UV Irradiation and Heat Shock Mediate JNK Activation via Alternate Pathways (*)
- § To whom correspondence should be addressed.
Abstract
To elucidate cellular pathways involved in Jun-NH
-terminal kinase (JNK) activation by different forms of stress, we have compared the effects of UV irradiation, heat shock,
and H2O2. Using mouse fibroblast cells (3T3-4A) we show that while H2O2 is ineffective, UV and heat shock (HS) are potent inducers of JNK. The cellular pathways that mediate JNK activation after
HS or UV exposure are distinctly different as can be concluded from the following observations: (i) H2O2 is a potent inhibitor of HS-induced but not of UV-induced JNK activation; (ii) Triton X-100-treated cells abolish the ability
of UV, but not HS, to activate JNK; (iii) the free radical scavenger N-acetylcysteine inhibits UV- but not HS-mediated JNK activation; (iv) N-acetylcysteine inhibition is blocked by H2O2 in a dose-dependent manner; (v) a Cockayne syndrome-derived cell line exhibits JNK activation upon UV exposure, but not upon
HS treatment. The significance of Jun phosphorylation by JNK after treatment with UV, HS, or H2O2 was evaluated by measuring Jun phosphorylation in vivo and also its binding activity in gel shifts. HS and UV, which are potent inducers of JNK, increased the level of c-Jun phosphorylation
when this was measured by [
P]orthophosphate labeling of 3T3-4A cultures. H2O2 had no such effect. Although H2O2 failed to activate JNK in vitro and to phosphorylate c-Jun in vivo, all three forms of stress were found to be potent inducers of binding to the AP1 target sequence. Overall, our data indicate
that both membrane-associated components and oxidative damage are involved in JNK activation by UV irradiation, whereas HS-mediated
JNK activation, which appears to be mitochondrial-related, utilizes cellular sensors.
Footnotes
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↵* This work was supported by Grants CA-51995 and CA-59908 from the National Cancer Institute. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵1 The abbreviations used are:
- HS
-
heat shock
- EMSA
-
electrophoretic mobility shift assay
- PAGE
-
polyacrylamide gel electrophoresis
- JNK
-
c-Jun amino-terminal protein kinase
- GST
-
glutathione S-transferase
- CS
-
Cockayne syndrome.
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↵2 V. Adler, L. Dolan, V. Dhar, and Z. Ronai, unpublished observations.
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- Received August 3, 1995.
- © 1995 by The American Society for Biochemistry and Molecular Biology, Inc.
