Adenosine diphosphate-ribosylation factor 6 is required for epidermal growth factor-induced glioblastoma cell proliferation

Ming Li, Jide Wang, Samuel S.M. Ng, Chu Yan Chan, Ming Liang He, Fang Yu, Lihui Lai, Chao Shi, Yangchao Chen, David T. Yew, Hsiang Fu Kung, Marie Chia Mi Lin

Research output: Contribution to journalArticlepeer-review

25 Scopus citations


BACKGROUND: Epidermal growth factor (EGF) signaling plays a pivotal role in gliomagenesis. The authors previously demonstrated that adenosine diphospate-ribosylation factor 6 (ARF6), a member of the Ras-related small guanosine-50-triphospate-binding protein family, is required for EFA6A-induced glioma cell migration and invasion. However, the role of ARF6 in EGF signaling is unknown. METHODS: The authors analyzed messenger RNA (mRNA) levels of ARF6 and EGF receptor (EGFR) in 16 high-grade glioma samples and in 6 low-grade glioma samples by reverse transcriptase-polymerase chain reaction analysis. To determine whether EGF induces ARF6 expression in human glioblastoma U87 cells through transcriptional regulation and EGFR activation, the levels of ARF6 were assayed in EGF-treated U87 cells that were preincubated with a transcriptional inhibitor (actinomycin D) and an EGFR tyrosine kinase inhibitor (PD153035), respectively. The downstream signaling of EGFR-mediated ARF6 up-regulation also was investigated using specific inhibitors of mitogen-activated protein kinase (MEK), phosphatidylinositol 30 kinase (PI3K), and Janus kinase 2. The involvement of SP1 in the downstream signaling was studied by using an SP1 inhibitor (mithramycin A). Small-interfering RNAs (siRNAs) targeting ARF6 were used to investigate the effects of ARF6 on EGF-mediated glioma cell proliferation. RESULTS: The results demonstrated that ARF6 and EGFR mRNA levels were elevated in glioma tissues. Furthermore, EGF stimulated ARF6 expression in U87 cells in a dose-dependent and time-dependant manner. This stimulation was caused by increased transcription of ARF6 and by activation of the MEK/extracellular signal-regulated kinase 1 and 2 (ERK1/2) and PI3K signaling pathways. It is noteworthy that SP1 was essential for EGF-induced ARF6 up-regulation. Finally, EGF-induced glioblastoma cell proliferation depended on ARF6, because the suppression of ARF6 by siRNA or by a dominant-negative mutant significantly inhibited EGF-induced cell proliferation. CONCLUSIONS: The results of the current study suggested that EGF-induced ARF6 expression plays a significant role in glioma cell proliferation.

Original languageEnglish (US)
Pages (from-to)4959-4972
Number of pages14
Issue number21
StatePublished - Nov 1 2009
Externally publishedYes


  • Adenosine diphosphate-ribosylation factor 6
  • Epidermal growth factor
  • Glioma
  • SP1
  • Signaling pathway


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