Transcriptomisc Reveals Myometrial Topologically Associated Domains linked to the Onset of Human Term Labor

Changes in cell phenotype are thought to occur through the expression of groups of co-regulated genes within Topologically Associated Domains (TADs). In this paper we locate genes expressed within the myometrium of the human uterus during the onset of term labor into TADs. Transformation of the myometrial cells of the uterus into a contractile phenotype during term human labor is the result of a complex interaction of different epigenomic and genomic layers. Recent work suggests that the transcription factor RelA lies at the top of this regulatory network. Using deep RNAseq analysis of myometrial samples obtained at term from women undergoing caesarean section prior to or after the onset of labor we have identified evidence for how other gene expression regulatory elements interact with transcription factors in the labor phenotype transition. Gene set enrichment analysis of our RNAseq data identified three modules of enriched genes (M1, M2 and M3) which in gene ontology studies are linked to matrix degradation, smooth muscle and immune gene signatures. These genes were predominantly located within chromosomal TADs suggesting co-regulation of expression. Our transcriptomic analysis also identified significant differences in the expression of long-non-coding RNAs (lncRNA), microRNAs (miRNA) and transcription factors that were predicted to target genes within the TADs. Additionally, network analysis revealed 14 new lncRNA (MCM3AP-AS1, TUG1, MIR29B2CHG, HCG18, LINC00963, KCNQ1OT1, NEAT1, HELLPAR, SNHG16, NUTM2B-AS1, MALAT1, PSMA3-AS1, GABPB1-AS1, NORAD, NKILA) and 4 miRNA (mir-145, mir-223, mir-let-7a, mir-132) as top gene hubs with 4 transcription factors (NFKB1, RELA, ESR1) as master regulators. Together, these factors are likely to be involved in co-regulatory networks driving a myometrial transformation to generate an estrogen sensitive phenotype. We conclude that lncRNA and miRNA targeting the ESR1 and NFKB pathways play a key role in the initiation of human labor. For the first time we perform an integrative analysis to present a multi-level genomic signature in the myometrium for spontaneous term labor. The samples were collected under our University of Newcastle John Hunter Hospital, Newcastle, Australia and the Singapore KK Women’s and Children’s Hospital ethics committees (Ethics approval H3820602). Pregnant women gave written informed consent to donate a biopsy of their myometrium at Caesarean section at term according to the institutional guidelines and regulations. All women were between 36 completed weeks and 40 weeks gestational age. Tissues were sampled using a surgical technique standardized such that the bladder is reflected off the lower segment of the uterus (the uterovesical-fold) and the uterine incision is made in the upper part of the lower segment. Candidates for whole transcriptome analysis using RNAseq were chosen from the two distal, ie non-laboring (n=5), and laboring (n=7) groups as well as women clinically classified as laboring from the middle band to represent mothers in early labor (n=5).