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Browsing School of Physics (ETDs) by Author "Gentle, N ."

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    Chromatin accessibility changes during early monocyte-to-macrophage differentiation
    (University of the Witwatersrand, Johannesburg, 2024) Xu, Yi Fan; Meyer, V.; Gentle, N .
    The differentiation of monocytes into macrophages is a crucial process that enhances the local immune response against infection by recruiting monocytes to local tissues and transforming them into macrophages. The changes in gene expression associated with this process are known to be regulated by various mechanisms, including the chromatin accessibility landscape. Previous in vitro studies have shown that promonocytic THP-1 cells can differentiate into macrophage-like cells following treatment with phorbol 12-myristate 13-acetate (PMA). While previous studies have attempted to track the differentiation process over time, there has been a lack of research specifically focusing on earlier time points. Therefore, in this study, we used various publicly available RNA-seq, ATAC-seq and ChIP-seq datasets to describe the early events involved in monocyte-to-macrophage differentiation, using THP-1 cells treated with 100 ng/ml PMA for 24 hours as the model system. ATAC-seq data were aligned to the reference human genome (GRCh38) using Bowtie2 and chromatin accessibility peaks were identified using HMMRATAC. Differentially accessible chromatin regions (|L2FC| > 2; FDR < 0.05) were identified using DiffBind, and were annotated based on their cis-regulatory features. These included promoter regions (based on the GENCODE v40 annotations of the human genome) and THP-1-specific enhancers (defined as known enhancers within the GeneHancer database with an overlapping, THP-1-specific, H3K27ac mark). These cis-regulatory features were then associated with genes found to be significantly differentially expressed in response to PMA treatment (|L2FC| > 2; p.adj < 0.05), following quantification of gene expression using Salmon and differential gene expression analysis using DESeq2. The results of this study revealed that the early response to PMA in THP-1 cells is linked to changes in both gene expression and chromatin accessibility. These changes in both gene expression and chromatin accessibility were shown to be linked with inflammatory responses and cell migration activities. Although there was only a limited association between changes in gene expression and chromatin accessibility at the 24-hour time point, opening of chromatin at promoter and enhancer regions and increased gene expression was observed for many genes previously reported to be involved in the process of monocyte-to-macrophage differentiation, including CSF1, CSF1R, and IL-1α/β. This suggests that changes in chromatin accessibility at cis-regulatory elements taking place early in the differentiation process drive the changes in gene expression necessary for monocyte-to-macrophage differentiation