MicroRNA miR–142–3p inhibits breast cancer cell invasiveness and stem cell properties by targeting integrin alpha V, KLF4 and multiple cytoskeletal elements

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Abstract

MicroRNAs are post-transcriptional regulators of gene expression which influence tumorigenesis and tumor progression. miR-142-3p expression is dysregulated in several breast cancer subtypes, but its function at the molecular level is not known in detail. Here, we demonstrate that miR-142-3p is regulated in response to irradiation (2Gy) in human breast cancer cells, suggesting a potential role in the response to radiotherapy. Supported by transcriptomic Affymetrix array analysis and confirmatory investigations at the mRNA and protein level, we demonstrate that overexpression of miR-142-3p in MDA-MB-231, MDA-MB-468 and MCF-7 breast cancer cells leads to downregulation of Integrin- WASL, RAC1, and CFL2, molecules implicated in cytoskeletal regulation and cell motility. ROCK2, IL6ST, KLF4, PGRMC2 and ADCY9 were identified as additional targets in a subset of cell lines. miR- 142-3p overexpressing cells showed decreased Matrigel invasiveness and decreased activity of the stemness-associated enzyme ALDH. Confocal immunofluorescence microscopy, nanoscale atomic force microscopy and digital holographic microscopy revealed an altered actin distribution, a loss of membrane protrusions and a reduced cell volume and size. Interference with additional components of the extracellular matrix environment (Syndecan-1, Hyaluronan) resulted in similar alterations of membrane protrusion formation. Luciferase activation assays confirmed direct miR-142-3p-dependent regulation of the 3’-untranslated region of ITGAV and WASL. siRNA-knockdown of ITGAV and WASL resulted in a significant reduction of cellular invasiveness, highlighting their contribution to the invasion phenotype. While WASL-knockdown significantly reduced the number of membrane protrusions compared to controls, knockdown of ITGAV resulted in a decreased cell volume. Our data identify ITGAV, WASL, and several additional cytoskeleton- associated molecules as novel invasionpromoting targets of miR-142-3p in breast cancer. Whether regulation of the pluripotency-associated transcription factor KLF4 and alterations in ALDH1 activity are linked to alterations in matrixdependent signaling, or due to direct regulation by miR-142-3p is currently under investigation.

Speakers from the University of Münster

Götte, Martin

Department of Gynecology and Obstetrics