Understanding the regulatory mechanisms governing hematopoiesis and how it is dysregulated in malignancy requires integrative approaches spanning multiple modalities. This dissertation presents three complementary strategies to understand cellular regulation: (1) single-cell CRISPR-based perturbation of post-transcriptional regulators to study alternative polyadenylation (APA), (2) single-cell multiomic profiling of spleens from patients with myelofibrosis to uncover the landscape of malignant hematopoiesis and its interaction with the microenvironment, and (3) ex vivo perturbation of transcription factors in primary hematopoietic stem and progenitor cells to understand how chromatin remodeling dictates lineage commitment. Together, these approaches span cellular regulation from chromatin accessibility to RNA processing and highlight how cell identity and function are controlled at multiple levels. Our findings reveal that alternative polyadenylation is not globally regulated but occurs in coordinated modules of transcripts dictated by sequence content and other functional characteristics, which has important implications for how post-transcriptional regulation affects cellular state. We identify malignant hematopoiesis in spleens of patients with myelofibrosis, which exhibits substantial lineage bias and massive increases in inflammation that alter interactions with the splenic microenvironment. Through transcription factor perturbations in hematopoietic stem and progenitor cells, we identify chromatin signatures that underpin how fate decisions are made and can be leveraged to interpret patient-specific disease states. Collectively, this work demonstrates the power of single-cell studies spanning multiple modalities and in diverse contexts as elements of a broader framework to understand both normal and malignant hematopoiesis.