LncRNA TTN-AS1/miR-134-5p/PAK3 axis regulates the radiosensitivity of human large intestine cancer cells through the P21 pathway and AKT/GSK-3β/β-catenin pathway

Cell Biol Int. 2020 Nov;44(11):2284-2292. doi: 10.1002/cbin.11436. Epub 2020 Aug 10.

Abstract

Radiotherapy is an important adjuvant treatment for large intestine cancer even though it does not cause any response in many patients. The present study aimed to investigate the effects of the TTN antisense RNA 1 (TTN-AS1) long noncoding RNA (lncRNA) on radiotherapy dynamics of large intestine cancer cells and to explore the underlying molecular mechanisms. TTN-AS1 expression was evaluated by reverse-transcription quantitative polymerase chain reaction, western blot, and cellular immunofluorescence, and flow cytometry analysis was used to measure apoptosis. Radiotherapy was simulated in vitro by exposing cancer cells to X-ray. TTN-AS1 was highly expressed in large intestine cancer cells after an X-ray exposition for 24 hr. TTN-AS1 knockdown improved the radiosensitivity of large intestine cancer cells and promoted apoptosis by increasing Bax/Bcl2 protein expression and the active-caspase 3/caspase 3 ratios following X-ray treatment. In addition, TTN-AS1 negatively regulated miR-134-5p expression, and miR-134-5p-mimic transfection decreased PAK3 protein expression in large intestine cancer cells. Importantly, TTN-AS1 promoted PAK3 and P21 protein expression in HT29 cells after X-ray treatment. Moreover, the knockdown of P21 protein expression improved radiosensitivity and promoted X-ray-induced apoptosis of HT29 cells. Finally, PAK3 knockdown expression decreased the p-AKT/AKT and p-GSK-3β/GSK-3β ratios and promoted the β-catenin transfer from the nucleus to the cytoplasm. These data suggest that the TTN-AS1 lncRNA promoted resistance to radiotherapy of large intestine cancer cells by increasing PAK3 expression via miR-134-5p inhibition, and this may be related to the P21 and AKT/GSK-3β/β-catenin pathway.

Keywords: HT29 cell; TTN-AS1; apoptosis; large intestine cancer; radiotherapy.

MeSH terms

  • Apoptosis / drug effects
  • Cell Line, Tumor
  • Cell Movement / genetics
  • Cell Proliferation / drug effects
  • Colorectal Neoplasms / genetics
  • Colorectal Neoplasms / pathology
  • Connectin / genetics
  • Cyclin-Dependent Kinase Inhibitor p21 / genetics
  • Cyclin-Dependent Kinase Inhibitor p21 / metabolism
  • Gene Expression Regulation, Neoplastic / genetics
  • Glycogen Synthase Kinase 3 beta / metabolism
  • Humans
  • Intestinal Neoplasms / genetics*
  • Intestinal Neoplasms / metabolism
  • Intestine, Large / metabolism
  • Intestine, Large / pathology
  • MicroRNAs / genetics*
  • MicroRNAs / metabolism
  • Oligoribonucleotides, Antisense / genetics
  • Proto-Oncogene Proteins c-akt / metabolism
  • RNA, Long Noncoding / genetics*
  • RNA, Long Noncoding / metabolism
  • Radiation Tolerance / genetics
  • Signal Transduction / genetics
  • Signal Transduction / physiology
  • beta Catenin / metabolism
  • p21-Activated Kinases / genetics

Substances

  • CDKN1A protein, human
  • Connectin
  • Cyclin-Dependent Kinase Inhibitor p21
  • MIRN134 microRNA, human
  • MicroRNAs
  • Oligoribonucleotides, Antisense
  • RNA, Long Noncoding
  • TTN protein, human
  • beta Catenin
  • Glycogen Synthase Kinase 3 beta
  • Proto-Oncogene Proteins c-akt
  • p21-Activated Kinases