Biology subjects

Bassal, M. A.

Publications and source records attributed to Bassal, M. A..

5 recordsLinked to original sources

bioRxiv2020

NAD modulates DNA methylation and cell differentiation

Nutritional intake impacts the human epigenome by directing epigenetic pathways in normal cell development via as yet unknown molecular mechanisms. Consequently, imbalance in the nutritional intake is able to dysregulate the epigenetic profile and drive cells towards malignant transformation. Herein, we present a novel epigenetic effect of the essential nutrient, NAD. We demonstrate that impairment of DNMT1 enzymatic activity by NAD-promoted ADP-ribosylation, leads to demethylation and transcriptional activation of CEBPA gene, suggesting the existence of an unknown NAD-controlled region within the locus. In addition to the molecular events, NAD treated cells exhibit significant morphological and phenotypical changes that correspond to myeloid differentiation. Collectively, these results delineate a novel role for NAD in cell differentiation and indicate novel nutri-epigenetic strategy to regulate and control gene expression in human cells.

molecular biology↗

bioRxiv2020

Myeloid lncRNA LOUP Mediates Opposing Regulatory Effects of RUNX1 and RUNX1-ETO in t(8;21) AML

The mechanism underlying cell type-specific gene induction conferred by ubiquitous transcription factors as well as disruptions caused by their chimeric derivatives in leukemia is not well understood. Here we investigate whether RNAs coordinate with transcription factors to drive myeloid gene transcription. In an integrated genome-wide approach surveying for gene loci exhibiting concurrent RNA- and DNA-interactions with the broadly expressed transcription factor RUNX1, we identified the long noncoding RNA LOUP. This myeloid-specific and polyadenylated lncRNA induces myeloid differentiation and inhibits cell growth, acting as a transcriptional inducer of the myeloid master regulator PU.1. Mechanistically, LOUP recruits RUNX1 to both the PU.1 enhancer and the promoter, leading to the formation of an active chromatin loop. In t(8;21) acute myeloid leukemia, wherein RUNX1 is fused to ETO, the resulting oncogenic fusion protein RUNX1-ETO limits chromatin accessibility at the LOUP locus, causing inhibition of LOUP and PU.1 expression. These findings highlight the important role of the interplay between cell type-specific RNAs and transcription factors as well as their oncogenic derivatives in modulating lineage-gene activation and raise the possibility that RNA regulators of transcription factors represent alternative targets for therapeutic development. KEY POINTSO_LIlncRNA LOUP coordinates with RUNX1 to induces PU.1 long-range transcription, conferring myeloid differentiation and inhibiting cell growth. C_LIO_LIRUNX1-ETO limits chromatin accessibility at the LOUP locus, causing inhibition of LOUP and PU.1 expression in t(8;21) AML. C_LI

cancer biology↗

bioRxiv2020

Targeted intragenic demethylation initiates chromatin rewiring for gene activation

Aberrant DNA methylation in the region surrounding the transcription start site is a hallmark of gene silencing in cancer. Currently approved demethylating agents lack specificity and exhibit high toxicity. Herein we show, using the p16 gene as an example, that targeted demethylation of the promoter-exon 1-intron 1 (PrExI) region initiates an epigenetic wave of local chromatin remodeling and distal long-range interactions, culminating in gene-locus specific activation. Through development of CRISPR-DiR (DNMT1-interacting RNA), in which ad hoc edited guides block methyltransferase activity in a locus-specific fashion, we demonstrate that demethylation is coupled to epigenetic and topological changes. These results suggest the existence of a specialized "demethylation firing center (DFC)" which can be switched on by an adaptable and selective RNA-mediated approach for locus-specific transcriptional activation. One Sentence SummaryLocus demethylation via CRISPR-DiR reshapes chromatin structure and specifically reactivates its cognate gene.

genetics↗

bioRxiv2020

Pseudogene-mediated DNA demethylation leads to oncogene activation

Despite being one of the leading causes of cancer-related deaths, there is an unmet clinical need for hepatocellular carcinoma (HCC) patients. The lack of effective treatment is, at least in part, due to our lack of understanding of the molecular pathogenesis of this disease. Oncofetal protein SALL4 is re-activated in patients with aggressive HCC along with other solid tumors and hematologic malignancies. This study identifies a previously unrecognized mechanism of SALL4 reactivation which is mediated by pseudogene-induced demethylation. Using a locus-specific demethylating technology, we identified the critical CpG region for SALL4 expression. We showed that SALL4 pseudogene 5 hypomethylates this region through interaction with DNMT1, resulting in SALL4 upregulation. Intriguingly, pseudogene 5 is significantly upregulated in a hepatitis B virus (HBV) model prior to SALL4 induction, and both are increased in HBV-HCC patients. Our results suggest that pseudogene-mediated demethylation represents a unique mechanism of oncogene activation in cancer. SignificanceOur study provides a mechanistic link between HBV infection, activation of the oncogene SALL4, and HCC. We reveal a previously undescribed capability of a pseudogene to epigenetically activate an oncogene by demethylation in a locus-specific manner.

cancer biology↗

bioRxiv2020

Zinc finger protein SALL4 functions through an AT-rich motif to regulate heterochromatin formation

SummaryThe zinc finger transcription factor SALL4 is highly expressed in embryonic stem cells, down-regulated in most adult tissues, but reactivated in many aggressive cancers. This unique expression pattern makes SALL4 an attractive target for designing therapeutic strategies. However, whether SALL4 binds DNA directly to regulate gene expression is unclear and many of its targets in cancer cells remain elusive. Here, through an unbiased screen of protein binding microarray (PBM) and Cleavage Under Targets and Release Using Nuclease (CUT&RUN) experiments, we identified and validated the DNA binding domain of SALL4 and its consensus binding sequence. Combined with RNA-seq analyses after SALL4 knockdown, we discovered hundreds of new SALL4 target genes that it directly regulates in aggressive liver cancer cells, including genes encoding a family of Histone 3 Lysine 9-specific Demethylases (KDMs). Taken together, these results elucidated the mechanism of SALL4 DNA binding and revealed novel pathways and molecules to target in SALL4-dependent tumors.Competing Interest StatementThe authors have declared no competing interest.View Full Text

molecular biology↗