CircRNA LRIG3 knockdown inhibits the progression of hepatocellular carcinoma by regulating miR-223-3p/MAP2K6 axis and inactivating MAPK/ERK pathway

Abstract

Background: Emerging evidence suggests that circular RNAs (circRNAs) play critical roles in tumorigenesis. However, the roles and molecular mechanism of circRNA leucine-rich repeat immunoglobulin domain-containing protein 3 (circ_LRIG3) in hepatocellular carcinoma (HCC) have not been investigated.Methods: The expression levels of circ_LRIG3, microRNA-223-3p (miR-223-3p), and mitogen-activated protein kinase kinase 6 (MAP2K6) were determined by quantitative real-time polymerase chain reaction (qRT-PCR). Flow cytometry was applied to determine the cell cycle distribution and cell apoptosis. Cell proliferation, migration and invasion were assessed by methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay and transwell assay, respectively. Western blot assay was employed to measure the protein levels of snail, E-cadherin, MAP2K6, mitogen-activated protein kinase (MAPK), phospho-MAPK (p-MAPK), extracellular signal-regulated kinases (ERKs), and phospho-ERKs (p-ERKs). The relationship between miR-223-3p and circ_LRIG3 or MAP2K6 was predicted by bioinformatics tools and verified by dual-luciferase reporter assay. A xenograft tumor model was established to confirm the functions of circ_LRIG3 in vivo.Results: Circ_LRIG3 and MAP2K6 expression were enhanced while miR-223-3p abundance was reduced in HCC tissues and cells. Knockdown of circ_LRIG3 inhibited the progression of HCC cells via reducing cell proliferation, metastasis and increasing apoptosis. MiR-223-3p was a target of circ_LRIG3 and its downregulation reversed the inhibitory effect of circ_LRIG3 knockdown on progression of HCC cells. Moreover, MAP2K6 could bind to miR-223-3p, and MAP2K6 upregulation also abolished the suppressive impact of circ_LRIG3 interference on progression of HCC cells. Additionally, silence of circ_LRIG3 suppressed the activation of MAPK/ERK pathway and tumor growth by upregulating miR-223-3p and downregulating MAP2K6.Conclusion: Circ_LRIG3 knockdown inhibited HCC progression through regulating miR-223-3p/MAP2K6 axis and inactivating MAPK/ERK pathway, providing a potential therapeutic approach for patients with HCC.

Lrig3 restricts the size of the colon stem cell compartment

“…Lrig3 is also regulated at the post-transcriptional level, as there are reports of a circular form of Lrig3 (circRNA Lrig3 ) that is highly expressed in human hepatocellular carcinoma (HCC). Detection of blood plasma circ-Lrig3 is a highly sensitive and specific diagnostic indicator for HCC (23). Mechanistically, downregulation of circRNA Lrig3 represses both the MAPK/ERK and Smad pathways to prevent the progression of HCC and the loss of circRNA Lrig3 suppresses proliferation, invasion of HCC cells in vitro and blocks tumor growth of HCC in vivo (12, 24).…”

“…Using human colon cancer cell lines, Zang and colleagues were able to show that Lrig3 represses metastasis-associated cell motility by inhibiting slug via inactivating ERK signaling (29). While the data on Lrig3 expression in solid tumors is expanding rapidly (5, 6, 12, 17, 23, 24, 29), the role of Lrig3 in the gastrointestinal tract at the genome, transcriptional and protein level is largely unknown and represents a valuable avenue for future research for both normal and cancerous colon tissue.…”

“…Generally, mice, sheep and pig are smaller if they have a loss-of-function in Lrig3 (1, 11, 15). Only recently have contributions to primary literature uncovered mechanistic roles for Lrig3 in the progression of many cancer cell types: cervical, hepatocellular, prostate, glioblastoma and colon (5, 6, 12, 17, 23, 24, 28).…”