Effects of phosphorylation of the neurofilament L protein on filamentous structures.

Abstract

Effects of phosphorylation of the neurofilament L protein (NF-L) on the reassembly system were studied by both sedimentation experiments and low-angle rotary shadowing. Bovine spinal cord NF-L was phosphorylated with 3-4 mol/mol protein by either the catalytic subunit of cAMP-dependent protein kinase or protein kinase C. Phosphorylated NF-L could not assemble into filaments. Phosphorylation by either cAMP-dependent protein kinase or protein kinase C inhibited the same step of the reassembly process. Phosphorylated NF-L remained as an 8-chain complex even in favorable conditions for reassembly. The extent of the effect of phosphorylation on the filamentous structure of NF-L was also investigated by using the catalytic subunit of cAMP-dependent protein kinase. The amount of unassembled NF-L increased linearly with increased phosphorylation in the sedimentation experiments. Structural observations indicated that 1 or 2 mol of phosphorylation is enough to inhibit reassembly and to induce disassembly, and the disassembly process was also observed. The filaments were shown to unravel with disassembly. Star-like clusters, which we reported as being the initial stage of reassembly, were also identified.

PKN Associates and Phosphorylates the Head-Rod Domain of Neurofilament Protein

“…Dosemeci and co-workers (49, 52) reported that bovine NF preparation contains known Ca 2ϩ /calmodulin-dependent kinase, PKC, and PKA as well as activatorindependent kinase activities. In vitro phosphorylation of NF at the amino-terminal domain by PKA or PKC inhibits the assembly process of NF and induces disassembly of preexisting filaments (53). In the present study, we revealed that PKN phosphorylated NF at the amino-terminal domain, was able to inhibit the assembly of NFL in vitro, and that PKN was another candidate for being a regulator of NF assembly.…”

The Extensive Nitration of Neurofilament Light Chain in the Hippocampus Is Associated with the Cognitive Impairment Induced by Amyloid β in Mice

“…Hyperphosphorylation of the serine residues of NFL could lead to disruption of the subtle regulation of the NF network (Hisanaga et al, 1990;Nixon and Shea, 1992). After being nitrated in vitro, NFL is not able to form the NF assembly (Crow et al, 1997).…”

Neurofilaments at a glance

“…Phosphorylation of the NF head domain is mediated by the second messenger dependent kinases protein kinase A (PKA) and C (PKC) (Sihag et al, 1988;Sihag and Nixon, 1989;Sihag and Nixon, 1990) and possibly also by Cam kinase II (Hashimoto et al, 2000). The occurrence of NF head phosphorylation in the cell body soon after subunit synthesis reflects its suggested role in maintaining the disassembled state of NFs, as head phosphorylation of NFL inhibits NF assembly (Hashimoto et al, 2000;Hisanaga et al, 1990;Sihag et al, 1999;Sihag and Nixon, 1991). Phosphorylation of the NF head domain is also known to modulate their interaction with fodrin, an important protein of the sub-axolemmal cytoskeleton (Frappier et al, 1987).…”