Related Experiment Videos
Regulation of the gadd45beta promoter by NF-kappaB
Rongguan Jin1, Enrico De Smaele, Francesca Zazzeroni
1The Gwen Knapp Center for Lupus and Immunology Research, and The Ben May Institute for Cancer Research, Committees on Immunology and Cancer Biology, The University of Chicago, Chicago, Illinois 60637, USA.
Abstract:
In addition to coordinating immune and inflammatory responses, NF-kappaB/Rel transcription factors control cell survival. The NF-kappaB antiapoptotic function is crucial to oncogenesis, cancer chemoresistance, and to antagonize tumor necrosis factor (TNF) receptor-induced killing. Recently, we have shown that the suppression of the c-Jun-N-terminal kinase (JNK) cascade is a pivotal protective mechanism by NF-kappaB, and that this suppression involves the upregulation of gadd45beta/myd118. Induction of gadd45beta by stress and cytokines requires NF-kappaB; however, the regulatory mechanisms underlying this induction are not known. Here, we report that, in HeLa cells, the NF-kappaB subunit RelA is sufficient to activate gadd45beta expression, whereas Rel and p50 are not. Activation of gadd45beta by RelA depends on three kappaB elements at positions -447/-438 (kappaB-1), -426/-417 (kappaB-2), and -377/-368 (kappaB-3) of the gadd45beta promoter. Each of these sites binds to NF-kappaB complexes in vitro, and is required for optimal promoter transactivation. The data establish the direct participation of NF-kappaB in the regulation of Gadd45beta, thereby providing important mechanistic insights into the control of apoptosis by the transcription factor.
Insights
Nuclear factor-kappa B (NF-kappaB) directly regulates Gadd45beta expression via RelA, impacting cell survival and apoptosis. This finding offers insights into NF-kappaB
Area of Science:
- Molecular Biology
- Cellular Biology
- Cancer Research
Background:
- Nuclear factor-kappa B (NF-kappaB) transcription factors regulate immune responses and cell survival.
- NF-kappaB's antiapoptotic role is critical in cancer development and resistance to chemotherapy.
- NF-kappaB suppresses the c-Jun-N-terminal kinase (JNK) cascade, involving Gadd45beta (Growth arrest and DNA-damage-inducible 45 beta).
Purpose of the Study:
- To elucidate the regulatory mechanisms by which NF-kappaB induces Gadd45beta expression.
- To identify the specific NF-kappaB subunits and promoter elements involved in Gadd45beta regulation.
Main Methods:
- Reporter assays in HeLa cells to assess Gadd45beta promoter activity.
- Electrophoretic mobility shift assays (EMSAs) to study NF-kappaB binding to promoter elements.
- Mutagenesis of kappaB sites within the Gadd45beta promoter.
Main Results:
- The NF-kappaB subunit RelA, but not Rel or p50, was sufficient to activate Gadd45beta expression.
- Activation of Gadd45beta by RelA required three specific kappaB elements in the promoter region (-447/-438, -426/-417, -377/-368).
- These kappaB sites bound NF-kappaB complexes in vitro and were essential for optimal promoter activity.
Conclusions:
- NF-kappaB directly regulates Gadd45beta gene expression.
- RelA is the key NF-kappaB subunit mediating Gadd45beta induction.
- These findings provide mechanistic insights into how NF-kappaB controls apoptosis and cell survival.