A Novel Effect of MARCKS Phosphorylation by Activated PKC: The Dephosphorylation of Its Serine 25 in Chick Neuroblasts

Andrea Toledo, Flavio R. Zolessi, Cristina Arruti
PLoS ONE. 2013-04-25; 8(4): e62863
DOI: 10.1371/journal.pone.0062863

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1. PLoS One. 2013 Apr 25;8(4):e62863. doi: 10.1371/journal.pone.0062863. Print 2013.

A novel effect of MARCKS phosphorylation by activated PKC: the dephosphorylation
of its serine 25 in chick neuroblasts.

Toledo A(1), Zolessi FR, Arruti C.

Author information:
(1)Laboratorio de Cultivo de Tejidos, Sección Biología Celular, Departamento de
Biología Celular y Molecular, Facultad de Ciencias, Universidad de la República,
Montevideo, Uruguay.

MARCKS (Myristoylated Alanine-Rich C Kinase Substrate) is a peripheral membrane
protein, especially abundant in the nervous system, and functionally related to
actin organization and Ca-calmodulin regulation depending on its phosphorylation
by PKC. However, MARCKS is susceptible to be phosphorylated by several different
kinases and the possible interactions between these phosphorylations have not
been fully studied in intact cells. In differentiating neuroblasts, as well as
some neurons, there is at least one cell-type specific phosphorylation site:
serine 25 (S25) in the chick. We demonstrate here that S25 is included in a
highly conserved protein sequence which is a Cdk phosphorylatable region, located
far away from the PKC phosphorylation domain. S25 phosphorylation was inhibited
by olomoucine and roscovitine in neuroblasts undergoing various states of cell
differentiation in vitro. These results, considered in the known context of Cdks
activity in neuroblasts, suggest that Cdk5 is the enzyme responsible for this
phosphorylation. We find that the phosphorylation by PKC at the effector domain
does not occur in the same molecules that are phosphorylated at serine 25. The in
situ analysis of the subcellular distribution of these two phosphorylated MARCKS
variants revealed that they are also segregated in different protein clusters. In
addition, we find that a sustained stimulation of PKC by
phorbol-12-myristate-13-acetate (PMA) provokes the progressive disappearance of
phosphorylation at serine 25. Cells treated with PMA, but in the presence of
several Ser/Thr phosphatase (PP1, PP2A and PP2B) inhibitors indicated that this
dephosphorylation is achieved via a phosphatase 2A (PP2A) form. These results
provide new evidence regarding the existence of a novel consequence of PKC
stimulation upon the phosphorylated state of MARCKS in neural cells, and propose
a link between PKC and PP2A activity on MARCKS.

DOI: 10.1371/journal.pone.0062863
PMCID: PMC3636281
PMID: 23634231 [Indexed for MEDLINE]


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