Visit our Expo - Redox and Inflammation signaling 2012

Session XIV : Transcriptional and translational control Poster XIV, 29
The transcription factor Nrf2 controls the expression of heme oxygenase-1 and Phase IIrelated
genes in cells exposed to aqueous extracts of cigarette smoke
Arnd Hengstermann1, Constanze Knörr-Wittmann1, Stephan Gebel1, Jawed Alam2,
Thomas Müller1.
1PHILIP MORRIS Research Laboratories GmbH, Cologne, Germany. 2Department of
Molecular Genetics, Alton Ochsner Clinic Foundation, New Orleans, LA, U.S.A. E-mail:
Arnd.Hengstermann@pmintl.com
Exposure of cells and tissues to cigarette smoke (CS) triggers a pronounced anti-oxidant
response, which is hallmarked by the transcriptional up-regulation of heme oxygenase-1
(hmox1), initiating a self-protection mechanism resulting in the formation of endogenous
antioxidant molecules, i.e., biliverdin and bilirubin from intracellular heme moieties. To
characterize the regulatory elements involved in CS-mediated hmox1 expression, we studied
the expression of various hmox1 promoter/luciferase reporter constructs in NIH3T3 cells
exposed to aqueous extracts of CS. The results showed that the CS-dependent expression of
hmox1 is governed primarily by the distal enhancers 1 and 2, both of which contain three
canonical anti-oxidant responsive element (ARE)-like stress responsive elements. These sites
are potentially addressed by the transcription-factor Nrf2, a principal inducer of antioxidant
and Phase II-related genes. As shown by Western-blot analysis, Nrf2 was strongly stabilized
and became detectable in nuclear extracts in cells exposed to aqueous extracts of CS.
Furthermore, nuclear localization of Nrf2 coincided with increased DNA binding of a putative
Nrf2/MafK heterodimer to ARE, as determined by EMSA. Notably, siRNA-mediated knockdown
of Nrf2 expression significantly compromised both CS-induced hmox1 promoter
activation and HO-1 expression. Finally, by using this approach CS-induced expression of
Phase II-related genes NAD(P)H:quinone oxidoreductase and glutamate-cysteine ligase,
catalytic subunit was shown to be completely abrogated, as determined by Real Time
quantitative PCR.
Taken together, these results add to the understanding of the central role of Nrf2 in the
context of CS-mediated oxidative stress by orchestrating an efficient transcriptional response
aimed at resolving the stressing conditions.
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