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Published on: October 4, 2019
miR-200bc/429 cluster targets PLCgamma1 and differentially regulates proliferation and EGF-driven invasion than
S Uhlmann1, J D Zhang, A Schwäger
1Division of Molecular Genome Analysis, German Cancer Research Center, Heidelberg, Germany.
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-to-mesenchymal transition and transforming growth factor-beta-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 EGF-driven 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.
Insights
The miR-200 family of microRNAs differentially regulates breast cancer cell behavior. The miR-200bc/429 cluster, targeting PLCG1, shows stronger effects on invasion and cell cycle than the miR-200a/141 cluster.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- The miR-200 family is frequently downregulated in cancer.
- Its role in epidermal growth factor (EGF)-driven breast cancer progression is largely unknown.
- Previous studies focused on transforming growth factor-beta, not EGF.
Purpose of the Study:
- To investigate the differential roles of miR-200 family clusters in EGF-driven breast cancer.
- To compare the effects of miR-200a/141 and miR-200bc/429 clusters on cell invasion, viability, apoptosis, and cell cycle.
- To identify specific molecular targets responsible for observed phenotypic differences.
Main Methods:
- Gain-of-function studies with miR-200 family members.
- Genome-wide microarray profiling.
- Luciferase reporter assays to validate direct targets.
- Analysis of cell invasion, viability, apoptosis, and cell cycle progression.
Main Results:
- Both miR-200 clusters differentially regulate EGF-driven breast cancer cell invasion and viability.
- The miR-200bc/429 cluster exhibited stronger effects than the miR-200a/141 cluster.
- Phospholipase C gamma 1 (PLCG1) was identified as a direct target of the miR-200bc/429 cluster, influencing cell phenotypes.
Conclusions:
- The miR-200 family acts as a tumor suppressor in breast cancer.
- Differential targeting by miR-200 clusters, particularly PLCG1 by miR-200bc/429, impacts EGF-driven progression.
- Understanding these cluster-specific roles is crucial for therapeutic strategies.
