Justicidin A-Induced autophagy flux enhances apoptosis of human colorectal cancer cells via class III PI3K and Atg5 pathway

Shen Jeu Won, Cheng Hsin Yen, Hsiao Sheng Liu, Shan Ying Wu, Sheng Hui Lan, Ya Fen Jiang-Shieh, Chun Nan Lin, Chun-Li Su

Research output: Contribution to journalArticle

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Abstract

Our previous reports showed that justicidin A (JA), a novel and pure arylnaphthalide lignan isolated from Justicia procumbens, induces apoptosis of human colorectal cancer cells and hepatocellular carcinoma cells, leading to the suppression of both tumor cell growth in NOD-SCID mice. Here, we reveal that JA induces autophagy in human colorectal cancer HT-29 cells by conversion of autophagic marker LC3-I to LC3-II. Furthermore, LC3 puncta and autophagic vesicle formation, and SQSTM1/p62 suppression were observed. Administration of autophagy inhibitor (bafilomycin A1 and chloroquine) and transfection of a tandem fluorescent-tagged LC3 (mRFP-GFP) reporter plasmid (ptfLC3) demonstrated that JA induces autophagy flux in HT-29 cells. Expression of LC3, SQSTM1, Beclin 1, and nuclear DNA double-strand breaks (representing apoptosis) were also detected in the tumor tissue of HT-29 cells transplanted into NOD-SCID mice orally administrated with JA. In addition, the expression of autophagy signaling pathway-related molecules p-PDK1, p-mTOR, p-p70S6k/p-RPS6KB2 was decreased, whereas that of class III PI3K, Beclin 1, Atg5-Atg12, and mitochondrial BNIP3 was increased in response to JA. Pre-treatment of the cells with class III PI3K inhibitor 3-methyladenine or Atg5 shRNA attenuated JA-induced LC3-II expression and LC3 puncta formation, indicating the involvement of class III PI3K and Atg5. A novel mechanism was demonstrated in the anticancer compound JA; pre-treatment with 3-methyladenine or Atg5 shRNA blocked JA-induced suppression in cell growth and colony formation, respectively, via inhibition of apoptosis. In contrast, administration of apoptosis inhibitor Z-VAD did not affect JA-induced autophagy. Our data suggest the chemotherapeutic potential of JA for treatment of human colorectal cancer.

Original languageEnglish
Pages (from-to)930-946
Number of pages17
JournalJournal of Cellular Physiology
Volume230
Issue number4
DOIs
Publication statusPublished - 2015 Apr 1

Fingerprint

Autophagy
Phosphatidylinositol 3-Kinases
Colorectal Neoplasms
Cells
Apoptosis
Fluxes
HT29 Cells
Inbred NOD Mouse
SCID Mice
Cell growth
Small Interfering RNA
Tumors
Justicia
justicidin A
70-kDa Ribosomal Protein S6 Kinases
Lignans
Double-Stranded DNA Breaks
Chloroquine
Growth
Transfection

ASJC Scopus subject areas

  • Physiology
  • Clinical Biochemistry
  • Cell Biology

Cite this

Justicidin A-Induced autophagy flux enhances apoptosis of human colorectal cancer cells via class III PI3K and Atg5 pathway. / Won, Shen Jeu; Yen, Cheng Hsin; Liu, Hsiao Sheng; Wu, Shan Ying; Lan, Sheng Hui; Jiang-Shieh, Ya Fen; Lin, Chun Nan; Su, Chun-Li.

In: Journal of Cellular Physiology, Vol. 230, No. 4, 01.04.2015, p. 930-946.

Research output: Contribution to journalArticle

Won, Shen Jeu ; Yen, Cheng Hsin ; Liu, Hsiao Sheng ; Wu, Shan Ying ; Lan, Sheng Hui ; Jiang-Shieh, Ya Fen ; Lin, Chun Nan ; Su, Chun-Li. / Justicidin A-Induced autophagy flux enhances apoptosis of human colorectal cancer cells via class III PI3K and Atg5 pathway. In: Journal of Cellular Physiology. 2015 ; Vol. 230, No. 4. pp. 930-946.
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T1 - Justicidin A-Induced autophagy flux enhances apoptosis of human colorectal cancer cells via class III PI3K and Atg5 pathway

AU - Won, Shen Jeu

AU - Yen, Cheng Hsin

AU - Liu, Hsiao Sheng

AU - Wu, Shan Ying

AU - Lan, Sheng Hui

AU - Jiang-Shieh, Ya Fen

AU - Lin, Chun Nan

AU - Su, Chun-Li

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AB - Our previous reports showed that justicidin A (JA), a novel and pure arylnaphthalide lignan isolated from Justicia procumbens, induces apoptosis of human colorectal cancer cells and hepatocellular carcinoma cells, leading to the suppression of both tumor cell growth in NOD-SCID mice. Here, we reveal that JA induces autophagy in human colorectal cancer HT-29 cells by conversion of autophagic marker LC3-I to LC3-II. Furthermore, LC3 puncta and autophagic vesicle formation, and SQSTM1/p62 suppression were observed. Administration of autophagy inhibitor (bafilomycin A1 and chloroquine) and transfection of a tandem fluorescent-tagged LC3 (mRFP-GFP) reporter plasmid (ptfLC3) demonstrated that JA induces autophagy flux in HT-29 cells. Expression of LC3, SQSTM1, Beclin 1, and nuclear DNA double-strand breaks (representing apoptosis) were also detected in the tumor tissue of HT-29 cells transplanted into NOD-SCID mice orally administrated with JA. In addition, the expression of autophagy signaling pathway-related molecules p-PDK1, p-mTOR, p-p70S6k/p-RPS6KB2 was decreased, whereas that of class III PI3K, Beclin 1, Atg5-Atg12, and mitochondrial BNIP3 was increased in response to JA. Pre-treatment of the cells with class III PI3K inhibitor 3-methyladenine or Atg5 shRNA attenuated JA-induced LC3-II expression and LC3 puncta formation, indicating the involvement of class III PI3K and Atg5. A novel mechanism was demonstrated in the anticancer compound JA; pre-treatment with 3-methyladenine or Atg5 shRNA blocked JA-induced suppression in cell growth and colony formation, respectively, via inhibition of apoptosis. In contrast, administration of apoptosis inhibitor Z-VAD did not affect JA-induced autophagy. Our data suggest the chemotherapeutic potential of JA for treatment of human colorectal cancer.

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