Understanding corepressor complex function in the epigenetic regulation of macrophage gene expression

<p dir="ltr">Macrophages play a major role in chronic inflammatory diseases such as type 2 diabetes, atherosclerosis, and cancer. Transcription factors, chromatin modifiers, and other coregulators coordinate inflammation and metabolism in macrophages through epigenetic mechanisms of transcriptional repression and activation. Among these, the HDAC3 corepressor complex is a key regulator of macrophage function. This thesis focuses on three of its subunits: G protein pathway suppressor 2 (GPS2), nuclear receptor corepressor (NCOR), and silencing mediator of retinoic acid and thyroid hormone receptor (SMRT). While these subunits together form the so-called core corepressor complex, the common functions of the core complex and the distinct roles of the subcomplexes that they form have remained poorly understood. Therefore, the overall aim of this thesis was to identify both common and distinct roles of these complex subunits in regulating gene expression in macrophages, and to uncover the mechanisms underlying their context-specific functions.</p><p dir="ltr">Previous studies have shown that GPS2, in a subcomplex with SMRT, acts as an anti-inflammatory chromatin-modifying coregulator in classically activated (M1) macrophages (1,2). In Study I, we investigated the role of GPS2 in alternatively activated (M2) macrophages. To explore this, we primarily used the mouse macrophage cell line RAW264.7 and bone marrow-derived macrophages depleted of GPS2, NCOR, or SMRT, and applied transcriptomic, cistromic, epigenomic, and DNA topology analyses. This study revealed that GPS2 is critical for the epigenetic regulation of M2 macrophage activation, as GPS2 antagonizes enhancer activation mediated by the lysine demethylase KDM1A. Importantly, this regulatory role of GPS2 was found to be dependent on its cooperation with NCOR and SMRT, highlighting a shared function of the corepressor complex in controlling enhancer accessibility and transcriptional regulation in M2 macrophages.</p><p dir="ltr">NCOR and SMRT are homologous subunits of the corepressor complex, yet previous research has shown that their depletion results in distinct phenotypes. However, their individual roles leading to these distinct outcomes have not been systematically investigated in macrophages. In Study II, we focused on identifying the shared and unique genome-wide roles of NCOR and SMRT in RAW264.7 cells and bone marrow-derived macrophages, using integrative analysis of cistrome, epigenome, and transcriptome data. NCOR depletion primarily affected metabolic gene programs, whereas SMRT depletion had a stronger impact on inflammation gene regulation. Remarkably, these alterations included selective changes in chromatin accessibility and H3K27ac signatures, with SMRT depletion exerting a dominant effect. These differences could not be attributed to differential chromatin recruitment, as NCOR and SMRT exhibit genome-wide co-occupancy in macrophages. Instead, cistrome analyses in NCOR- and SMRT-depleted macrophages revealed a functional hierarchy between the two corepressors. SMRT appears to play a predominant role, as its depletion abolishes chromatin binding of both NCOR and GPS2, and even seems to induce translocation of the entire core corepressor complex to the cytoplasm. In contrast, NCOR depletion has a more limited effect on both SMRT and GPS2, primarily altering SMRT binding preferences to distinct cis-regulatory elements and reducing GPS2 chromatin binding.</p><p dir="ltr">Overall, this thesis provides insights into the role of the corepressor complex subunits GPS2, NCOR, and SMRT in the epigenetic regulation of inflammation- and metabolism-related gene expression in macrophages. These findings may contribute to future strategies for therapeutic targeting of macrophage epigenetic states in chronic diseases such as diabetes, atherosclerosis, and cancer.</p><h3>List of scientific papers</h3><p dir="ltr">I. Antagonistic action of GPS2 and KDM1A at enhancers governs alternative macrophage activation by interleukin 4. Zhiqiang Huang, <b>Astradeni Efthymiadou</b>, Ning Liang, Rongrong Fan, Eckardt Treuter. Nucleic Acids Research; 2023 Feb 22; Volume 51, Issue 3, Pages 1067-1086, PMID: 36610795. <a href="https://doi.org/10.1093/nar/gkac1230 " rel="noreferrer" target="_blank">https://doi.org/10.1093/nar/gkac1230 </a></p><p dir="ltr">II. SMRT anchors the HDAC3 corepressor complex to chromatin to regulate inflammatory and metabolic pathways in macrophages<b>. </b><b>Astradeni Efthymiadou</b>, Chaode Gu, Cheng Wang, Hongwei Wang, Ziyi Li, Oihane Garcia-Irigoyen, Rongrong Fan, Eckardt Treuter, Zhiqiang Huang. Nucleic Acids Research; 2025 Sep 23; Volume 53, Issue 17, gkaf880, PMID: 40966497. <a href="https://doi.org/10.1093/nar/gkaf880 " rel="noreferrer" target="_blank">https://doi.org/10.1093/nar/gkaf880 </a></p>