Regulation of the sphingosine-recycling pathway for ceramide generation by oxidative stress, and its role in controlling c-Myc/Max function

Biochem J. 2006 Jan 15;393(Pt 2):513-21. doi: 10.1042/BJ20051083.


In the present study, the regulation of the sphingosine-recycling pathway in A549 human lung adenocarcinoma cells by oxidative stress was investigated. The generation of endogenous long-chain ceramide in response to exogenous C6-cer (C6-ceramide), which is FB1 (fumonisin B1)-sensitive, was employed to probe the sphingosine-recycling pathway. The data showed that ceramide formation via this pathway was significantly blocked by GSH and NAC (N-acetylcysteine) whereas it was enhanced by H2O2, as detected by both palmitate labelling and HPLC/MS. Similar data were also obtained using a novel approach that measures the incorporation of 17Sph (sphingosine containing 17 carbons) of 17C6-cer (C6-cer containing a 17Sph backbone) into long-chain 17C16-cer in cells by HPLC/MS, which was significantly decreased and increased in response to GSH and H2O2 respectively. TNF (tumour necrosis factor)-a, which decreases the levels of endogenous GSH, increased the generation of C16-cer in response to C6-cer, and this was blocked by exogenous GSH or NAC, or by the overexpression of TPx I (thioredoxin peroxidase I), an enzyme that reduces the generation of intracellular ROS (reactive oxygen species). Additional data showed that ROS regulated both the deacylation and reacylation steps of C6-cer. At a functional level, C6-cer inhibited the DNA-binding function of the c-Myc/Max oncogene. Inhibition of the generation of longchain ceramide in response to C6-cer by FB1 or NAC significantly blocked the modulation of the c-Myc/Max function. These data demonstrate that the sphingosine-recycling pathway for the generation of endogenous long-chain ceramide in response to exogenous C6-cer is regulated by ROS, and plays an important biological role in controlling c-Myc function.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Acylation
  • Basic-Leucine Zipper Transcription Factors / genetics
  • Basic-Leucine Zipper Transcription Factors / metabolism*
  • Cell Line, Tumor
  • Ceramides / biosynthesis*
  • Ceramides / chemistry
  • Ceramides / metabolism
  • Gene Expression Regulation
  • Golgi Apparatus / metabolism
  • Humans
  • Oxidative Stress*
  • Protein Binding
  • Proto-Oncogene Proteins c-myc / genetics
  • Proto-Oncogene Proteins c-myc / metabolism*
  • Reactive Oxygen Species / metabolism
  • Sphingosine / metabolism*
  • Substrate Specificity
  • Thioredoxin Reductase 1
  • Thioredoxin-Disulfide Reductase / metabolism
  • Time Factors
  • Tumor Necrosis Factor-alpha / metabolism
  • Tumor Necrosis Factor-alpha / pharmacology


  • Basic-Leucine Zipper Transcription Factors
  • Ceramides
  • Myc associated factor X
  • Proto-Oncogene Proteins c-myc
  • Reactive Oxygen Species
  • Tumor Necrosis Factor-alpha
  • TXNRD1 protein, human
  • Thioredoxin Reductase 1
  • Thioredoxin-Disulfide Reductase
  • Sphingosine