br caption on next page
(caption on next page)
Fig. 4. Identification of enriched miRNA Prostaglandin E2 from mRNA-miRNA and lncRNA-miRNA interaction networks and stage specific mRNA-miRNA-lncRNA association network generation. miRNA-mRNA interaction networks are being depicted for miRNA family enrichment of validated interacting miRNA partners of (4a) mRNAs selected from microarray, (4b) mRNAs selected from TCGA and (4c) a selected lncRNA LINC00263. Panel d shows Venn diagram of enriched miRNA candidates of let-7 family. Stage I specific mRNA-miRNA-lncRNA interaction network is shown in 4e and Stage III specific mRNA-miRNA-lncRNA interaction network is shown in 4f. During stage specific mRNA-miRNA-lncRNA triad generation, for experimentally valid interaction in some CC cell lines, miRNAs are shown in orange and mRNAs in pink colour in 4e and for the other valid interactions miRNAs were shown in yellow, while mRNAs were depicted in blue. For all conditions, lncRNAs were depicted in red. In 4a, enriched miRNA families were shown in specific colors like miR-30 were shown in yellow, miR-17 in green, let-7 in pink and miR-130 in cyan. In panels a and b, red color shows mRNA and blue depicts miRNAs. In 4b and 4c, green points depict candidates of let-7 family miRNA, while in 4c, red points depict the lncRNA, LINC000263. (For interpretation of the references to color in this figure, the reader is referred to the web version of this article.)
mRNA selection from microarray, LAMP3, was also found to be a sur-vival marker at early stages and its upregulation also promoted CC metastasis (Kanao et al., 2005). GAGE5, GAGE2B and GAGE4 are found important in our study during mRNA selection from stage II vs stage III, but a previous study showed that GAGE family genes like GAGE-3/6 were upregulated in CC in late stages (Sarcevic et al., 2003). Emergence of downregulated AKR1C2 as an important mRNA found during stage
III vs stage IV based mRNA selection from microarray in this study may be biologically relevant as HPV16 E7 transactivation on aldo-keto
The results showing the significance of electron transport chain and oxidative phosphorylation pathway in lncRNA associated pathway
Fig. 5. Pathway, gene ontology and DSigDB Analysis of selected RNAs.
Bar plots of (5a) Reactome 2016 (5b) GO Biological Process 2018 (5c) GO Cellular Component 2018 and (5d) DSigDB analysis of selected mRNAs and plot for (5e) Wiki-pathway enrichment of selected lncRNAs and their co-expressed mRNAs.
S. Banerjee and D. Karunagaran
analysis correlate well with E6-mediated activation of genes associated with those pathways (Evans et al., 2016). Emergence of Hs Proteasome Degradation pathway can also be biologically validated as E6 in-activates p53 through proteasomal degradation in CC too (Tomaić, 2016). Oxidative phosphorylation was also found to be the preferential ATP production mechanism from glutamine in HeLa cells (Jose et al., 2011). Receptor tyrosine kinase signaling pathway is another lncRNA associated pathway obtained in the study that was found to be pro-longed due to E6 (Spangle and Munger, 2013). HPV oncogenes also dysregulated the associated collagen and ECM dynamics via transcrip-tional regulation independently and synergistically with estrogen (Spurgeon et al., 2017). Enhanced EGFR internalization by E6 of HPV16 and mTORC1 activation supports the role of HPV in lncRNA mediated
CC progression during pathway identification (Spangle and Munger, 2013). It may be noted from our results that the pathways enriched by selected lncRNAs are mainly modulated by HPV oncogenes, recognized to be the major cause for CC. As many selected mRNAs in this study were found to be hypermethylated like HBB, HBA2, SLC2A1, S100A7, CXCL10 etc., emergence of Decitabine, a DNA hypomethylating agent, beside another convention chemotherapeutic drug 5-Fluorouracil during DSigDB analysis supports the biological significance of this analysis, since E6 and E7 modulate the expression of many proteins by epigenetic regulation (Durzynska et al., 2017). In metastatic CC, Cis-platin- decitabine combination is in Phase II clinical trial (Pohlmann et al., 2002).