Proline dehydrogenase is essential for proline protection against hydrogen peroxide-induced cell death

Sathish K Natarajan, Weidong Zhu, Xinwen Liang, Lu Zhang, Andrew J. Demers, Matthew C Zimmerman, Melanie A. Simpson, Donald F Becker

Research output: Contribution to journalArticle

55 Citations (Scopus)

Abstract

Proline metabolism has an underlying role in apoptotic signaling that influences tumorigenesis. Proline is oxidized to glutamate in the mitochondria, with the rate-limiting step catalyzed by proline dehydrogenase (PRODH). PRODH expression is inducible by p53, leading to increased proline oxidation, reactive oxygen species formation, and induction of apoptosis. Paradoxical to its role in apoptosis, proline also protects cells against oxidative stress. Here we explore the mechanism of proline protection against hydrogen peroxide stress in melanoma WM35 cells. Treatment of WM35 cells with proline significantly increased cell viability, diminished oxidative damage of cellular lipids and proteins, and maintained ATP and NADPH levels after exposure to hydrogen peroxide. Inhibition or siRNA-mediated knockdown of PRODH abolished proline protection against oxidative stress, whereas knockdown of Δ1- pyrroline-5-carboxylate reductase, a key enzyme in proline biosynthesis, had no impact on proline protection. Potential linkages between proline metabolism and signaling pathways were explored. The combined inhibition of the mammalian target of rapamycin complex 1 (mTORC1) and mTORC2 eliminated proline protection. A significant increase in Akt activation was observed in proline-treated cells after hydrogen peroxide stress along with a corresponding increase in the phosphorylation of the forkhead transcription factor class O3a (FoxO3a). The role of PRODH in proline-mediated protection was validated in the prostate carcinoma cell line PC3. Knockdown of PRODH in PC3 cells attenuated phosphorylated levels of Akt and FoxO3a and decreased cell survival during hydrogen peroxide stress. The results provide evidence that PRODH is essential in proline protection against hydrogen peroxide-mediated cell death and that proline/PRODH helps activate Akt in cancer cells.

Original languageEnglish (US)
Pages (from-to)1181-1191
Number of pages11
JournalFree Radical Biology and Medicine
Volume53
Issue number5
DOIs
StatePublished - Sep 1 2012

Fingerprint

Proline Oxidase
Cell death
Proline
Hydrogen Peroxide
Cell Death
Cells
Oxidative stress
Metabolism
Pyrroline Carboxylate Reductases
Cell Survival
Oxidative Stress
Apoptosis
Forkhead Transcription Factors
Phosphorylation
Mitochondria

Keywords

  • Akt
  • Free radicals
  • Oxidative stress
  • Proline
  • Proline dehydrogenase
  • Pyrroline-5-carboxylate reductase

ASJC Scopus subject areas

  • Biochemistry
  • Physiology (medical)

Cite this

Proline dehydrogenase is essential for proline protection against hydrogen peroxide-induced cell death. / Natarajan, Sathish K; Zhu, Weidong; Liang, Xinwen; Zhang, Lu; Demers, Andrew J.; Zimmerman, Matthew C; Simpson, Melanie A.; Becker, Donald F.

In: Free Radical Biology and Medicine, Vol. 53, No. 5, 01.09.2012, p. 1181-1191.

Research output: Contribution to journalArticle

Natarajan, Sathish K ; Zhu, Weidong ; Liang, Xinwen ; Zhang, Lu ; Demers, Andrew J. ; Zimmerman, Matthew C ; Simpson, Melanie A. ; Becker, Donald F. / Proline dehydrogenase is essential for proline protection against hydrogen peroxide-induced cell death. In: Free Radical Biology and Medicine. 2012 ; Vol. 53, No. 5. pp. 1181-1191.
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AB - Proline metabolism has an underlying role in apoptotic signaling that influences tumorigenesis. Proline is oxidized to glutamate in the mitochondria, with the rate-limiting step catalyzed by proline dehydrogenase (PRODH). PRODH expression is inducible by p53, leading to increased proline oxidation, reactive oxygen species formation, and induction of apoptosis. Paradoxical to its role in apoptosis, proline also protects cells against oxidative stress. Here we explore the mechanism of proline protection against hydrogen peroxide stress in melanoma WM35 cells. Treatment of WM35 cells with proline significantly increased cell viability, diminished oxidative damage of cellular lipids and proteins, and maintained ATP and NADPH levels after exposure to hydrogen peroxide. Inhibition or siRNA-mediated knockdown of PRODH abolished proline protection against oxidative stress, whereas knockdown of Δ1- pyrroline-5-carboxylate reductase, a key enzyme in proline biosynthesis, had no impact on proline protection. Potential linkages between proline metabolism and signaling pathways were explored. The combined inhibition of the mammalian target of rapamycin complex 1 (mTORC1) and mTORC2 eliminated proline protection. A significant increase in Akt activation was observed in proline-treated cells after hydrogen peroxide stress along with a corresponding increase in the phosphorylation of the forkhead transcription factor class O3a (FoxO3a). The role of PRODH in proline-mediated protection was validated in the prostate carcinoma cell line PC3. Knockdown of PRODH in PC3 cells attenuated phosphorylated levels of Akt and FoxO3a and decreased cell survival during hydrogen peroxide stress. The results provide evidence that PRODH is essential in proline protection against hydrogen peroxide-mediated cell death and that proline/PRODH helps activate Akt in cancer cells.

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