In the past several years, advances in high-throughput DNA sequencing have enabled massive tumor genome sequencing studies to identify recurrent mutations across many different cancer types and thousands of patients. These mutational surveys have reinforced the paradigm that cancer is a disease of the genome through the identification of inactivating mutations in well studied tumor-suppressors and activating mutations in well studied oncogenes, particularly in adult cancers. A new class of recurrent mutations has emerged as well, inactivating genes that encode chromatin regulators; these are disproportionately prevalent in pediatric and hematological malignancies. The molecular consequences of chromatin regulator mutations on a genome-wide scale, and moreover, how other genetic insults drive chromatin dysregulation and potentially enhance tumorigenesis, were until now completely unknown. In the chapters that follow, we show that a translocation-derived transcription factor chimera in Ewing Sarcoma acquired chromatin modifying activity such that it acts as a pioneer factor, altering chromatin configuration and inducing transcriptional dysregulation. We also demonstrate how alterations in chromatin link aberrancies in transcript processing with histone methyltransferase loss in clear cell Renal Cell Carcinoma through functional studies of chromatin accessibility and RNA processing in primary human tumors. Lastly, we describe how simple modifications to our experimental assay of chromatin accessibility permit the usage of archival (FFPE) human specimens. Together, in addition to contributing a greater understanding of chromatin biology and dysregulation in human cancers, this work will enable large-scale studies of the causes and roles of chromatin dysregulation in other models of human disease. It has also led to the initiation of high-throughput screens for compounds that affect chromatin accessibility, and subsequently tumor cell proliferation. Future work will utilize chromatin accessibility information as a novel clinical diagnostic and prognostic to guide and enhance patient treatment.