MDMs derived from 1 healthy individual were divided into three parts: the control cultured with medium, the stimulated cells cultured with LPS, and the rescued cells pretreated with 30 nM THZ1 for 0.5 h prior to LPS stimulation. Total RNA was prepared as described in the quantitative real-time PCR experiments. The isolated RNA was used to prepare RNA-seq libraries using TruSeq RNA Library Prep Kit v2 (Illumina) following the manufacturer’s instruction. The libraries were sequenced on an Illumina HiSeq X-ten sequencing System with a read length of 150 base pairs.

Raw RNA-seq reads were first preprocessed the in-house Perl scripts and sickle software (version, 1.200). The preprocessed reads were aligned against the human genome (GRCH38, https://asia.ensembl.org/info/data/ftp/index.html) using HISAT2 (v2.1.0) [22]. Gene-level read counts were summarized with HTSeq script (version 0.6.0) [23]. Genes with more than 50 reads in at least one library were retained and used to identify DEGs with the Bioconductor package DEGseq [24]. A DEG met the following criteria: average expression abundance of the gene at least in one sample more than 1 Fragments per Kilobase Million (FPKM), false discovery rate (FDR, an adjusted P value after multiple testing of Benjamini-Hochberg [25]) < 0.01 and abs (fold change) > 2. Heatmaps in Fig. 3a were plotted using heatmap.2 in gplots, and Fig. 3d was created using the Heatmap script in R package ComplexHeatmap.

CDK7 inhibitor suppresses the gene profiles of inflammation via RNA Pol II. a Heatmap of 701 DEGs in MDMs following 8-h LPS stimulation pretreated with 30 nM THZ1 or not. Heatmap displayed the Log2 fold change in gene expression versus vehicle control or LPS-stimulated cells. b The top 10 enriched GO biological processes of 701 LPS-stimulated DEGs and 361 THZ1-sensitive DEGs. Individual bars represent the P value after Benjamini-Hochberg correction for enrichment of GO biological processes. c There were 361 THZ1-sensitive DEGs enriched using gene set enrichment analysis. d Heatmaps of DEGs from the top 3 inhibited GO molecular functions and the TFs described in e. e Involvement network between upregulated TFs and biological processes. Only the relatively central TFs involved in more than 3 GO terms are shown. f Quantitative RT-PCR analysis of partial TFs described in e. g Peak plot and heatmap of the RNA Pol II ChIP-seq density around the transcription start sites (TSS) and transcription end sites (TES) based on 361 THZ1-sensitive DEGs in control mTHP-1 (gray) and LPS-stimulated mTHP-1 pretreated with THZ1 (blue) or not (red). h Gene tracks of RNA Pol II binding density at the representative gene loci after treatment as in g. Data are the mean ± SD, n = 3 in f

The DAVID suite of online tools (http://david.abcc.ncifcrf.gov/tools.jsp) was used to determine the biological process ontology defined by the Gene Ontology Consortium. Gene Set Enrichment Analysis was performed using a gene list pre-ranked by fold change upon LPS treatment from mTHP-1 cells. The network in Fig. 3e was created using the functional Network in R package FGNet.

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