studies of clinical isolates to explicate how mutations and differentiation status coordinately regulate cell state, clonal expansion and malignant transformation. We will model sequential activation/inactivation of somatic mutations in defined cell compartments and perform functional studies to uncover critical gene networks and therapeutic dependencies. This will include single cell studies in primary patient samples, use of innovative models which allow for sequential mutational activation/inactivation, and functional genomic approaches to delineate mechanisms of transformation and novel therapeutic dependencies. Moreover, we will delineate crosstalk between different cell types in normal and malignant hematopoiesis and how these interactions impact therapeutic response. These tools will be distributed widely to enable studies of tissue development/homeostasis, cell state changes, transformation, and therapeutic dependencies.