Chromodomain helicase DNA binding protein CHD2 interacts with TET2 and regulates lineage defining transcription in hematopoietic stem and progenitor cells

Abstract Introduction. TET2 mutations (TET2MT) are among the most frequent somatic lesions in myeloid malignancies, found at ~30% in MDS/AML and over 66% in CMML. TET2MT is also common in clonal hematopoiesis of indeterminate potential (CHIP), a pre-malignant condition that increases the risk of myeloid neoplasms (MN) and other diseases including treatment response in certain solid tumors. However, TET2MT alone is insufficient for malignant transformation, as they are frequently observed in CHIP without apparent disease. Thus, strongly suggesting the presence of additional mechanisms of lineage perturbation and evolution to MN. Chromodomain Helicase DNA-binding Protein 2 (CHD2) is an ATP-dependent DNA helicase that modulates gene expression via nucleosome repositioning. Despite its essential role in transcriptional fidelity and genomic integrity, CHD2 function remains poorly characterized in hematopoiesis and leukemogenesis. Here we report that CHD2 directly interacts with TET2 and co-occupies chromatin regulatory sites to maintain epigenetic homeostatic hematopoiesis. In the absence of TET2, CHD2 is co-opted to sustain oncogenic transcriptional programs that skew differentiation to myeloid precursors and prevent differentiation seen in TET2MT AML. Methods. CRISPR-Cas9 was used to generate clonal isogenic TET2 knockout (TET2KO) and TET2WT cells derived from K562 and THP1. Endogenous immunoprecipitation (IP) of CHD2 and TET2 was performed to purify associated protein complexes, followed by analysis and identification of the CHD2-TET2 interactome with high performance liquid chromatography coupled with tandem mass spectrometry (LCMS/MS). To investigate functional consequences of TET2 loss on CHD2 activity, we conducted integrative multi-omics analysis combining proteomics, publicly available chromatin immunoprecipitation sequencing (ChIP-seq), and RNA sequencing (RNA-seq). Genome-wide co-occupancy of CHD2 and TET2 was evaluated through comparative analysis of published ChIP-seq datasets. Results. Reciprocal IP of TET2/CHD2 in K562 and CMK cells revealed robust interaction between CHD2-TET2. Global proteomic comparisons using LCMS/MS revealed significant overlap between CHD2 and TET2 interactomes. 268 proteins were unique to CHD2 IPs from TET2WT that reduced to 106 in isogeneic TET2KO cells with CHD2 retaining a conserved overlapping interaction network in both cases. Notably, CHD2 IPs were enriched for components of the CoREST complex (RCOR1, RCOR3, GSE1, KDM1A), suggesting a potential role in transcriptional repression. In contrast, SWI/SNF components were particularly enriched in TET2 IP, highlighting TET2's role as a critical hub for chromatin remodeling. TET2 interactome was markedly broader, comprising over 3,000 proteins, including key regulators of polycomb repressive complexes (PRC1/2) and lineage-defining transcription factors like GATA1, GATA2, RUNX1 and STAT5A/B. Our analysis of ChIP-seq data showed 95% overlap between CHD2- and TET2-bound genes. RNAseq analysis in matched isogeneic cells revealed that these co-targeted genes are differentially expressed in the TET2KO cells. The oncogenic pathways were enriched among genes upregulated in TET2KO, while RNA processing and chromatin homeostasis genes were down regulated compared to TET2WT. ChIP-seq track analysis showed CHD2 and TET2 binding at promoter-proximal nucleosome-depleted regions, supporting a cooperative regulatory mechanism. These findings suggest that in TET2-proficient cells, CHD2 is directed to key regulatory elements to maintain transcriptional fidelity that is essential for homoeostasis hematopoiesis. In TET2-deficient cells, CHD2 is hijacked to sustain aberrant transcription that may promote myeloid skewing and leukemogenesis. Conclusions. For the first time, here we define a previously unrecognized CHD2-TET2 interactome and its potential functional role in the hematopoiesis. This, in part, may provide a probable mechanism of myeloid skewing in TET2MT HSPCs. These findings may provide rational for the dependency of TET2MT clones on CHD2 for malignant evolution. By integrating proteomic, genomic and functional analyses, we elucidate the role of CHD2 in transcriptional control that define lineage fate in the absence of TET2. In addition, we address a significant knowledge gap in CHD2 biology that may form the basis for novel therapeutic approaches targeting epigenetic vulnerabilities in TET2MT associated disorders.