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Vol. 24, No. 3-4, 2009
Issue release date: 2009
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Cell Physiol Biochem 2009;24:219–230
(DOI:10.1159/000233248)
Original Paper

A Ceramide-binding C1 Domain Mediates Kinase Suppressor of Ras Membrane Translocation

Yin X.1 · Zafrullah M.1 · Lee H.1 · Haimovitz-Friedman A.2 · Fuks Z.2 · Kolesnick R.1
1Laboratory of Signal Transduction and2Department of Radiation Oncology, Memorial Sloan-Kettering Cancer Center, New York Cell Physiol Biochem 2009;24:219–230 (DOI:10.1159/000233248)

Abstract

Genetic and biochemical data support Kinase Suppressor of Ras 1 (KSR1) as a positive regulator of the Ras-Raf-MAPK pathway, functioning as a kinase and/or scaffold to regulate c-Raf-1 activation. Membrane translocation mediated by the KSR1 CA3 domain, which is homologous to the atypical PKC C1 lipid-binding domain, is a critical step of KSR1-mediated c-Raf-1 activation. In this study, we used an ELISA to characterize the KSR1 CA3 domain as a lipid-binding moiety. Purified GST-KSR1-CA3 protein effectively binds ceramide but not other lipids including 1,2-diacylglyceol, dihydroceramide, ganglioside GM1, sphingomyelin and phosphatidylcholine. Upon epidermal growth factor stimulation of COS-7 cells, KSR1 translocates into and is activated within glycosphingolipid-enriched plasma membrane platforms. Pharmacologic inhibition of ceramide generation attenuates KSR1 translocation and KSR1 kinase activation in COS-7 cells. Disruption of two cysteines, which are indispensable for maintaining ternary structure of all C1 domains and their lipid binding capability, mitigates ceramide-binding capacity of purified GST-KSR1-CA3 protein, and inhibits full length KSR1 membrane translocation and kinase activation. These studies provide evidence for a mechanism by which the second messenger ceramide can target proteins to subcellular compartments in the process of transmembrane signal transduction.

 goto top of outline Author Contacts

Richard Kolesnick MD
Memorial Sloan-Kettering Cancer Center
1275 York Avenue, New York, NY 10021 (USA)
Tel. +1 646 888-2174, Fax: +1 646 422-0281
E-Mail r-kolesnick@ski.mskcc.org


 goto top of outline Article Information

Accepted: May 23, 2009
Published online: August 03, 2009
Number of Print Pages : 12


 goto top of outline Publication Details

Cellular Physiology and Biochemistry (International Journal of Experimental Cellular Physiology, Biochemistry andPharmacology)

Vol. 24, No. 3-4, Year 2009 (Cover Date: 2009)

Journal Editor: F. Lang, Tübingen
ISSN: 1015–8987 (Print), eISSN: 1421–9778 (Online)

For additional information: http://www.karger.com/journals/cpb


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