Dsg2-Regulated miRNA and lncRNA Dynamics in Wound Healing and Tumor Development
Abstract
Both Dr. Dvorak and Sir Haddow described cancer as a wound that never heals, and while this comparison has been in place since the late 1900s, the intricate molecular underpinnings shared between these two contexts remain elusive. Previous endeavors have unveiled the multifaceted nature of the cadherin, desmoglein 2 (Dsg2). Dsg2 has been implicated as a key mediator of inflammation, mitogenic signaling, pro-growth and -migratory programming, and has thus emerged as a central player in both cancer and wound repair. miRNA sequencing (miRNA-seq) revealed that Dsg2 expression led to a decrease in miR-146a and a simultaneous rise in miR-155. These miRNAs have been implicated in the regulation of inflammation, but their roles in proliferation and migration have been less well-defined. MiR-146a, often termed the master regulator of inflammation, does so by targeting IRAK1 and TRAF6 in the NFkB pathway, resulting in the reduction of proinflammatory molecules, including the chemokine, interleukin 8 (IL-8). In the course of this research, one avenue of exploration led to the intersection of the Dsg2/miR-146a/IL-8 regulatory axis and the crucial subject of response to immune checkpoint inhibitors (ICIs) in head and neck squamous cell carcinoma (HNSCC). Patient pre-treatment biopsies unveiled an elevation of Dsg2 at baseline in HPV-positive (HPVpos) patients, who would ultimately remain unresponsive to ICI. Delving further, an analysis of patient plasma small extracellular vesicles (sEV) showed that miR-146a was lower in these non-responding HPVpos patients, while IL-8 level was increased. To understand the impact of miR-146a expression in HPVpos HNSCC, in vitro studies using HPVpos HNSCC cells revealed that restoration of miR-146a expression reduced the expression of IL-8 and increased cell death. To better understand the regulation of Dsg2, miRNet.ca and ScanmiRWalk were used to identify miRNAs that could decrease Dsg2 expression. Interestingly, miR-155 was determined to be up-regulated in response to Dsg2 expression. This counterintuitive relationship spurred an exploration of a regulatory mechanism referred to as the double negative hypothesis, involving the long non-coding RNA (lncRNA), Dsg2-antisense 1 (Dsg2-AS1). Dsg2-AS1 emerged as a sponge for miR-155, inhibiting its ability to target Dsg2, resulting in a surge in Dsg2 expression. Through RNA sequencing (RNAseq) and quantitative polymerase chain reaction (qPCR), it was demonstrated that Dsg2 and Dsg2-AS1 are co-expressed. And, using a dual luciferase assay, it was revealed that miR-155 was able to sponge the lncRNA, Dsg2-AS1. Western blot highlighted the increase in Dsg2 expression in response to miR-155 expression in the presence of Dsg2-AS1. Functionally, transwell migration assay and the in vitro scratch assay demonstrated an increase migration in response to miR-155 expression. Using publicly available RNAseq datasets, for both HNSCC and at the leading edge of wounds, it was demonstrated that this regulatory network may be at play. Overall, this study aimed to unveil the active regulatory loop involving miR-146a, Dsg2, miR-155, and Dsg2-AS1, shedding light on the complex and dynamic relationships that shape their interplay within the realms of tissue regeneration and malignant transformation.
Subject Area
Cellular biology|Oncology|Genetics|Molecular biology|Bioinformatics
Recommended Citation
Hill, Brianna Leigh, "Dsg2-Regulated miRNA and lncRNA Dynamics in Wound Healing and Tumor Development" (2024). ProQuest ETD Collection - Thomas Jefferson University. AAI31144058.
https://jdc.jefferson.edu/dissertations/AAI31144058
