BCL-6 and EZH2 cooperate to epigenetically repress anti-inflammatory miR-142-3p/5p in lupus CD4+T cells

Epigenetics plays an important role in the pathogenesis of several autoimmune diseases.^1–3 Indeed, a large body of evidence implicates abnormal T cell DNA methylation patterns in the pathogenies of lupus, a prototypical autoimmune systemic disease.^4,5 Specific DNA methylation marks have been associated with clinical subsets of lupus patients,^6–8 while others, such as methylation levels in apoptosis genes, might explain differences in disease susceptibility between ethnicities.⁹ Furthermore, demethylation of the IFI44L gene locus has recently been described as a novel promising highly sensitive and specific diagnostic tool in lupus.¹⁰

Recent work has suggested that early epigenetic changes in naïve CD4+T cells are associated with disease flares in lupus patients. A proinflammatory epigenetic shift in naïve CD4+T cells, preceding gene expression changes, primes naïve CD4+T cells for Th2/Th17/Tfh differentiation, and opposes inhibitory Treg pathways as the disease becomes more active.¹¹ The key Tfh transcription factor BCL-6 is progressively demethylated as the disease becomes more active in lupus patients.¹¹ This proinflammatory epigenetic shift may be associated with disease flares and is induced by overexpression of the epigenetic modulator EZH2.¹¹ EZH2 is a component of the polycomb repressive complex 2 (PRC2) and mediates trimethylation of histone H3 at lysine 27 (H3K27me3), a repressive epigenetic mark. Both in vitro and in vivo studies support a pathogenic role for EZH2 in lupus.^12,13

Qianjin Lu et al.¹⁴ elegantly demonstrated a novel pathogenic role for BCL-6 in lupus through EZH2-dependent epigenetic suppression of miR-142-3p/5p in CD4+T cells.

Lu and colleagues¹⁵ have previously shown that reduced levels of the microRNAs miR-142-3p/5p in lupus CD4+T cells cause T cell activation and B cell hyperstimulation. The researchers demonstrated that miR-142-3p/5p inhibit signaling lymphocytic activation molecule-associated protein (SAP), CD84, and IL-10 and that inhibiting miR-142-3p/5p in normal CD4+T cells promotes T cell activation and B cell hyperresponsiveness, similar to lupus CD4+T cells.¹⁵ Overexpression of miR-142-3p/5p in lupus CD4+T cells reduced T cell activation and B cell stimulation. Importantly, increased repressive epigenetic marks at miR-142 regulatory sequences in lupus CD4+T cells, including increased levels of H3K27me3, were observed.¹⁵

In a follow-up study published in this issue, Lu and colleagues¹⁴ elucidated the mechanisms regulating miR-142-3p/5p in lupus CD4+T cells. Because previous studies revealed binding of BCL-6 to the miR-142 promoter region in B cells, the researchers investigated the role of BCL-6 in lupus CD4+T cells. They reported significant upregulation of BCL-6 in lupus CD4+T cells and a correlation between BCL-6 levels and anti-dsDNA antibody titers and lupus disease activity as measured by SLEDAI scores. BCL-6 overexpression using plasmid transfection in normal CD4+T cells inhibited miR-142-3p/5p expression, and BCL-6 knockdown in lupus CD4+T cells restored miR-142-3p/5p expression levels. Taken together, these data strongly support a role for BCL-6 overexpression in suppressing miR-142-3p/5p levels in lupus CD4+T cells. Next, Lu and colleagues demonstrated increased recruitment of BCL-6 to miR-142 promoter elements in lupus CD4+T cells and that BCL-6 overexpression is associated with increased levels of repressive histone marks, such as H3K27me3, and reduced levels of histone marks associated with transcriptional accessibility such as histone acetylation (H3K9/K14ac). Furthermore, they showed that BCL-6 recruits EZH2 and HDAC5 to the miR-142 promoter sequence, providing an explanation for the observed increased H3K27me3 levels, decreased H3K9/K14ac levels, and suppression of miR-142-3p/5p expression in lupus CD4+T cells. BCL-6 knockdown in lupus CD4+T cells resulted in a significant reduction in CD40L, IL-21, and ICOS expression and reduced IgG production from autologous B cells in coculture experiments. Taken together, these data indicate that BCL-6 induces a pathogenic effect in lupus CD4+T cells, at least in part, via recruitment of EZH2 and the resulting epigenetic repression of anti-inflammatory microRNAs such as miR-142-3p/5p.

These important data provide novel insights into the role of Tfh cells in lupus, as BCL-6, IL-21, and ICOS are all key regulators involved in Tfh cell differentiation and function. The findings also identify a novel mechanism for a therapeutic approach targeting BCL-6 and/or miR-142-3p/5p in lupus patients. Furthermore, the results of this study emphasize the role of EZH2 in the pathogenesis of lupus. Previous studies showed that EZH2 induces a proinflammatory epigenetic shift and the expression of junctional adhesion molecule A (JAM-A) in lupus CD4+T cells.¹² Blocking EZH2 in vitro inhibited JAM-A overexpression and abnormal lupus CD4+T cell adhesion in vitro, and ameliorated lupus-like disease in animal models.^12,13 The findings by Lu and colleagues help further elucidate the pathogenic role of EZH2 in lupus. Their data demonstrated the recruitment of EZH2 to regulatory elements in miR-142, inducing H3K27me3 and silencing of miR-142-3p/5p, and resulting in increased IL-21 production, ICOS and CD40L overexpression, and B cell stimulation (Fig. 1).

Fig. 1.

Epigenetic repression inhibits miR-142-3p/5p expression in lupus CD4+T cells. The promoter region of miR-142 recruits BCL-6, which is overexpressed in lupus CD4+T cells. BCL-6 recruits EZH2 and HDAC5, resulting in increased H3K27me3 (repressive mark) and decreased H3K9/K14ac (activating marks), respectively. These changes lead to epigenetic repression of miR-142 and reduced miR-142-3p/5p expression in lupus CD4+T cells. MiR-142-3p/5p is anti-inflammatory, and when suppressed, CD4+T cells overexpress inflammatory molecules, including CD40L, ICOS, and IL-21