miR-200bc/429 cluster targets PLCγ1 and differentially regulates proliferation and EGF-driven invasion than miR-200a/141 in breast cancer

Abstract

The genes encoding microRNAs of the human miR-200 family map to fragile chromosomal regions and are frequently downregulated upon tumor progression. Although having been reported to regulate epithelial-tomesenchymal transition and transforming growth factorbeta-driven cell invasion, the role of the miR-200 family in EGF-driven breast cancer cell invasion, viability, apoptosis and cell cycle progression is still unknown. In particular, there is no study comparing the roles of the two clusters of this miRNA family. In this study, we show for the first time that miR-200 family members differentially regulate EGF-driven invasion, viability, apoptosis and cell cycle progression of breast cancer cells. We showed that, all miR-200 family members regulate EGFdriven invasion, with the miR-200bc/429 cluster showing stronger effects than the miR-200a/141 cluster. Furthermore, expression of the miR-200a/141 cluster results in G1 arrest supported by increased p27/Kip1 and decreased cyclin dependent kinase 6 expression. In contrast, expression of the 200bc/429 cluster decreases G1 population and increases G2/M phase, in line with the observed reduction of p27/Kip1 and upregulation of the inhibitory phosphorylation of Cdc25C, respectively. To test the hypothesis that phenotypical differences observed between the two clusters are caused by differential targeting spectrums, we performed genome-wide microarray profiling in combination with gain-of-function studies. This identified phospholipase C gamma 1 (PLCG1), which was downregulated only by the miR-200bc/429 cluster, as a potential candidate contributing to these phenotypical differences. Luciferase reporter assays validated PLCG1 as a direct functional target of miR-200bc/429 cluster, but not of miR-200a/141 cluster. Finally, loss of PLCG1 in part mimicked the effect of miR-200bc/429 overexpression in viability, apoptosis and EGF-driven cell invasion of breast cancer cells. Our results suggest that the miR-200 family has a tumor-suppressor function by negatively regulating EGF-driven cell invasion, viability and cell cycle progression in breast cancer.

MicroRNA-520/373 family functions as a tumor suppressor in estrogen receptor negative breast cancer by targeting NF-κB and TGF-β signaling pathways

“…Recent progress in cancer biology has revealed that miRNAs are frequently deregulated in various human cancers, including breast cancer. For instance, elevated miR-21 expression has been reported in breast tumors (Volinia et al, 2006), while miR-335 and miR-200c have been shown to inhibit metastatic cell invasion (Tavazoie et al, 2008;Uhlmann et al, 2010). These findings demonstrate the ability of miRNAs to modulate the development of malignancies by regulating critical cancer-related genes and signaling pathways.…”

“…miR-520/373 family modulates NF-jB and TGF-b signaling I Keklikoglou et al cancer cells in vivo, a chorioallantoic membrane (CAM) assay was performed (Zijlstra et al, 2002;van der Horst et al, 2004). MDA-MB-231 cells were transfected with miR-520/373 family mimics or with small interfering RNAs targeting RELA and TGFBR2 genes and were inoculated on the upper CAM of 10-day-old chicken embryos.…”

MicroRNA-200c plays an oncogenic role in nasopharyngeal carcinoma by targeting PTEN

“…25 MiR-200c inhibits matrix invasion by breast cancer cell MDA-MB-231, 26 while miR-200b/c/429 cluster significantly reduces cell growth and promotes apoptosis. 27 Meanwhile, miR-200 family also inhibits EMT by downregulating ZEB1 and ZEB2. [28][29][30] These studies all suggest anti-cancer roles of miR-200 family.…”

miR-200a Regulates Nrf2 Activation by Targeting Keap1 mRNA in Breast Cancer Cells

“…The miR-200 family has been found to play a critical role in the maintenance of the epithelial phenotype by targeting ZEB1, SIP1, and SIRT1, thus preventing the silencing of E-cadherin (30,31). Within the miR-200 family, there are two clusters: miR-200b/200a/429 located on chromosome 1p36, and miR-200c/141 located on chromosome 12p13 (32).…”