Neurofilament-associated protein phosphatase 2A: Its possible role in preserving neurofilaments in filamentous states

Abstract

Neurofilament phosphatase (NF-phosphatase) activity, which dephosphorylates NF proteins phosphorylated by cyclic AMP-dependent protein kinase (A-kinase), was detected in NF fractions prepared from bovine spinal cords. This phosphatase was suggested to be associated with NFs by gel filtration and sedimentation analysis and was further demonstrated by dephosphorylation-dependent binding assay of NFs to microtubules. The NF-associated NF-phosphatase was identified as a type of protein phosphatase 2A (PP2A) by (i) its complete inhibition with 100 nM okadaic acid, at which concentration the purified type 1 protein phosphatase (PP1) was inhibited only 25%; (ii) the absence of effect of inhibitor-2, a specific inhibitor of PP1, on the NF-phosphatase activity; and (iii) the detection of 38-kDa catalytic and 65-kDa regulatory subunits of PP2A by immunoblotting. The NF-associated PP2A was partially solubilized from NFs by a high concentration of MgSO4, and the solubilized PP2A was suggested by gel filtration to be a dimeric holoenzyme consisting of a 38-kDa catalytic and a 65-kDa regulatory subunit. Phosphorylated NF-L, which is assembly incompetent, was induced to assemble into filaments by dephosphorylation with PP2A. These results suggest a role of NF-associated PP2A in preserving filamentous forms of NF in neurons.

Role of Janus kinase-2 in insulin-mediated phosphorylation and inactivation of protein phosphatase-2A and its impact on upstream insulin signalling components

“…Our observations coincide with a number of recent studies documenting PP-2A association with different kinases [12][13][14][15][16][17][18][19][20][21][22]. For example, studies by Heriche et al [12] indicate that casein kinase 2α associates with PP-2A and causes activation of PP-2A.…”

“…For example, PP-2A has been shown to bind the β2-adrenergic receptor [11], casein kinase 2α [12], homeobox gene 11 [13], tau [14], translation termination eukaryotic release factor 1 [15], adenovirus e4orf4 protein [16], α4 protein [17], neurofilament proteins [18,19] and small t and middle t antigens encoded by DNA tumour viruses [20,21]. Recent studies by Westphal et al [22] identified a calmodulin IV (CaMIV)-PP-2A complex in which PP-2A dephosphorylates…”

Vimentin Dephosphorylation by Protein Phosphatase 2A Is Modulated by the Targeting Subunit B55

“…PP2A is the major protein phosphatase acting on neurofilaments and found associated with this type of IF (Saito et al, 1995). To corroborate an association of PP2A with vimentin, fractions highly enriched in IFs were prepared from Hs68 fibroblasts ( Figure 7A) and immunoblotted for the presence of PP2A subunits.…”

“…We have previously reported that cytokeratins 8 and 18 become hyperphosphorylated when type 2A but not type 1 enzymes are inhibited . PP2A was also identified as a neurofilament phosphatase and, significantly, associates with this type of IF (Saito et al, 1995). Whether PP2A is always targeted to IF by B55 remains to be established.…”

Enzymatic Form and Cytoskeletal Form of Bifunctional Tetrahymena 49kDa Protein Is Regulated by Phosphorylation

“…On the other hand, there are examples that assembly of filaments is as well as regulated by phosphorylation and dephosphorylation. Neurofilaments are major intermediate filaments expressed in neurons, and their assembly is regulated by phosphorylation and dephosphorylation (Gonda et al , 1990;Saito et al , 1995). Reorganization of vimentin and glial fibrillary acidic protein during mitosis are also regulated by phosphorylation (Inagaki et al , 1987;Matsuzawa et al , 1995).…”