Lithium chloride inhibits the phosphorylation of newly synthesized neurofilament protein, NF-M, in cultured chick sensory neurons.
The middle and high molecular weight members of the neurofilament triplet, NF-M and NF-H, undergo extensive posttranslational polyphosphorylation, a process requiring 24 h or more for completion. We have investigated ways of perturbing this process in intact cells and have found that phosphorylation of newly synthesized NF-M in cultured chick sensory neurons is inhibited by Li+. [35S]Methionine pulse-chase experiments were carried out with pure neuronal cultures, and the phosphorylation of newly synthesized NF-M was monitored by following the accompanying change, with chase time, in apparent size and charge of the polypeptide. Addition of LiCl to the medium inhibited this mobility shift in a dose-dependent manner over concentrations between 2 and 25 mM. Incorporation of 32P into NF-M, as well as NF-H, was also inhibited, whereas incorporation into the low molecular weight neurofilament protein, beta-tubulin, and total protein was unaffected. Protein synthesis was not altered. Exposure to 25 mM LiCl for up to 72 h was not toxic, and the inhibition of NF-M phosphorylation was completely reversible. When 25 mM Li+ was added after NF-M had become partially phosphorylated, further progression was blocked, but there was no net dephosphorylation or degradation of NF-M. Additional experiments suggest that this action of Li+ is probably not due to effects on second messenger levels or to effects on tubulin metabolism and assembly state presented in our accompanying article, but rather to interference by Li+ itself, with the phosphorylation of NF-M and NF-H by specific neurofilament kinase(s).
Bennett
G S
GS
Department of Anatomy and Cell Biology, University of Florida, Gainesville 32610.
Laskowska
D
D
DiLullo
C
C
D000818:Animals; D002478:Cells, Cultured; D002642:Chick Embryo; D002712:Chlorides / Q000494:pharmacology*; D003599:Cytoskeleton / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D007381:Intermediate Filament Proteins / Q000037:antagonists & inhibitors* / Q000096:biosynthesis; D008094:Lithium / Q000494:pharmacology*; D018021:Lithium Chloride; D008870:Microtubules / Q000187:drug effects; D016900:Neurofilament Proteins; D009475:Neurons, Afferent / Q000378:metabolism*; D010710:Phosphates / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D013997:Time Factors
D002712:Chlorides; D007381:Intermediate Filament Proteins; D016900:Neurofilament Proteins; D010710:Phosphates; C061195:neurofilament protein M; D008094:Lithium; D018021:Lithium Chloride