Elsevier

Neuroscience

Volume 151, Issue 3, 6 February 2008, Pages 771-779
Neuroscience

Activating transcription factor 3 up-regulated by c-Jun NH2-terminal kinase/c-Jun contributes to apoptosis induced by potassium deprivation in cerebellar granule neurons

https://doi.org/10.1016/j.neuroscience.2007.10.057Get rights and content

Abstract

Cerebellar granule neurons (CGNs) depend on potassium depolarization for survival and undergo apoptosis when deprived of depolarizing concentration of potassium. Activating transcription factor 3 (ATF3), a stress-inducible protein, belongs to the ATF/CREB family of transcription factors family and is involved in cell growth and apoptosis. However, the role of ATF3 in neuronal apoptosis remains unknown. Here, we showed that ATF3 was up-regulated under potassium deprivation in CGNs, and this induction was preceded by a rapid and sustained activation of c-Jun NH2-terminal kinase/c-Jun signaling pathway, which plays a fundamental role in neuronal apoptosis. Furthermore, ATF3 up-regulation was abolished by inhibition of JNK or knockdown of c-Jun. Finally, knockdown of ATF3 by RNA interference protected CGNs from potassium deprivation–induced apoptosis. Taken together, our results indicate that ATF3 is a downstream target of JNK/c-Jun pathway and contributes to apoptosis induced by potassium deprivation in rat CGNs.

Section snippets

Neuronal culture and potassium deprivation

Rat CGNs were prepared from 7 to 8-day-old Sprague–Dawley rat pups (15–19 g) as described previously (Li et al., 2000, Ma et al., 2007). Briefly, neurons were dissociated from freshly dissected cerebella by mechanical disruption in the presence of trypsin and DNase and then seeded at a density of 1.5×106 cells/ml in basal modified Eagle medium (BME) containing 10% fetal bovine serum and 25 mM KCl (25 K+S). Cytosine arabinoside (10 μM) was added 24 h after seeding to limit the growth of

JNK/c-Jun is activated and ATF3 is up-regulated during apoptosis induced by potassium deprivation in CGNs

To determine whether potassium deprivation evokes the activation of JNK/c-Jun in CGNs, DIV7 neurons were switched to 25 K or 5 K for various duration of time (0.5, 1, 2, 4 h), and then processed for Western blotting with antibodies against phospho-JNK and phospho-c-Jun. Consistent with previous studies (Coffey et al., 2002, Ma et al., 2007), potassium deprivation (5 K vs. 25 K) led to an increase in the level of phospho-JNK starting at 0.5 h without detectable changes in total JNK level (Fig. 1

Discussion

Although the induction of ATF3 during neuronal apoptosis has been widely reported, the role of ATF3 in neuronal apoptosis has not been well studied. To the best of our knowledge, this is the first report that utilizes siRNA targeting ATF3 for analysis of the role of endogenous ATF3 during neuronal apoptosis. By knockdown of ATF3, we demonstrate that the up-regulation of ATF3 mediated by JNK/c-Jun pathway contributes to apoptosis induced by potassium deprivation in CGNs. JNK/c-Jun pathway plays

Conclusion

In summary, our results suggest that JNK/c-Jun mediates the up-regulation of ATF3 and ATF3 contributes to apoptosis induced by potassium deprivation in CGNs.

Acknowledgment

This work was supported by the National Natural Science Foundation of China (grants 30570562, 30629002 and U0632006); the Natural Science Foundation of Guangdong Province (grant 5100982); Y.-P.C. was financially supported by Hong Kong University/NFSC Young Research Award.

References (43)

  • C. Murakata et al.

    Mixed lineage kinase activity of indolocarbazole analogues

    Bioorg Med Chem Lett

    (2002)
  • R. Murata et al.

    Extracorporeal shockwaves induce the expression of ATF3 and GAP-43 in rat dorsal root ganglion neurons

    Auton Neurosci

    (2006)
  • K. Nobori et al.

    ATF3 inhibits doxorubicin-induced apoptosis in cardiac myocytes: a novel cardioprotective role of ATF3

    J Mol Cell Cardiol

    (2002)
  • N. Ohba et al.

    Biphasic expression of activating transcription factor-3 in neurons after cerebral infarction

    Brain Res Mol Brain Res

    (2003)
  • Y.X. Pan et al.

    Interaction of RNA-binding Proteins HuR and AUF1 with the Human ATF3 mRNA 3′-untranslated region regulates its amino acid limitation-induced stabilization

    J Biol Chem

    (2005)
  • A.G. Pearson et al.

    ATF3 enhances c-Jun-mediated neurite sprouting

    Mol Brain Res

    (2003)
  • L. Shi et al.

    Activity deprivation-dependent induction of the proapoptotic BH3-only protein Bim is independent of JNK/c-Jun activation during apoptosis in cerebellar granule neurons

    Neurosci Lett

    (2005)
  • H. Tsujino et al.

    Activating transcription factor 3 (ATF3) induction by axotomy in sensory and motoneurons: A novel neuronal marker of nerve injury

    Mol Cell Neurosci

    (2000)
  • J. Whitfield et al.

    Dominant-negative c-Jun promotes neuronal survival by reducing BIM expression and inhibiting mitochondrial cytochrome c release

    Neuron

    (2001)
  • T. Yin et al.

    Tissue-specific pattern of stress kinase activation in ischemic/reperfused heart and kidney

    J Biol Chem

    (1997)
  • B.L. Bennett et al.

    SP600125, an anthrapyrazolone inhibitor of Jun N-terminal kinase

    Proc Natl Acad Sci U S A

    (2001)
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