© The Rockefeller University Press,
0021-9525/1997//703 $5.00
The Journal of Cell Biology, Volume 137, Number 3,
, 1997 703-714
Tyrosine Phosphorylation at a Site Highly Conserved in the L1 Family of Cell Adhesion Molecules Abolishes Ankyrin Binding and Increases Lateral Mobility of Neurofascin
Timothy D. Garver*,
Qun Ren
,
Shmuel Tuvia*, and
Vann Bennett*,
,
* Howard Hughes Medical Institute,
Department of Cell Biology, and
Department of Biochemistry,
Duke University Medical Center, Durham, North Carolina 27710
This paper presents evidence that a member of the L1 family of ankyrin-binding cell adhesion molecules is a substrate for protein tyrosine kinase(s) and phosphatase(s), identifies the highly conserved FIGQY tyrosine in the cytoplasmic domain as the principal site of phosphorylation, and demonstrates that phosphorylation of the FIGQY tyrosine abolishes ankyrin-binding activity. Neurofascin expressed in neuroblastoma cells is subject to tyrosine phosphorylation after activation of tyrosine kinases by NGF or bFGF or inactivation of tyrosine phosphatases with vanadate or dephostatin. Furthermore, both neurofascin and the related molecule Nr-CAM are tyrosine phosphorylated in a developmentally regulated pattern in rat brain. The FIGQY sequence is present in the cytoplasmic domains of all members of the L1 family of neural cell adhesion molecules. Phosphorylation of the FIGQY tyrosine abolishes ankyrin binding, as determined by coimmunoprecipitation of endogenous ankyrin and in vitro ankyrin-binding assays. Measurements of fluorescence recovery after photobleaching demonstrate that phosphorylation of the FIGQY tyrosine also increases lateral mobility of neurofascin expressed in neuroblastoma cells to the same extent as removal of the cytoplasmic domain. Ankyrin binding, therefore, appears to regulate the dynamic behavior of neurofascin and is the target for regulation by tyrosine phosphorylation in response to external signals. These findings suggest that tyrosine phosphorylation at the FIGQY site represents a highly conserved mechanism, used by the entire class of L1-related cell adhesion molecules, for regulation of ankyrin-dependent connections to the spectrin skeleton.
1. Abbreviations used in this paper: Ng-CAM, neuroglial cell adhesion molecule; Nr-CAM, Ng-CAM–related cell adhesion molecule.
Dr. Oxana Tsygankova is gratefully acknowledged for initial studies demonstrating tyrosine phosphorylation of neurofascin and for preparation of B104 cell lines stably transfected with first generation of HA-tagged neurofascin constructs. Dr. Fen Zhang is gratefully acknowledged for preparation of initial HA-tagged neurofascin constructs.
We also wish to thank Dr. Tobias Meyer for invaluable assistance in our photobleaching experiments.
Please address all correspondence to Vann Bennett, Howard Hughes Medical Institute, Department of Cell Biology, and Department of Biochemistry, Duke University Medical Center, Durham, NC 27710; Tel.: (919) 684-3538; Fax: (919) 684-3590.

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