Study of proline-directed protein kinases involved in phosphorylation of the heavy neurofilament subunit.
The high-molecular-mass neurofilament subunit (NFH) is normally hypophosphorylated in the neuronal perikaryon and undergoes extensive phosphorylation after entering the initial axon segment. Aberrant hyperphosphorylation of perikaryal NFH is a common feature of many neurological diseases. In a previous study (), we demonstrated a correlation between phosphorylation of perikaryal NFH and induction of stress-activated protein kinase (SAPK)-gamma. In this report, we present direct evidence showing that the in vivo activation of SAPKs by an upstream activator (MEKK-1) caused extensive NFH phosphorylation. We also show that stress-activated p38 kinases were not involved in the phosphorylation of perikaryal NFH in cultured dorsal root ganglion neurons and that this process was reversible. SAPKgamma was shown to be located in both the cell body and the neurites of the cultured neurons, suggesting that it is likely to be involved in the phosphorylation of cytoplasmic substrates. These could include neuritic NFH, which is highly phosphorylated despite the demonstrated lack of cyclin-dependent kinase-5 activity in these neurons. Neuritic NFH was also highly phosphorylated in neuronal cultures devoid of Schwann cells, indicating that this form of post-translational modification does not require cues stemming from Schwann cell-axon contacts. Collectively, these findings provide significant new insights into mechanisms involved in NFH phosphorylation in normal neurons and in disease states characterized by aberrant phosphorylation of neurofilaments.
Giasson
B I
BI
McGill University, Department of Biochemistry, Montréal, Québec, Canada H3G 1Y6.
Mushynski
W E
WE
D016475:3T3 Cells / Q000502:physiology; D000818:Animals; D001369:Axons / Q000737:chemistry / Q000201:enzymology; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002478:Cells, Cultured; D051360:Cyclin-Dependent Kinase 5; D018844:Cyclin-Dependent Kinases; D005727:Ganglia, Spinal / Q000737:chemistry / Q000166:cytology / Q000201:enzymology; D051379:Mice; D020929:Mitogen-Activated Protein Kinase Kinases; D020928:Mitogen-Activated Protein Kinases; D016900:Neurofilament Proteins / Q000737:chemistry / Q000378:metabolism*; D009474:Neurons / Q000737:chemistry / Q000201:enzymology* / Q000648:ultrastructure; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D051381:Rats; D013312:Stress, Physiological / Q000378:metabolism; D014162:Transfection; D048051:p38 Mitogen-Activated Protein Kinases
D016900:Neurofilament Proteins; C051774:neurofilament protein H; D011494:Protein Kinases; D051360:Cyclin-Dependent Kinase 5; D017346:Protein Serine-Threonine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C495906:Cdk5 protein, mouse; C495907:Cdk5 protein, rat; D018844:Cyclin-Dependent Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D020929:Mitogen-Activated Protein Kinase Kinases