PARP-1 promotes autophagy via the AMPK/mTOR pathway in CNE-2 human nasopharyngeal carcinoma cells following ionizing radiation, while inhibition of autophagy contributes to the radiation sensitization of CNE-2 cells

Mol Med Rep. 2015 Aug;12(2):1868-76. doi: 10.3892/mmr.2015.3604. Epub 2015 Apr 9.

Abstract

It was previously reported that poly-(adenosine diphosphate-ribose) polymerase-1 (PARP-1) regulated ionizing radiation (IR)-induced autophagy in CNE-2 human nasopharyngeal carcinoma cells. The present study aimed to investigate whether PARP-1-mediated IR-induced autophagy occurred via activation of the liver kinase B1 (LKB1)/adenosine monophosphate-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) signaling pathway in CNE-2 cells. In addition, the effect of PARP-1 and AMPK inhibition on the radiation sensitization of CNE-2 cells was investigated. CNE-2 cells were treated with 10 Gy IR in the presence or absence of the AMPK activator 5-amino-1-β-D-ribofuranosyl-1H-imid-azole-4-carboxamide (AICAR). In addition, IR-treated CNE-2 cells were transfected with lentivirus-delivered small-hairpin RNA or treated with the AMPK inhibitor Compound C. Western blot analysis was used to assess the protein expression of PARP-1, phosphorylated (p)-AMPK, microtubule-associated protein 1 light chain 3 (LC3)-II and p-P70S6K. Cell viability and clone formation assays were performed to determine the effect of PARP-1 silencing and AMPK inhibition on the radiation sensitization of CNE-2 cells. The results showed that IR promoted PARP-1, p-AMPK and LC3-II protein expression as well as decreased p-P70S6K expression compared with that of the untreated cells. In addition, AICAR increased the expression of p-AMPK and LC3-II as well as decreased p-P70S6K expression compared with that of the IR-only group; however, AICAR did not increase PARP-1 expression. Furthermore, PARP-1 gene silencing decreased the expression of PARP-1, p-AMPK and LC3-II as well as increased p-P70S6K expression. Compound C decreased p-AMPK and LC3-Ⅱ expression as well as increased p-P70S6K expression; however, Compound C did not increase PARP-1 expression. Western blot analysis detected limited expression of p-LKB1 in all treatment groups. Cell viability and clone formation assays revealed that PARP-1 or AMPK inhibition reduced the proliferation of CNE-2 cells following IR. In conclusion, the present study demonstrated that PARP-1 promoted autophagy via the AMPK/mTOR pathway; in addition, PARP-1 or AMPK inhibition contributed to the radiation sensitization of CNE-2 cells following IR. However, it remains to be elucidated whether PARP-1 is an upstream mediator of the LKB1 pathway in CNE‑2 cells following IR.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • AMP-Activated Protein Kinases / antagonists & inhibitors
  • AMP-Activated Protein Kinases / chemistry
  • AMP-Activated Protein Kinases / metabolism*
  • Aminoimidazole Carboxamide / analogs & derivatives
  • Aminoimidazole Carboxamide / pharmacology
  • Autophagy / drug effects
  • Autophagy / radiation effects*
  • Carcinoma
  • Cell Line, Tumor
  • Cell Survival / drug effects
  • Cell Survival / radiation effects
  • Humans
  • Microscopy, Fluorescence
  • Microtubule-Associated Proteins / genetics
  • Microtubule-Associated Proteins / metabolism
  • Nasopharyngeal Carcinoma
  • Nasopharyngeal Neoplasms / metabolism
  • Nasopharyngeal Neoplasms / pathology
  • Phosphorylation / drug effects
  • Phosphorylation / radiation effects
  • Poly (ADP-Ribose) Polymerase-1
  • Poly(ADP-ribose) Polymerases / chemistry
  • Poly(ADP-ribose) Polymerases / genetics
  • Poly(ADP-ribose) Polymerases / metabolism*
  • RNA Interference
  • RNA, Small Interfering / metabolism
  • Radiation, Ionizing*
  • Ribonucleotides / pharmacology
  • Ribosomal Protein S6 Kinases, 70-kDa / genetics
  • Ribosomal Protein S6 Kinases, 70-kDa / metabolism
  • Signal Transduction / drug effects
  • Signal Transduction / radiation effects
  • TOR Serine-Threonine Kinases / metabolism*

Substances

  • MAP1LC3A protein, human
  • Microtubule-Associated Proteins
  • RNA, Small Interfering
  • Ribonucleotides
  • Aminoimidazole Carboxamide
  • PARP1 protein, human
  • Poly (ADP-Ribose) Polymerase-1
  • Poly(ADP-ribose) Polymerases
  • Ribosomal Protein S6 Kinases, 70-kDa
  • TOR Serine-Threonine Kinases
  • AMP-Activated Protein Kinases
  • AICA ribonucleotide