JIAN XU LABORATORY

Enhancer Biology in Development and Cancer

A fundamental question in biology is how gene expression programs are established and maintained to preserve or alter cell identity. This question is intimately related to how transcription factors interact with the appropriate gene-specific cis-regulatory elements within chromatin.

Enhancers are cis-acting DNA sequences that determine cell identity by regulating when, where and how genes are expressed. Despite their importance, the regulatory components controlling the structure and function of enhancers during lineage development and disease pathogenesis remain largely unknown.

We recently developed the CRISPR/Cas9-based ‘CAPTURE’ system to unbiasedly identify chromatin interactions that regulate non-coding genomic elements including enhancers. Using biotinylated dCas9 and programmable sgRNAs, we were able to selectively isolate the native genomic locus-associated protein complexes and 3D chromatin structures (Liu et al., 2017). The CAPTURE system provides a ‘first-in-kind’ approach to simultaneously identify cis-element-regulating proteins and DNA structures, and compare the results to establish the causality of gene expression during development and disorders.

We also developed dCas9-based enhancer-targeting epigenetic perturbation systems, enCRISPRi and enCRISPRa, for high throughput analyses of enhancer function in lineage differentiation and cancer pathogenesis (Li et al., 2020). Building on these discoveries, it is our long-term goal to demystify the cis-regulatory logics as fundamental principles that control stem cell development and disorders.

 Ongoing studies in our lab include:

• Elucidating the molecular determinants of lineage-specifying enhancers
• Functional and mechanistic analysis of pathogenic non-coding variants
• Developing genomic and epigenomic editing technologies in dissecting genome structure and function

Epigenetics and Metabolic Control of Hematopoiesis and Leukemia

How epigenetics and metabolism cooperate to control normal development and disease processes is a fundamentally important question with significant clinical implication. Many epigenetic enzymes catalyzing DNA or histone modifications are susceptible to changes in co-substrates of cellular metabolism, but little is known about whether and how altered epigenetics influences metabolism during cancer progression.

We recently described that inactivation of the histone methyltransferase EZH2 promotes leukemic transformation by aberrant activation of branched-chain amino acid (BCAA) metabolism in leukemia-initiating cells, establishing a new molecular link between altered epigenetics and metabolism in cancer progression (Gu et al., 2019). This study was among the first to show that epigenetic alterations rewire intracellular metabolism during cancer transformation, causing epigenetic and metabolic vulnerabilities in cancer-initiating cells.

In other studies, we compared the proteomic and transcriptomic changes in human primary hematopoietic stem/progenitor cells and erythroid cells, and uncovered pathways related to mitochondrial biogenesis enhanced through protein translation, establishing a new mechanism for post-transcriptional control of mitochondria related to hematologic defects in mitochondrial diseases and aging (Liu et al., 2017).

Building on these findings, we are examining the epigenetic and metabolic liabilities of blood stem cells in development and pathological conditions. Ongoing studies include:

• BCAA metabolism in stem cell function and cancer development
• Metabolic control of normal and pathological erythropoiesis
• Crosstalk between epigenetic gene regulation and intracellular metabolism in hematopoiesis and leukemia

Non-Coding Genome in Normal and Cancer Stem Cells

Advances in genome sequencing are poised for applications in personalized medicine. However, current understanding of disease-causing genetic alterations is largely based on protein-coding DNA sequences consisting of only ~1% of human genome. It remains unclear how alterations within non-coding genome, consisting of various regulatory elements and mobile DNA sequences, contribute to disease pathophysiology. Similarly, genotype-phenotype association studies continue to elucidate non-coding genomic regions that are altered in human diseases, although identification of causal elements remains challenging impeding drug development and therapeutics.

We aim to develop experimental and computational methodologies, and integrate with in vitro and in vivo disease modeling towards a systems-level view of disease-associated non-coding genetic elements in development and diseases. This is possible by mapping genetic and epigenetic events that discriminate the normal and neoplastic genomes, coupling epigenetic changes with genetic lesions and gene expression programs, and conducting mechanistic studies of individual candidates in disease models.

By comparing the regulatory composition of gene regulatory networks in normal and neoplastic hematopoiesis, we aim to investigate how non-coding regulatory genome, lineage-specifying regulators, epigenetic modulators and environmental signals cooperate to control lineage specification, and how dysregulation of these mechanisms contribute to cancer development.

Ongoing studies in our lab include:

• Oncogenic cooperation between coding and non-coding variants in cancer pathophysiology
• Integrative analysis of genomic structural variants as cancer drivers
• Roles and rules of genomic transposable elements in hematopoiesis and leukemia