hox-loci-focused-crisprsgrna-library-screening-identifying-critical

<em>HOX</em> Loci Focused CRISPR/sgRNA Library Screening Identifying Critical CTCF Boundaries

CCCTC-binding factor (CTCF)-mediated stable topologically associating domains (TADs) play a critical role in constraining interactions of DNA elements ...

University of Florida

 gene dysregulation is a common feature of acute myeloid leukemia (AML). The molecular mechanisms underlying aberrant  gene expression and associated AML pathogenesis remain unclear. The nuclear protein CCCTC-binding factor (CTCF), when bound to insulator sequences, constrains temporal  gene-expression patterns within confined chromatin domains for normal development. Here, we used targeted pooled CRISPR-Cas9-knockout library screening to interrogate the function of CTCF boundaries in the  gene loci. We discovered that the CTCF binding site located between  and  genes (CBS7/9) is critical for establishing and maintaining aberrant  gene expression in AML. Disruption of the CBS7/9 boundary resulted in spreading of repressive H3K27me3 into the posterior active  chromatin domain that subsequently impaired enhancer/promoter chromatin accessibility and disrupted ectopic long-range interactions among the posterior  genes. Consistent with the role of the CBS7/9 boundary in  locus chromatin organization, attenuation of the CBS7/9 boundary function reduced posterior  gene expression and altered myeloid-specific transcriptome profiles important for pathogenesis of myeloid malignancies. Furthermore, heterozygous deletion of the CBS7/9 chromatin boundary in the  locus reduced human leukemic blast burden and enhanced survival of transplanted AML cell xenograft and patient-derived xenograft mouse models. Thus, the CTCF boundary constrains the normal gene-expression program, as well as plays a role in maintaining the oncogenic transcription program for leukemic transformation. The CTCF boundaries may serve as novel therapeutic targets for the treatment of myeloid malignancies.

CTCF boundary remodels chromatin domain and drives aberrant  gene transcription in acute myeloid leukemia.

Arsenic exposure is a worldwide health concern associated with an increased risk of skin, lung and bladder cancer but arsenic trioxide (ATO; AsIII) is also an effective chemotherapeutic agent. The current use of AsIII in chemotherapy is limited to acute promyelocytic leukemia (APL). However, AsIII was suggested as a potential therapy for other cancer types including chronic myeloid leukemia (CML), especially when combined with other drugs. Here, we carried out a genome-wide CRISPR-based approach to identify modulators of AsIII toxicity in K562, a human CML cell line. We found that disruption of KEAP1, the inhibitory partner of the key antioxidant transcriptional factor Nrf2, or TXNDC17, a thioredoxin-like protein markedly increased AsIII tolerance. Loss of aquaporin 3 (AQP3), the zinc transporter ZNT1 (SLC30A1) and its regulator metal regulatory transcription factor 1 (MTF1) also enhanced tolerance to AsIII whereas loss of the multidrug resistance protein ABCC1 increased sensitivity to AsIII. Remarkably, disruption of any of multiple genes, EEFSEC, SECISBP2, SEPHS2, SEPSECS, PSTK, encoding proteins involved in selenocysteine metabolism increased resistance to AsIII. Our data suggest a model in which an intracellular interaction between selenium and AsIII may impact intracellular AsIII levels and toxicity. Together this work revealed a suite of cellular components/processes which modulate the toxicity of AsIII in CML cells. Targeting such processes simultaneously with AsIII treatment could potentiate AsIII in CML therapy.

Article	Year
CTCF boundary remodels chromatin domain and drives aberrant gene transcription in acute myeloid leukemia. Blood\| PubMed ID: 29760161	2018
Functional profiling identifies determinants of arsenic trioxide cellular toxicity. Toxicological sciences : an official journal of the Society of Toxicology\| PubMed ID: 30815697	2019

Amin Sobh

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