Expression of MARCKS effector domain mutants alters phospholipase D activity and cytoskeletal morphology of SK-N-MC neuroblastoma cells

Sherry C. Morash; Donna Douglas; Christopher R. McMaster; Harold W. Cook; David M. Byers

doi:10.1007/s11064-005-8220-6

Expression of MARCKS effector domain mutants alters phospholipase D activity and cytoskeletal morphology of SK-N-MC neuroblastoma cells

Sherry C. Morash, Donna Douglas, Christopher R. McMaster, Harold W. Cook, David M. Byers

Medicine

Medicine

Résultat de recherche: Article › examen par les pairs

6 Citations (Scopus)

Résumé

Stable overexpression of myristoylated alanine-rich C-kinase substrate (MARCKS) is known to enhance phorbol ester stimulation of phospholipase D (PLD) activity and protein kinase Cα (PKCα) levels in SK-N-MC neuroblastoma cells. In contrast, expression of MARCKS mutants (S152A or S156A) lacking key PKC phosphorylation sites within the central basic effector domain (ED) had no significant effect on PLD activity or PKCα levels relative to vector control cells. Like control cells, those expressing wild type MARCKS were elongated and possessed longitudinally oriented stress fibers, although these cells were more prone to detach from the substratum and undergo cell death upon phorbol ester treatment. However, cells expressing MARCKS ED mutants were irregularly shaped and stress fibers were either shorter or less abundant, and cell adhesion and viability were not affected. These results suggest that intact phosphorylation sites within the MARCKS ED are required for PLD activation and influence both membrane-cytoskeletal organization and cell viability.

Langue d'origine	English
Pages (de-à)	1353-1364
Nombre de pages	12
Journal	Neurochemical Research
Volume	30
Numéro de publication	11
DOI	https://doi.org/10.1007/s11064-005-8220-6
Statut de publication	Published - nov. 2005

Note bibliographique

Funding Information:
This work was supported by operating grants from the Canadian Institutes for Health Research (DMB, HWC, and CRM), the Lockwood Foundation (SCM), and a Canada Research Chairs award (CRM). The excellent technical assistance of Susan Powell, Robert Zwicker and Gladys Keddy is gratefully acknowledged. The authors thank Dr. Michael Frohman, State University of New York, Stony Brook for the generous gifts of the PLD1 antibody and HA-tagged hPLD1 and mPLD2 constructs, and Dr. Sylvain Bourgoin, Centre de Recherche du CHUL, Quebec, Canada for the PLD2 antibody.

ASJC Scopus Subject Areas

Biochemistry
Cellular and Molecular Neuroscience

PubMed: MeSH publication types

Journal Article
Research Support, Non-U.S. Gov't

Accès au document

10.1007/s11064-005-8220-6

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title = "Expression of MARCKS effector domain mutants alters phospholipase D activity and cytoskeletal morphology of SK-N-MC neuroblastoma cells",

abstract = "Stable overexpression of myristoylated alanine-rich C-kinase substrate (MARCKS) is known to enhance phorbol ester stimulation of phospholipase D (PLD) activity and protein kinase Cα (PKCα) levels in SK-N-MC neuroblastoma cells. In contrast, expression of MARCKS mutants (S152A or S156A) lacking key PKC phosphorylation sites within the central basic effector domain (ED) had no significant effect on PLD activity or PKCα levels relative to vector control cells. Like control cells, those expressing wild type MARCKS were elongated and possessed longitudinally oriented stress fibers, although these cells were more prone to detach from the substratum and undergo cell death upon phorbol ester treatment. However, cells expressing MARCKS ED mutants were irregularly shaped and stress fibers were either shorter or less abundant, and cell adhesion and viability were not affected. These results suggest that intact phosphorylation sites within the MARCKS ED are required for PLD activation and influence both membrane-cytoskeletal organization and cell viability.",

author = "Morash, {Sherry C.} and Donna Douglas and McMaster, {Christopher R.} and Cook, {Harold W.} and Byers, {David M.}",

note = "Funding Information: This work was supported by operating grants from the Canadian Institutes for Health Research (DMB, HWC, and CRM), the Lockwood Foundation (SCM), and a Canada Research Chairs award (CRM). The excellent technical assistance of Susan Powell, Robert Zwicker and Gladys Keddy is gratefully acknowledged. The authors thank Dr. Michael Frohman, State University of New York, Stony Brook for the generous gifts of the PLD1 antibody and HA-tagged hPLD1 and mPLD2 constructs, and Dr. Sylvain Bourgoin, Centre de Recherche du CHUL, Quebec, Canada for the PLD2 antibody.",

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AU - McMaster, Christopher R.

AU - Cook, Harold W.

AU - Byers, David M.

N1 - Funding Information: This work was supported by operating grants from the Canadian Institutes for Health Research (DMB, HWC, and CRM), the Lockwood Foundation (SCM), and a Canada Research Chairs award (CRM). The excellent technical assistance of Susan Powell, Robert Zwicker and Gladys Keddy is gratefully acknowledged. The authors thank Dr. Michael Frohman, State University of New York, Stony Brook for the generous gifts of the PLD1 antibody and HA-tagged hPLD1 and mPLD2 constructs, and Dr. Sylvain Bourgoin, Centre de Recherche du CHUL, Quebec, Canada for the PLD2 antibody.

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N2 - Stable overexpression of myristoylated alanine-rich C-kinase substrate (MARCKS) is known to enhance phorbol ester stimulation of phospholipase D (PLD) activity and protein kinase Cα (PKCα) levels in SK-N-MC neuroblastoma cells. In contrast, expression of MARCKS mutants (S152A or S156A) lacking key PKC phosphorylation sites within the central basic effector domain (ED) had no significant effect on PLD activity or PKCα levels relative to vector control cells. Like control cells, those expressing wild type MARCKS were elongated and possessed longitudinally oriented stress fibers, although these cells were more prone to detach from the substratum and undergo cell death upon phorbol ester treatment. However, cells expressing MARCKS ED mutants were irregularly shaped and stress fibers were either shorter or less abundant, and cell adhesion and viability were not affected. These results suggest that intact phosphorylation sites within the MARCKS ED are required for PLD activation and influence both membrane-cytoskeletal organization and cell viability.

AB - Stable overexpression of myristoylated alanine-rich C-kinase substrate (MARCKS) is known to enhance phorbol ester stimulation of phospholipase D (PLD) activity and protein kinase Cα (PKCα) levels in SK-N-MC neuroblastoma cells. In contrast, expression of MARCKS mutants (S152A or S156A) lacking key PKC phosphorylation sites within the central basic effector domain (ED) had no significant effect on PLD activity or PKCα levels relative to vector control cells. Like control cells, those expressing wild type MARCKS were elongated and possessed longitudinally oriented stress fibers, although these cells were more prone to detach from the substratum and undergo cell death upon phorbol ester treatment. However, cells expressing MARCKS ED mutants were irregularly shaped and stress fibers were either shorter or less abundant, and cell adhesion and viability were not affected. These results suggest that intact phosphorylation sites within the MARCKS ED are required for PLD activation and influence both membrane-cytoskeletal organization and cell viability.

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Expression of MARCKS effector domain mutants alters phospholipase D activity and cytoskeletal morphology of SK-N-MC neuroblastoma cells

Résumé

Note bibliographique

ASJC Scopus Subject Areas

PubMed: MeSH publication types

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