Binding of glyceraldehyde-3-phosphate dehydrogenase to the cis-acting element of structure-anchored repression in ccn2 mRNA

Seiji Kondo, Satoshi Kubota, Yoshiki Mukudai, Takashi Nishida, Yasuto Yoshihama, Tatsuo Shirota, Satoru Shintani, Masaharu Takigawa

Research output: Contribution to journalArticle

33 Citations (Scopus)

Abstract

CCN2/connective tissue growth factor (CTGF) can be induced by hypoxia and promotes tumor angiogenesis. Our previous studies revealed that hypoxia-induced gene expression of human ccn2 mRNA is regulated post-transcriptionally in human chondrosarcoma-derived cell line, HCS-2/8, in which a minimal cis-element, entitled CAESAR, in the 3'-untranslated region (UTR) of ccn2 mRNA and a 35-kDa protein counterpart play an important role by determining the stability of ccn2 mRNA. In the present study, we identified this corresponding protein as glyceraldehyde-3-phosphate dehydrogenase (GAPDH) by utilizing RNA affinity chromatography combined with mass spectrometry. The results of an RNA binding assay revealed the specific binding of GAPDH to this cis-element. To further characterize the interaction between GAPDH and ccn2 mRNA, we examined the roles of redox conditions and glycolytic coenzyme in the binding of GAPDH to the ccn2 mRNA. An oxidizing agent, diamide, abolished the GAPDH-RNA interaction in a concentration-dependent manner; whereas this effect could be reversed by subsequent treatment with 2-mercaptoethanol (2-ME). In addition, nicotinamide-adenine dinucleotide (NAD), a coenzyme of GAPDH, inhibited the GAPDH-RNA binding. Taken together, these findings suggest that the glycolytic enzyme GAPDH regulates the gene expression of ccn2 mRNA in trans by acting as a sensor of oxidative stress and redox signals, leading to CCN2 overexpression under the condition of hypoxia and promotion of angiogenesis.

Original languageEnglish
Pages (from-to)382-387
Number of pages6
JournalBiochemical and Biophysical Research Communications
Volume405
Issue number3
DOIs
Publication statusPublished - Feb 18 2011

Keywords

  • 3'-UTR
  • CCN2
  • GAPDH
  • Hypoxia
  • MRNA stability

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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