title: hsa-mir-30c promotes the invasive phenotype of metastatic breast cancer cells by targeting NOV/CCN3.
abstract: BACKGROUND
  For treatment and prevention of metastatic disease, one of the premier challenges is the identification of pathways and proteins to target for clinical intervention. Micro RNAs (miRNAs) are short, non-coding RNAs, which regulate cellular activities by either mRNA degradation or translational inhibition. Our studies focused on the invasive properties of hsa-mir30c based on its high expression in MDA-MB-231 metastatic cells and our bioinformatic analysis of the Cancer Genome Atlas that identified aberrant hsa-mir-30c to be associated with poor survival.
  
  
  METHODS
  Contributions of hsa-mir-30c to breast cancer cell invasion were examined by Matrigel invasion transwell assays following modulation of hsa-mir-30c or hsa-mir-30c* levels in MDA-MB-231 cells. hsa-mir-30c in silico predicted targets linked to cell invasion were screened for targeting by hsa-mir-30c in metastatic breast cancer cells by RT-qPCR. The contribution to invasion by a target of hsa-mir-30c, Nephroblastoma overexpressed (NOV), was characterized by siRNA and invasion assays. Significant effects were determined using Student's T-tests with Welch's correction for unequal variance.
  
  
  RESULTS
  MCF-7 and MDA-MB-231 cells were used as models of poorly invasive and late-stage metastatic disease, respectively. By modulating the levels of hsa-mir-30c in these cells, we observed concomitant changes in breast cancer cell invasiveness. From predicted targets of hsa-mir-30c that were related to cellular migration and invasion, NOV/CCN3 was identified as a novel target of hsa-mir-30c. Depleting NOV by siRNA caused a significant increase in the invasiveness of MDA-MB-231 cells is a regulatory protein associated with the extracellular matrix.
  
  
  CONCLUSIONS
  NOV/CCN3 expression, which protects cells from invasion, is known in patient tumors to inversely correlate with advanced breast cancer and metastasis. This study has identified a novel target of hsa-mir-30c, NOV, which is an inhibitor of the invasiveness of metastatic breast cancer cells. Thus, hsa-mir-30c-mediated inhibition of NOV levels promotes the invasive phenotype of MDA-MB-231 cells and significantly, the miR-30/NOV pathways is independent of RUNX2, a known target of hsa-mir-30c that promotes osteolytic disease in metastatic breast cancer cells. Our findings allow for mechanistic insight into the clinical observation of poor survival of patients with elevated hsa-mir-30c levels, which can be considered for miRNA-based translational studies.
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Ave, North, Worcester 01655, MA, USA ; Current address: Center for Computational Molecular Biology, Department of Molecular Biology, Cell Biology, and Biochemistry, and Department of Computer Science, Brown University, 115 Waterman Street, Providence 02912, RI, USA.
authors.forename: Jason R
authors.initials: JR
authors.lastname: Dobson
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Ave, North, Worcester 01655, MA, USA ; Current address: Heisenberg-Group for Molecular Skeletal Biology, Department of Trauma, Hand, and Reconstructive Surgery, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.
authors.forename: Hanna
authors.initials: H
authors.lastname: Taipaleenmäki
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Ave, North, Worcester 01655, MA, USA.
authors.forename: Yu-Jie
authors.initials: YJ
authors.lastname: Hu
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Ave, North, Worcester 01655, MA, USA ; Current address: Department of Biochemistry and Vermont Cancer Center, University of Vermont College of Medicine, 89 Beaumont Avenue, Burlington 05405-0068, VT, USA.
authors.forename: Deli
authors.initials: D
authors.lastname: Hong
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Ave, North, Worcester 01655, MA, USA ; Current address: Departments of Orthopedic Surgery and Biochemistry & Molecular Biology, Mayo Clinic, 200 First Street SW, Medical Sciences Building 3-69, Rochester 55905, MN, USA.
authors.forename: Andre J
authors.initials: AJ
authors.lastname: van Wijnen
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Ave, North, Worcester 01655, MA, USA ; Current address: Department of Biochemistry and Vermont Cancer Center, University of Vermont College of Medicine, 89 Beaumont Avenue, Burlington 05405-0068, VT, USA.
authors.forename: Janet L
authors.initials: JL
authors.lastname: Stein
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Ave, North, Worcester 01655, MA, USA ; Current address: Department of Biochemistry and Vermont Cancer Center, University of Vermont College of Medicine, 89 Beaumont Avenue, Burlington 05405-0068, VT, USA.
authors.forename: Gary S
authors.initials: GS
authors.lastname: Stein
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Ave, North, Worcester 01655, MA, USA ; Current address: Department of Biochemistry and Vermont Cancer Center, University of Vermont College of Medicine, 89 Beaumont Avenue, Burlington 05405-0068, VT, USA.
authors.forename: Jane B
authors.initials: JB
authors.lastname: Lian
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Ave, North, Worcester 01655, MA, USA ; Current address: Department of Anatomy and Cell Biology, Rush University Medical Center, Armour Academic Center, 600 S, Paulina Street, Suite 507, Chicago 60612, IL, USA.
authors.forename: Jitesh
authors.initials: J
authors.lastname: Pratap
keywords: Hsa-mir-30c breast cancer; Invasion; Metastasis; NOV/CCN3; miRNA