MicroRNA-155 Is Regulated by the Transforming Growth Factor β/Smad Pathway and Contributes to Epithelial Cell Plasticity by Targeting RhoA

Abstract

Transforming growth factor ␤ (TGF-␤) signaling facilitates metastasis in advanced malignancy. While a number of protein-encoding genes are known to be involved in this process, information on the role of microRNAs (miRNAs) in TGF-␤-induced cell migration and invasion is still limited. By hybridizing a 515-miRNA oligonucleotide-based microarray library, a total of 28 miRNAs were found to be significantly deregulated in TGF-␤-treated normal murine mammary gland (NMuMG) epithelial cells but not Smad4 knockdown NMuMG cells. Among upregulated miRNAs, miR-155 was the most significantly elevated miRNA. TGF-␤ induces miR-155 expression and promoter activity through Smad4. The knockdown of miR-155 suppressed TGF-␤-induced epithelial-mesenchymal transition (EMT) and tight junction dissolution, as well as cell migration and invasion. Further, the ectopic expression of miR-155 reduced RhoA protein and disrupted tight junction formation. Reintroducing RhoA cDNA without the 3 untranslated region largely reversed the phenotype induced by miR-155 and TGF-␤. In addition, elevated levels of miR-155 were frequently detected in invasive breast cancer tissues. These data suggest that miR-155 may play an important role in TGF-␤-induced EMT and cell migration and invasion by targeting RhoA and indicate that it is a potential therapeutic target for breast cancer intervention.Metastasis accounts for the majority of deaths of cancer patients, and thus, it is crucial to understand the molecular and cellular mechanisms that cause primary tumors to metastasize. The most critical step in the conversion of primary tumors to metastases is attributed to the process known as epithelialmesenchymal transition (EMT). EMT is a remarkable example of cellular plasticity that involves the dissolution of epithelial tight junctions, the intonation of adherens junctions, the remodeling of the cytoskeleton, and the loss of apical-basal polarity (49, 55). In cells undergoing EMT, the loss of epithelial cell adhesion and cytoskeletal components is coordinated with a gain of mesenchymal components and the initiation of a migratory phenotype.Transforming growth factor ␤ (TGF-␤) has emerged as a key regulator of EMT in late-stage carcinomas, where it promotes invasion and metastasis (54). TGF-␤ binds to a heteromeric complex of transmembrane serine/threonine kinases, the type I and II TGF-␤ receptors (T␤RI and T␤RII). Following ligand binding to T␤RII, the type I receptor is recruited to the ligand-receptor complex, where the constitutively active T␤RII transactivates T␤RI. Activated T␤RI phosphorylates the receptor-specific Smad2 and Smad3. Phosphorylated Smad2/ Smad3 associates with Smad4 as a heteromeric complex and translocates to the nucleus. This complex binds directly to Smad-binding elements and associates with a plethora of transcription factors, coactivators or corepressors, thus leading to the transcriptional induction or repression of a diverse array of genes (54). A number of genes that are associated with tumor growth and metastasis have been show...

Mutant p53 drives invasion in breast tumors through up-regulation of miR-155

“…Collectively, this study provides evidence that miR-155 targeted therapies may provide an attractive approach to treat mutant p53 expressing breast tumors. In contrast to our findings, a previous report suggests that ectopic expression of miR-155 did not drive a mesenchymal phenotype in the NMuMG murine mammary epithelial cell line (25).…”

“…Therefore, it is tempting to speculate a role for TGF-β in the regulation of miR-155 expression. This is indeed the case, as miR-155 has been previously shown to be upregulated upon TGF-β treatment (25) …”

“…In fact, findings from Xiang et al suggest that miR-155 suppresses the ability of 4T1 cells to undergo EMT. Indeed, it appears that the biological activity of miR-155 in human cell lines (10,24) is vastly different to that observed upon miR-155 expression in mammary cell lines that originate from mice (25,26), suggesting a lack of conservation of miR-155 seed sequences in its target 3'UTRs across these two species.…”

miR-421, miR-155 and miR-650: Emerging Trends of Regulation of Cancer and Apoptosis

“…C/EBPβ mediated expression of E-cadherin and therefore loss of C/EBP resulted in epithelial to mesenchymal transition (Johansson et al, 2013). There is a direct piece of evidence that suggests that miR-155 is controlled by SMAD4 in TGF treated breast cancer cells (Kong et al, 2008). mutant p53-expressing tumors displayed substantially enhanced expression of miR-155 (Neilsen et al, 2013).…”

The microRNA network and tumor metastasis

“…Apart from the miR-200 family, some other miRNAs have important regulatory roles in EMT. miR-155, encoding within a region known as Bic (B-cell integration cluster) and identified originally as a frequent integration site for avian leucosis virus (Tam et al, 1997;Lagos-Quintana et al, 2002), can not only regulate the generation of immunoglobulin class-switched plasma cells (Vigorito et al, 2007) but also augment the epithelial cell plasticity as part of EMT by targeting RhoA (Kong et al, 2008). In addition, Valastyan et al (2009) depicted how miR-31, a pleiotropically acting miRNA, inhibited breast cancer metastasis, and a few pro-metastatic genes had been verified as its targets, such as Fzd3, ITGA5, M-RIP, Matrix metalloproteinases 16 (MMP16), RDX and RhoA.…”