Knockdown of miR-182 promotes apoptosis via regulating RIP1 deubiquitination in TNF-α-treated triple-negative breast cancer cells

Tumour Biol. 2016 Oct;37(10):13733-13742. doi: 10.1007/s13277-016-5174-z. Epub 2016 Jul 30.

Abstract

Overexpression of microRNA-182 (miR-182) is found in multiple cancers, but the association of miR-182 expression with the sensitivity of triple-negative breast cancer (TNBC) cells to tumor necrosis factor-alpha (TNF-α) remains unknown. In this study, up-regulation of miR-182 was validated in TNBC patients and cell lines. Knockdown of miR-182 was observed to hinder the proliferation of BT-549 cells. More importantly, knockdown of miR-182 significantly promoted the apoptosis induced by TNF-α treatment in BT-549. JC-1 staining and western blot assays revealed that the K63-linked ubiquitin chains on receptor-interacting protein 1 (RIP1) were removed and the outer mitochondrial membrane potential (MMP) and permeability was altered upon combination of TNF-α with anti-miR-182. We then demonstrated that knockdown of miR-182 up-regulated the expression of cylindromatosis (CYLD) deubiquitinase, which promoted the formation of death-inducing signaling complex (DISC) and subsequent caspase-8 activation in TNF-α-treated BT-549 cells. Collectively, the results of the present study improve our understanding of the role of miR-182 in TNBC, knockdown of which facilitates the degradation of ubiquitin chains on RIP1, leading to the caspase-8 activation and apoptosis in TNF-α-treated TNBC cells. This may be valuable for the development of cancer therapy.

Keywords: CYLD; Deubiquitination; TNBC; TNF-α; miR-182.

MeSH terms

  • Apoptosis / drug effects
  • Biomarkers, Tumor / genetics
  • Biomarkers, Tumor / metabolism
  • Blotting, Western
  • Cell Proliferation / drug effects
  • Female
  • Flow Cytometry
  • Gene Expression Regulation, Neoplastic / drug effects
  • Humans
  • Immunoenzyme Techniques
  • Immunoprecipitation
  • MicroRNAs / antagonists & inhibitors
  • MicroRNAs / genetics*
  • Neoplasm Staging
  • Nuclear Pore Complex Proteins / genetics
  • Nuclear Pore Complex Proteins / metabolism*
  • Prognosis
  • RNA, Messenger / genetics
  • RNA-Binding Proteins / genetics
  • RNA-Binding Proteins / metabolism*
  • Real-Time Polymerase Chain Reaction
  • Reverse Transcriptase Polymerase Chain Reaction
  • Triple Negative Breast Neoplasms / drug therapy
  • Triple Negative Breast Neoplasms / genetics
  • Triple Negative Breast Neoplasms / metabolism
  • Triple Negative Breast Neoplasms / pathology*
  • Tumor Cells, Cultured
  • Tumor Necrosis Factor-alpha / pharmacology*
  • Ubiquitin / metabolism*
  • Ubiquitination / genetics*

Substances

  • AGFG1 protein, human
  • Biomarkers, Tumor
  • MicroRNAs
  • Mirn182 microRNA, human
  • Nuclear Pore Complex Proteins
  • RNA, Messenger
  • RNA-Binding Proteins
  • Tumor Necrosis Factor-alpha
  • Ubiquitin