Dual modification of Alzheimer’s disease PHF-tau protein by lysine methylation and ubiquitylation: a mass spectrometry approach

Abstract

In sporadic Alzheimer’s disease (AD), neurofibrillary lesion formation is preceded by extensive post-translational modification of the microtubule associated protein tau. To identify the modification signature associated with tau lesion formation at single amino acid resolution, immunopurified paired helical filaments were isolated from AD brain and subjected to nanoflow liquid chromatography–tandem mass spectrometry analysis. The resulting spectra identified monomethylation of lysine residues as a new tau modification. The methyl-lysine was distributed among seven residues located in the projection and microtubule binding repeat regions of tau protein, with one site, K254, being a substrate for a competing lysine modification, ubiquitylation. To characterize methyl lysine content in intact tissue, hippocampal sections prepared from post mortem late-stage AD cases were subjected to double-label confocal fluorescence microscopy using anti-tau and anti-methyl lysine antibodies. Anti-methyl lysine immunoreactivity colocalized with 78 ± 13% of neurofibrillary tangles in these specimens. Together these data provide the first evidence that tau in neurofibrillary lesions is post-translationally modified by lysine methylation.Electronic supplementary materialThe online version of this article (doi:10.1007/s00401-011-0893-0) contains supplementary material, which is available to authorized users.

Tau prions from Alzheimer’s disease and chronic traumatic encephalopathy patients propagate in cultured cells

“…Finally, it is also possible that posttranslational modifications (PTMs) on tau facilitate protein misfolding into distinct prion strains. A large number of PTMs have been identified on tau isolated from both human and Tg mouse samples, including multiple sites for phosphorylation, acetylation, ubiquitination, and O-glycosylation (60)(61)(62). However, none of these differences have been consistently detected in distinct patient groups in a manner that would explain the differences observed here.…”

Tau-induced neurodegeneration: mechanisms and targets

“…T h e m e c h a n i s m l e a d i n g n o r m a l t a u t o b e c o m e hyperphosphorylated remains unknown and posttranslational modifications besides phosphorylation could regulate tau function and aggregation [58] , such as ubiquitination [59] , glycation [60] , glycosylation [61] , nitration [62] , polyamination [63] , proteolysis [64] , acetylation [58] , and methylation [65] .…”

“…Moreover, in other biochemical pathways, certain acetylated Lys residues can be alternatively methylated, suggesting that the web of tau post-translational modifi cations is potentially complex, with Lys-directed modifi cations playing key regulatory roles with respect to rates of tau turnover and aggregation [58] . With the mass spectrometry approach expanding the search criteria to include both acetyl-and methyl-lysine modifications, it was found that seven Lys residues (K44, K163, K174, K180, K254, K267, and K290) in PHF-tau immunopurifi ed from AD brain are monomethylated, and reveal tau methylation as a new tau post-translational modification accompanying PHF deposition in vivo [65] .…”

“…Moreover, in other biochemical pathways, certain acetylated Lys residues can be alternatively methylated, suggesting that the web of tau post-translational modifi cations is potentially complex, with Lys-directed modifi cations playing key regulatory roles with respect to rates of tau turnover and aggregation [58] . With the mass spectrometry approach expanding the search criteria to include both acetyl-and methyl-lysine modifications, it was found that seven Lys residues (K44, K163, K174, K180, K254, K267, and K290) in PHF-tau immunopurifi ed from AD brain are monomethylated, and reveal tau methylation as a new tau post-translational modification accompanying PHF deposition in vivo [65] .The tau phosphorylation state is also modulated by competing modifications of hydroxy amino-acids such as O-linked beta-N-acetylglucosaminylation (O-GlcNAcylation), which is a key regulatory posttranslational modification that is reversible and often reciprocal with phosphorylation of serine and threonine at the same or nearby residues. Gong and coworkers demonstrated that inhibition of O-GlcNAcylation leads to hyperphosphorylation of tau in cultured cells and in rat brain slices [47,71] .…”

Neurochemical Profile of Dementia Pugilistica

“…[82][83][84] Recently, methylation of tau at residue K254, a competing site for ubiquitination, has been added to the post-translational modifications exhibited by these proteins. 85 To assess the various isoforms…”