IEX-1 directly interferes with RelA/p65 dependent transactivation and regulation of apoptosis.
The early response gene IEX-1 plays a complex role in the regulation of apoptosis. Depending on the cellular context and the apoptotic stimulus, IEX-1 is capable to either enhance or suppress apoptosis. To further dissect the molecular mechanisms involved in the modulation of apoptosis by IEX-1, we analysed the molecular crosstalk between IEX-1 and the NF-kappaB pathway. Using GST-pulldown assays, a direct interaction of IEX-1 with the C-terminal region of the subunit RelA/p65 harbouring the transactivation domain of the NF-kappaB transcription factor was shown. This interaction negatively regulates RelA/p65 dependent transactivation as shown by GAL4-and luciferase assay and was confirmed for the endogenous proteins by co-immunoprecipitation experiments. Using deletion constructs, we were able to map the C-terminal region of IEX-1 as the critical determinant of the interaction with RelA/p65. We could further show, that IEX-1 mediated NF-kappaB inhibition accounts for the reduced expression of the anti-apoptotic NF-kappaB target genes Bcl-2, Bcl-xL, cIAP1 and cIAP2, thereby sensitizing cells for apoptotic stimuli. Finally, ChIP-assays revealed that IEX-1 associates with the promoter of these genes. Altogether, our findings suggest a critical role of IEX-1 in the NF-kappaB dependent regulation of apoptotic responses.
Arlt
Alexander
A
Laboratory of Molecular Gastroenterology and Hepatology, 1st Department of Medicine, University Hospital Schleswig-Holstein, Campus Kiel, Kiel, Germany. aarlt@1med.uni-kiel.de
Rosenstiel
Philip
P
Kruse
Marie-Luise
ML
Grohmann
Frauke
F
Minkenberg
Jörg
J
Perkins
Neil D
ND
Fölsch
Ulrich R
UR
Schreiber
Stefan
S
Schäfer
Heiner
H
D017209:Apoptosis / Q000235:genetics*; D051017:Apoptosis Regulatory Proteins / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D015398:Signal Transduction; D051996:Transcription Factor RelA / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D015533:Transcriptional Activation
D051017:Apoptosis Regulatory Proteins; C099815:IER3 protein, human; D008565:Membrane Proteins; D051996:Transcription Factor RelA