Ceramide recruits and activates protein kinase C ζ (PKCζ) within structured membrane microdomains

Todd E. Fox, Kristy L. Houck, Sean M. O'Neill, Murali Nagarajan, Thomas C. Stover, Pawel T. Pomianowski, Onur Unal, Jong Yun, Stanley J. Naides, Mark Kester

Research output: Contribution to journalArticle

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Abstract

We have previously demonstrated that hexanoyl-D-erythro-sphingosine (C 6-ceramide), an anti-mitogenic cell-permeable lipid metabolite, limited vascular smooth muscle growth by abrogating trauma-induced Akt activity in a stretch injury model of neointimal hyperplasia. Furthermore, ceramide selectively and directly activated protein kinase Cζ (PKCζ) to suppress Akt-dependent mitogenesis. To further analyze the interaction between ceramide and PKCζ, the ability of ceramide to localize within highly structured lipid microdomains (rafts) and activate PKCζ was investigated. Using rat aorta vascular smooth muscle cells (A7r5), we now demonstrate that C6-ceramide treatment results in an increased localization and phosphorylation of PKCζ within caveolin-enriched lipid microdomians to inactivate Akt. In addition, ceramide specifically reduced the association of PKCζ with 14-3-3, a scaffold protein localized to less structured regions within membranes. Pharmacological disruption of highly structured lipid microdomains resulted in abrogation of ceramide-activated, PKCζ-dependent Akt inactivation, whereas molecular strategies suggest that ceramide-dependent PKCζ phosphorylation of Akt3 at Ser34 was necessary for ceramide-induced vascular smooth muscle cell growth arrest. Taken together, these data demonstrate that structured membrane microdomains are necessary for ceramide-induced activation of PKCζ and resultant diminished Akt activity, leading to vascular smooth muscle cell growth arrest.

Original languageEnglish (US)
Pages (from-to)12450-12457
Number of pages8
JournalJournal of Biological Chemistry
Volume282
Issue number17
DOIs
StatePublished - Apr 27 2007

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Membrane Microdomains
Ceramides
Protein Kinase C
Membranes
Vascular Smooth Muscle
Muscle
Smooth Muscle Myocytes
Lipids
Phosphorylation
Cell growth
Growth
Caveolins
Wounds and Injuries
Metabolites
Scaffolds
Hyperplasia
Aorta
Rats
Chemical activation
Cells

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Fox, T. E., Houck, K. L., O'Neill, S. M., Nagarajan, M., Stover, T. C., Pomianowski, P. T., ... Kester, M. (2007). Ceramide recruits and activates protein kinase C ζ (PKCζ) within structured membrane microdomains. Journal of Biological Chemistry, 282(17), 12450-12457. https://doi.org/10.1074/jbc.M700082200
Fox, Todd E. ; Houck, Kristy L. ; O'Neill, Sean M. ; Nagarajan, Murali ; Stover, Thomas C. ; Pomianowski, Pawel T. ; Unal, Onur ; Yun, Jong ; Naides, Stanley J. ; Kester, Mark. / Ceramide recruits and activates protein kinase C ζ (PKCζ) within structured membrane microdomains. In: Journal of Biological Chemistry. 2007 ; Vol. 282, No. 17. pp. 12450-12457.
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Fox, TE, Houck, KL, O'Neill, SM, Nagarajan, M, Stover, TC, Pomianowski, PT, Unal, O, Yun, J, Naides, SJ & Kester, M 2007, 'Ceramide recruits and activates protein kinase C ζ (PKCζ) within structured membrane microdomains', Journal of Biological Chemistry, vol. 282, no. 17, pp. 12450-12457. https://doi.org/10.1074/jbc.M700082200

Ceramide recruits and activates protein kinase C ζ (PKCζ) within structured membrane microdomains. / Fox, Todd E.; Houck, Kristy L.; O'Neill, Sean M.; Nagarajan, Murali; Stover, Thomas C.; Pomianowski, Pawel T.; Unal, Onur; Yun, Jong; Naides, Stanley J.; Kester, Mark.

In: Journal of Biological Chemistry, Vol. 282, No. 17, 27.04.2007, p. 12450-12457.

Research output: Contribution to journalArticle

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T1 - Ceramide recruits and activates protein kinase C ζ (PKCζ) within structured membrane microdomains

AU - Fox, Todd E.

AU - Houck, Kristy L.

AU - O'Neill, Sean M.

AU - Nagarajan, Murali

AU - Stover, Thomas C.

AU - Pomianowski, Pawel T.

AU - Unal, Onur

AU - Yun, Jong

AU - Naides, Stanley J.

AU - Kester, Mark

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Y1 - 2007/4/27

N2 - We have previously demonstrated that hexanoyl-D-erythro-sphingosine (C 6-ceramide), an anti-mitogenic cell-permeable lipid metabolite, limited vascular smooth muscle growth by abrogating trauma-induced Akt activity in a stretch injury model of neointimal hyperplasia. Furthermore, ceramide selectively and directly activated protein kinase Cζ (PKCζ) to suppress Akt-dependent mitogenesis. To further analyze the interaction between ceramide and PKCζ, the ability of ceramide to localize within highly structured lipid microdomains (rafts) and activate PKCζ was investigated. Using rat aorta vascular smooth muscle cells (A7r5), we now demonstrate that C6-ceramide treatment results in an increased localization and phosphorylation of PKCζ within caveolin-enriched lipid microdomians to inactivate Akt. In addition, ceramide specifically reduced the association of PKCζ with 14-3-3, a scaffold protein localized to less structured regions within membranes. Pharmacological disruption of highly structured lipid microdomains resulted in abrogation of ceramide-activated, PKCζ-dependent Akt inactivation, whereas molecular strategies suggest that ceramide-dependent PKCζ phosphorylation of Akt3 at Ser34 was necessary for ceramide-induced vascular smooth muscle cell growth arrest. Taken together, these data demonstrate that structured membrane microdomains are necessary for ceramide-induced activation of PKCζ and resultant diminished Akt activity, leading to vascular smooth muscle cell growth arrest.

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Fox TE, Houck KL, O'Neill SM, Nagarajan M, Stover TC, Pomianowski PT et al. Ceramide recruits and activates protein kinase C ζ (PKCζ) within structured membrane microdomains. Journal of Biological Chemistry. 2007 Apr 27;282(17):12450-12457. https://doi.org/10.1074/jbc.M700082200