Brain Advance Access published online on November 13, 2007
Brain, doi:10.1093/brain/awm260
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
The ß-amyloid protein of Alzheimer's disease increases neuronal CRMP-2 phosphorylation by a Rho-GTP mechanism
1Molecular Neurobiology Laboratory, Department of Biochemistry and Molecular Biology, Monash University, Clayton 3800, 2Department of Pathology, University of Melbourne, Parkville 3010, Victoria, Australia and 3Menzies Research Institute, Hobart, Tasmania 7000, Australia
Correspondence to:
David H. Small, PhD, Molecular Neurobiology Laboratory, Department of Biochemistry and Molecular Biology, Monash University, Clayton 3800, Victoria, Australia. E-mail: david.small{at}med.monash.edu.au
Neuritic abnormalities are a major hallmark of Alzheimer's disease (AD) pathology. Accumulation of ß-amyloid protein (Aß) in the brain causes changes in neuritic processes in individuals with this disease. In this study, we show that Aß decreases neurite outgrowth from SH-SY5Y human neuroblastoma cells. To explore molecular pathways by which Aß alters neurite outgrowth, we examined the activation and localization of RhoA and Rac1 which regulate the level and phosphorylation of the collapsin response mediator protein-2 (CRMP-2). Aß increased the levels of the GTP-bound (active) form of RhoA in SH-SY5Y cells. This increase in GTP-RhoA correlated with an increase in an alternatively spliced form of CRMP-2 (CRMP-2A) and its threonine phosphorylated form. Both a constitutively active form of Rac1 (CA-Rac1) and the Rho kinase inhibitor, Y27632, decreased levels of the CRMP-2A variant and decreased threonine phosphorylation caused by Aß stimulation. The amount of tubulin bound to CRMP-2 was decreased in the presence of Aß but Y27632 increased the levels of tubulin bound to CRMP-2. Increased levels of both RhoA and CRMP-2 were found in neurons surrounding amyloid plaques in the cerebral cortex of the APP(Swe) Tg2576 mice. We found that there was an increase in threonine phosphorylation of CRMP-2 in Tg2576 mice and the increase correlated with a decrease in the ability of CRMP-2 to bind tubulin. The results suggest that Aß-induced neurite outgrowth inhibition may be initiated through a mechanism in which Aß causes an increase in Rho GTPase activity which, in turn, phosphorylates CRMP-2 to interfere with tubulin assembly in neurites.
Key Words: RhoA; Rac1; collapsing response mediator protein-2; amyloid ß peptide; neurite dystrophy of Alzheimer's disease
Abbreviations: APP, amyloid precursor protein; MCI, mild cognitive impairment; PAK, p21-activated kinase; HRP, horse-radish peroxidase; AFM, atomic force microscopy
Received July 24, 2007. Revised September 21, 2007. Accepted September 27, 2007.