Multiple interactions of aluminum with neurofilament subunits: regulation by phosphate-dependent interactions between C-terminal extensions of the high and middle molecular weight subunits.
Exposure of individual purified neurofilament (NF) proteins to AlCl3 alters their electrophoretic properties in a time- and concentration-dependent manner, as visualized by their failure to migrate into SDS gels. Co-incubation of purified high (NF-H) and middle (NF-M) but not low (NF-L) molecular weight NF subunits prevents this AlCl3-induced alteration in electrophoretic migration. This latter finding suggested that specific interactions between NF-H and NF-M other than filament formation influenced their interaction with AlCl3. Co-incubation of the 160 kDa alpha-chymotryptic cleavage product of NF-H (corresponding to the highly phosphorylated C-terminal sidearm domain) with native NF-M prevented alteration in subunit electrophoretic migration by AlCl3. By contrast, intact, dephosphorylated NF-H subunits were unable to prevent AlCl3-induced alteration of native NF-M electrophoretic migration. Taken together, these findings suggest that phosphate-dependent interactions between the sidearm extensions of NF-H and NF-M diminish the ability of AlCl3 to associate with either subunit in a manner that alters their electrophoretic migration. This interaction of NF-H and NF-M sidearms is SDS-sensitive, while AlCl3-induced alteration in electrophoretic migration of individual subunits is SDS-resistant. Addition of SDS to mixtures of NF-H and NF-M subunits disrupted the protective effect, and promoted AlCl3-induced alterations in subunit electrophoretic migration. These findings support and extend the current hypothesis that the ability of aluminum to interact with NF subunits is a function of subunit phosphorylation, assembly, and extent of neurofilament-neurofilament cross-linking.
Shea
T B
TB
Laboratory for Molecular Neuroscience, McLean Hospital, Belmont, Massachusetts 02178.
Beermann
M L
ML
D000535:Aluminum / Q000737:chemistry*; D000818:Animals; D002417:Cattle; D002918:Chymotrypsin; D003599:Cytoskeleton / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D006868:Hydrolysis; D015151:Immunoblotting; D008970:Molecular Weight; D016900:Neurofilament Proteins / Q000737:chemistry*; D010710:Phosphates / Q000737:chemistry*; D010766:Phosphorylation; D013116:Spinal Cord / Q000737:chemistry
D016900:Neurofilament Proteins; D010710:Phosphates; D000535:Aluminum; D002918:Chymotrypsin