Effect of Pseudophosphorylation and Cross-linking by Lipid Peroxidation and Advanced Glycation End Product Precursors on Tau Aggregation and Filament Formation

Björn Kuhla,Cathleen Haase,Katharina Flach,Hans-Joachim Lüth,Thomas Arendt,Gerald Münch

doi:10.1074/jbc.m609521200

Abstract

Accumulation of hyperphosphorylated Tau protein as paired helical filaments in pyramidal neurons is a major hallmark of Alzheimer disease. Besides hyperphosphorylation, other modifications of the Tau protein, such as cross-linking, are likely to contribute to the characteristic features of paired helical filaments, including their insolubility and resistance against proteolytic degradation. In this study, we have investigated whether the four reactive carbonyl compounds acrolein, malondialdehyde, glyoxal, and methylglyoxal accelerate the formation of Tau oligomers, thioflavin T-positive aggregates, and fibrils using wild-type and seven pseudophosphorylated mutant Tau proteins. Acrolein and methylglyoxal were the most reactive compounds followed by glyoxal and malondialdehyde in terms of formation of Tau dimers and higher molecular weight oligomers. Furthermore, acrolein and methylglyoxal induced the formation of thioflavin T-fluorescent aggregates in a triple pseudophosphorylation-mimicking mutant to a slightly higher degree than wild-type Tau. Analysis of the Tau aggregates by electron microscopy study showed that formation of fibrils using wild-type Tau and several Tau mutants could be observed with acrolein and methylglyoxal but not with glyoxal and malondialdehyde. Our results suggest that reactive carbonyl compounds, particularly methylglyoxal and acrolein, could accelerate tangle formation in vivo and that this process could be slightly accelerated, at least in the case of methylglyoxal and acrolein, by hyperphosphorylation. Interference with the formation or the reaction of these reactive carbonyl compounds could be a promising way of inhibiting tangle formation and neuronal dysfunction in Alzheimer disease and other tauopathies.

Highlights

The reactivity of the four RCCs tested decreased in the following order: acrolein Ͼ methylglyoxal Ͼ glyoxal Ϸ malondialdehyde
It was suggested that carbonyl-derived post-translational modifications of neurofilaments may account for the biochemical properties of neurofibrillary tangles, possibly as a result of extensive cross-links [22, 39]
By using more reactive carbonyl compounds than glucose in our experiments, we could induce formation of AGE/ALE-modified Tau oligomers and, with acrolein and methylglyoxal, paired helical filaments (PHF)-like structures, which indicated that intensive cross-linking by AGEs or ALEs favor formation of PHFs

Summary

Introduction

The reactivity of the four RCCs tested decreased in the following order: acrolein Ͼ methylglyoxal Ͼ glyoxal Ϸ malondialdehyde. Only acrolein and methylglyoxal, the two most reactive RCCs, formed PHFs in a concentration-dependent manner. It is possible that MDA and glyoxal would have formed oligomers and PHF-like filaments if the reaction had proceeded for a much longer time period. It can be assumed that the reaction of 10 mM acrolein or methylglyoxal with Tau was nearly complete, which could explain the high content of large PHF-like structures that were detected by electron microscopy. When 1 mM acrolein or methylglyoxal were used, more protofibrils and less PHF-like structures were observed. The conversion of Tau proteins with 10 mM MDA and glyoxal within 96 h could have been incomplete, and this may be the cause of why only a few small filament

Methods

Results

Conclusion