Phosphorylation of neurofilament heavy-chain side-arm fragments by cyclin-dependent kinase-5 and glycogen synthase kinase-3alpha in transfected cells.
The side-arm domain of neurofilament heavy-chain (NF-H) is heavily phosphorylated in axons. Much of this phosphate is located within a multiphosphorylation repeat (MPR) domain situated toward the carboxy terminus of the molecule. The MPR domain contains the repeat motif KSP of which there are two broad categories, KSPXX and KSPXK. In mouse NF-H, the KSPXK repeats are situated toward the latter part of the MPR domain. We have expressed in mammalian cells fragments of mouse NF-H side-arm containing all of the MPR domain, the latter part of the MPR domain containing the KSPXK repeats, and the complementary amino-terminal part of the MPR domain, which contains the KSPXX repeats. By cotransfecting these fragments with the neurofilament kinases cyclin-dependent kinase-5 (cdk-5)/p35 and glycogen synthase kinase-3alpha (GSK-3alpha), we show that cdk-5 induces cellular phosphorylation of the KSPXK-containing fragment of NF-H. Using the transfected fragments, we also map the epitopes for several commonly utilised NF-H monoclonal antibodies and describe the effects that phosphorylation by cdk-5 and GSK-3alpha have on their reactivities.
Bajaj
N P
NP
Department of Neurology, Institute of Psychiatry, London, England, U.K.
Miller
C C
CC
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D019556:COS Cells; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism*; D051360:Cyclin-Dependent Kinase 5; D018844:Cyclin-Dependent Kinases; D038362:Glycogen Synthase Kinase 3; D038341:Glycogen Synthase Kinases; D051379:Mice; D016900:Neurofilament Proteins / Q000235:genetics / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D012091:Repetitive Sequences, Nucleic Acid; D014162:Transfection
D016900:Neurofilament Proteins; D010446:Peptide Fragments; D038341:Glycogen Synthase Kinases; D051360:Cyclin-Dependent Kinase 5; D017346:Protein Serine-Threonine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C495906:Cdk5 protein, mouse; D018844:Cyclin-Dependent Kinases; D038362:Glycogen Synthase Kinase 3