PROJECT SUMMARY Research Plan: An impaired hematopoietic differentiation process underlies bone marrow malignancies like leukemia, but we still lack the mechanistic understanding of the sequence of regulatory events that misleads the differentiation process. Since epigenomic regulatory patterns are major features of leukemic development, understanding the chromatin dynamics of a failed (malignant) hematopoietic differentiation process can help define the molecular basis of leukemia. A prerequisite to such an understanding is a framework that allows investigation of the progressive changes in the activity of the regulatory elements (RE) during hematopoietic differentiation. Single-cell CUT&Tag (scCUT&Tag) technology is well-suited for such studies as RE activity through histone modification profiles can be investigated in a lineage-specific manner. However, poor understanding of the cell-type-specific histone modification patterns makes the task challenging. To overcome this challenge, we designed scCUT&Tag-pro which allows simultaneous measurement of cell-surface protein and in-silico integration of gene-expression and chromatin accessibility. I will leverage this novel multimodal framework to investigate the RE and progressive changes in their activity during hematopoiesis. First, I will define a multimodal reference mapping framework for mouse hematopoiesis. This framework will allow me to integrate multiple histone modification profiles onto one reference and compare the chromatin states of the RE between a wild type (WT) and mouse model with loss of function in histone methyl transferase (HMT) (Aim 1). Second, since HMTs regulate transcription through the interaction network of RE. I will define a chromatin state aware map that dynamically links REs across developmental trajectories. I will use this framework to investigate the changes in the interaction of REs due to HMT loss (Aim 2). Third, since the transcriptional state of a cell emerges from the underlying gene regulatory network (GRN), I will integrate single-cell gene expression data with histone modification profiles and extend it to define a chromatin state aware model of GRN. I will compare the WT and HMT loss experiments and define the differential GRN (Aim 3). Altogether, this research proposal seeks to pioneer the computational methods for the integrated analyses of multimodal single-cell histone modifications and systematically dissect progressive changes in the system-level function of the regulatory circuits that misleads hematopoietic differentiation using mouse models with conditional HMT loss of function in the hematopoietic compartment. I have developed a 5-year career development plan to meet my goal of becoming an independent investigator in the multi-disciplinary field of computational cancer biology. The mentorship committee will also provide me the guidance in my research and academic job search. Given the excellent the outstanding record of training multiple independent scientists, New York Genome Center provides me an ideal environment to attain my scientific career goals.

PROJECT NARRATIVE An impaired hematopoietic differentiation process underlies bone marrow malignancies like leukemia, but we still lack the mechanistic understanding of the sequence of regulatory events that misleads the differentiation process. Understanding the chromatin dynamics of a failed (malignant) hematopoietic differentiation process, in comparison to a healthy process, can define the molecular basis of leukemia. This research proposal seeks to develop a suite of computational tools for the multimodal integration of epigenomic data and use the framework to mechanistically investigate the corrupted hematopoietic differentiation process through mouse models of leukemia.