Reversible phosphorylation of tau to form A68 in heat-shocked neuronal PC12 cells.
A68, the primary protein constituent of Alzheimer's disease-associated neurofibrillary tangles, is an abnormally phosphorylated form of the microtubule-associated protein tau. We find that A68 is formed in neuronal PC12 cells when the cells are subjected to a heat shock (45 degrees C for 30 min). A68 was identified by immunoprecipitation with two different anti-tau antibodies (tau-2 and Alz50). Upon separation by SDS-polyacrylamide gel electrophoresis, the tau immunoprecipitates from heat-shocked cells exhibited an additional polypeptide of reduced electrophoretic mobility (approximately 68 kDa) when compared to control cells. A68 was formed with heat shock in the presence of cycloheximide, suggesting that its production occurred by post-translational modification of existing polypeptides. The tau/A68 polypeptides were identified as phosphoproteins by incorporation of 32P into the immunoprecipitates. The phosphorylation of tau to form A68 was reversed with recovery of the intact cells from the heat shock. Finally, immunoprecipitation of lysates from heat-shocked cells with antibodies to heat shock protein (hsp) 72/73 resulted in co-precipitation of tau with hsp 72, which indicates a stable complex formation between these two proteins. On the other hand, A68 remained unassociated with hsp during the heat shock. These results suggest that tau is reversibly phosphorylated to form A68 in neuronal PC12 cells under conditions of stress.
Wallace
W
W
Department of Psychiatry, Mount Sinai School of Medicine, New York, NY.
Johnson
G
G
Sugar
J
J
Merril
C R
CR
Refolo
L M
LM
D000818:Animals; D000906:Antibodies; D001345:Autoradiography; D004591:Electrophoresis, Polyacrylamide Gel; D006358:Hot Temperature; D007700:Kinetics; D009419:Nerve Tissue Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism*; D016716:PC12 Cells; D010766:Phosphorylation; D013462:Sulfur Radioisotopes; D013997:Time Factors; D016875:tau Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*
D000906:Antibodies; C489131:Mapt protein, rat; D009419:Nerve Tissue Proteins; D013462:Sulfur Radioisotopes; D016875:tau Proteins