Insights into Interleukin-10-activated STAT3 transcriptional function via genome-wide chromatin occupancy and gene expression analysis

Interleukin-10 (IL-10) acts as a potent immune-modulator mainly via simultaneous suppression of pro-inflammatory cytokines expression and induction of anti-inflammatory molecules by activated innate immune cells [1]. Modulation of cytokine production by IL-10 takes place mainly at the transcriptional level [2, 3] and requires STAT3 activation [4, 5]. Furthermore, we have shown that histone acetylation plays a role in IL-10 ability to modulate transcription of LPS-induced IL-1ra and CXCL8 genes [6, 7]. On these bases, we aimed to establish whether chromatin modification represents a general mechanism whereby IL- 10 achieves its modulatory activity on gene transcription. ChIPseq analysis for H3K27acetylation and for STAT3 recruitment was conducted in CD14+ monocytes cultured in the presence of IL-10, LPS, LPS+IL-10 for 60 min or left untreated. Clustering analysis identified ten clusters differentially modulated in the experimental conditions described above. Focusing on the effect of IL-10 on LPS-induced H3K27 acetylation we found 35% of the LPS-induced H3K27ac peaks, preferentially associated with intergenic and intronic region, were inhibited, 5% were potentiated, and 3% were not affected by IL-10. Genome-wide analysis of STAT3 occupancy identified 836 STAT3 peaks induced by IL-10, preferentially located within the gene body. Most importantly, cross analysis of the two ChIP-seq data sets highlighted that: 1) No STAT3 recruitment was observed in genomic region where LPS-induced H3K27ac was not modified by IL-10; 2) 0.8% of the genomic region where LPS-induced H3K27ac was inhibited by IL-10 were found to overlap with STAT3 peaks; 3) 8% of the genomic region where LPS-induced H3K27ac was potentiated by IL-10 were found to overlap with STAT3 peaks. This study provides mechanistic evidence that IL-10 induces covalent histone modification on a genome-wide scale, by promoting or inhibiting histone acetylation. Correlation of ChIP-seq with RNA-seq data will aid in the understanding of IL-10-induced transcriptional regulation of multiple genes.