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Molecular and Cellular Biology, December 2006, p. 9209-9219, Vol. 26, No. 24
0270-7306/06/$08.00+0     doi:10.1128/MCB.00478-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Dimerization of the IB Kinase-Binding Domain of NEMO Is Required for Tumor Necrosis Factor Alpha-Induced NF-B Activity ^,

Ralf B. Marienfeld,^1,2, Lysann Palkowitsch,^2, and Sankar Ghosh^1*

Section of Immunobiology and Department of Molecular Biophysics and Biochemistry, Yale University Medical School, New Haven, Connecticut 06520,¹ Department of Physiological Chemistry, Ulm University, Albert Einstein Allee 11, 89081 Ulm, Germany²

Received 18 March 2006/ Returned for modification 20 April 2006/ Accepted 15 September 2006

Previous studies have demonstrated that peptides corresponding to a six-amino-acid NEMO-binding domain from the C terminus of IB kinase alpha (IKK) and IKKß can disrupt the IKK complex and block NF-B activation. We have now mapped and characterized the corresponding amino-terminal IKK-binding domain (IBD) of NEMO. Peptides corresponding to the IBD were efficiently recruited to the IKK complex but displayed only a weak inhibitory potential on cytokine-induced NF-B activity. This is most likely due to the formation of sodium dodecyl sulfate- and urea-resistant NEMO dimers through a dimerization domain at the amino terminus of NEMO that overlaps with the region responsible for binding to IKKs. Mutational analysis revealed different -helical subdomains within an amino-terminal coiled-coil region are important for NEMO dimerization and IKKß binding. Furthermore, NEMO dimerization is required for the tumor necrosis factor alpha-induced NF-B activation, even when interaction with the IKKs is unaffected. Hence, our data provide novel insights into the role of the amino terminus of NEMO for the architecture of the IKK complex and its activation.

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* Corresponding author. Mailing address: Section of Immunobiology and Department of Molecular Biophysics and Biochemistry, Yale University Medical School, 310 Cedar St., New Haven, CT 06520. Phone: (203) 737-4419. Fax: (203) 737-1764. E-mail: sankar.ghosh{at}yale.edu.

Published ahead of print on 25 September 2006.

Supplemental material for this article may be found at http://mcb.asm.org/.

These authors contributed equally to this work.

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Molecular and Cellular Biology, December 2006, p. 9209-9219, Vol. 26, No. 24
0270-7306/06/$08.00+0     doi:10.1128/MCB.00478-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

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