JBC Transcription and Nuclear Factor Monoclonals

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Originally published In Press as doi:10.1074/jbc.M411045200 on October 25, 2004

J. Biol. Chem., Vol. 279, Issue 53, 55262-55270, December 31, 2004
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Phosphorylation of NG2 Proteoglycan by Protein Kinase C-{alpha} Regulates Polarized Membrane Distribution and Cell Motility*

Irwan T. Makagiansar{ddagger}, Scott Williams, Kimberlee Dahlin-Huppe, Jun-ichi Fukushi§, Tomas Mustelin, and William B. Stallcup

From the Cancer Research Center, The Burnham Institute, La Jolla, California 92037

Protein kinase C (PKC)-{alpha} phosphorylation of recombinant NG2 cytoplasmic domain and phorbol ester-induced PKC-dependent phosphorylation of full-length NG2 expressed in U251 cells are both blocked by mutation of Thr2256, identifying this residue as a primary phosphorylation site. In untreated U251/NG2 cells, NG2 is present along with ezrin and {alpha}3{beta}1 integrin in apical cell surface protrusions. Phorbol ester treatment causes redistribution of all three components to lamellipodia, accompanied by increased cell motility. U251 cells expressing NG2 with a valine substitution at position 2256 are resistant to phorbol ester treatment: NG2 remains in membrane protrusions and cell motility is unchanged. In contrast, NG2 with a glutamic acid substitution at position 2256 redistributes to lamellipodia even without phorbol ester treatment, rendering transfected U251 cells spontaneously motile. PKC-{alpha}-mediated NG2 phosphorylation at Thr2256 is therefore a key step for initiating cell polarization and motility.


Received for publication, September 27, 2004

* This work was supported by National Institutes of Health Grants PO1-HD25938 and RO1-CA95287 (to W. B. S.) and RO1 Grants AI35603, AI48032, AI53585, AI55741, and CA96949 (to T. M.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

§ Present address: Dept. of Orthopedic Surgery, Kyushu University, Fukuoka 812-8582, Japan.

{ddagger} To whom correspondence should be addressed: the Cancer Research Center, Burnham Institute, 10901 North Torrey Pines Rd., La Jolla, CA 92037. Tel.: 858-646-3100 (ext. 3220); Fax: 858-646-3197; E-mail: irwanm{at}burnham.org.


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