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Lipid rafts are required for Kit survival and proliferation signals.

ABSTRACT: In addition to its physiologic role as central regulator of the hematopoietic and reproductive systems, the Kit receptor tyrosine kinase (RTK) is pathologically overexpressed in some forms of leukemia and constitutively activated by oncogenic mutations in mast-cell proliferations and gastrointestinal stromal tumors. To gain insight into the general activation and signaling mechanisms of RTKs, we investigated the activation-dependent dynamic membrane distributions of wild-type and oncogenic forms of Kit in hematopoietic cells. Ligand-induced recruitment of wild-type Kit to lipid rafts after stimulation by Kit ligand (KL) and the constitutive localization of oncogenic Kit in lipid rafts are necessary for Kit-mediated proliferation and survival signals. KL-dependent and oncogenic Kit kinase activity resulted in recruitment of the regulatory phosphatidylinositol 3-kinase (PI3-K) subunit p85 to rafts where the catalytical PI3-K subunit p110 constitutively resides. Cholesterol depletion by methyl-beta-cyclodextrin prevented Kit-mediated activation of the PI3-K downstream target Akt and inhibited cellular proliferation by KL-activated or oncogenic Kit, including mutants resistant to the Kit inhibitor imatinib-mesylate. Our data are consistent with the notion that Kit recruitment to lipid rafts is required for efficient activation of the PI3-K/Akt pathway and Kit-mediated proliferation.

SUBMITTER: Jahn T

PROVIDER: S-EPMC1976347 | biostudies-literature | 2007 Sep

REPOSITORIES: biostudies-literature

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Lipid rafts are required for Kit survival and proliferation signals.

Jahn Thomas T Leifheit Erica E Gooch Stacie S Sindhu Simran S Weinberg Kenneth K

Blood 20070606 6

In addition to its physiologic role as central regulator of the hematopoietic and reproductive systems, the Kit receptor tyrosine kinase (RTK) is pathologically overexpressed in some forms of leukemia and constitutively activated by oncogenic mutations in mast-cell proliferations and gastrointestinal stromal tumors. To gain insight into the general activation and signaling mechanisms of RTKs, we investigated the activation-dependent dynamic membrane distributions of wild-type and oncogenic forms ...[more]

PMID: 17554062

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Project description:BACKGROUND:KIT tyrosine kinase is expressed in mast cells, interstitial cells of Cajal, and hematopoietic cells. Permanently active KIT mutations lead these host cells to tumorigenesis, and to such diseases as mast cell leukemia (MCL), gastrointestinal stromal tumor (GIST), and acute myeloid leukemia (AML). Recently, we reported that in MCL, KIT with mutations (D816V, human; D814Y, mouse) traffics to endolysosomes (EL), where it can then initiate oncogenic signaling. On the other hand, KIT mutants including KITD814Y in GIST accumulate on the Golgi, and from there, activate downstream. KIT mutations, such as N822K, have been found in 30% of core binding factor-AML (CBF-AML) patients. However, how the mutants are tyrosine-phosphorylated and where they activate downstream molecules remain unknown. Moreover, it is unclear whether a KIT mutant other than KITD816V in MCL is able to signal on EL. METHODS:We used leukemia cell lines, such as Kasumi-1 (KITN822K, AML), SKNO-1 (KITN822K, AML), and HMC-1.1 (KITV560G, MCL), to explore how KIT transduces signals in these cells and to examine the signal platform for the mutants using immunofluorescence microscopy and inhibition of intracellular trafficking. RESULTS:In AML cell lines, KITN822K aberrantly localizes to EL. After biosynthesis, KIT traffics to the cell surface via the Golgi and immediately migrates to EL through endocytosis in a manner dependent on its kinase activity. However, results of phosphorylation imaging show that KIT is preferentially activated on the Golgi. Indeed, blockade of KITN822K migration to the Golgi with BFA/M-COPA inhibits the activation of KIT downstream molecules, such as AKT, ERK, and STAT5, indicating that KIT signaling occurs on the Golgi. Moreover, lipid rafts in the Golgi play a role in KIT signaling. Interestingly, KITV560G in HMC-1.1 migrates and activates downstream in a similar manner to KITN822K in Kasumi-1. CONCLUSIONS:In AML, KITN822K mislocalizes to EL. Our findings, however, suggest that the mutant transduces phosphorylation signals on lipid rafts of the Golgi in leukemia cells. Unexpectedly, the KITV560G signal platform in MCL is similar to that of KITN822K in AML. These observations provide new insights into the pathogenic role of KIT mutants as well as that of other mutant molecules.

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Lipid rafts are required for Kit survival and proliferation signals.

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Lipid rafts are required for Kit survival and proliferation signals.

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