Removal of Pattern-breaking Sequences in Microtubule Binding Repeats Produces Instantaneous Tau Aggregation and Toxicity

Asparouh Iliev Iliev,Sundar Ganesan,Gertrude Bunt,Fred Silvester Wouters

doi:10.1074/jbc.m604863200

Abstract

Aggregated and highly phosphorylated tau protein is a pathological hallmark of Alzheimer's disease (AD) and other tauopathies. We identified motifs of alternating polar and apolar amino acids within the microtubule-binding repeats of tau which were interrupted by small breaking stretches. Minimal mutation of these breaking sequences yielded a unique instantly aggregating tau mutant containing longer stretches of polar/apolar amino acids without losing its microtubule-binding capacity. These modifications produced rapid aggregation and cytotoxicity with accompanying occurrence of pathologic tau phosphoepitopes (AT8, AT180, AT270, AT100, Ser(422), and PHF-1) and conformational epitopes (MC-1 and Alz50) in cells. Similar to pathological tau in the pretangle state, toxicity appeared to occur early without the requirement for extensive fibril formation. Thus, our mutant protein provides a novel platform for the investigation of the molecular mechanisms for toxicity and cellular behavior of pathologically aggregated tau proteins and the identification of its interaction partners.

Highlights

Aggregates of hyperphosphorylated tau protein are a pathological hallmark of many neurodegenerative diseases, which include Alzheimer disease (AD),3 frontotemporal dementia with Parkinson syndrome linked to chromosome 17 (FTDP17), and others [1, 2]
It has been shown that the third microtubule-binding repeats (MTBRs) contains a sequence that aggregates in isolated form [6], and the MTBRs have been shown to be located in the core of fibrils by tryptophan fluorescence scanning spectroscopy [4]
Modification of MTBRs Producing an Instantly Aggregating Tau Mutant—We identified motifs of 4 or 5 amino acids consisting of alternating polar/apolar residues in the second, third, and fourth MTBR of the 441-amino acid-long form of human wild-type tau (WT-tau) (Fig. 1, upper panel)

Summary

Introduction

Aggregates of hyperphosphorylated tau protein are a pathological hallmark of many neurodegenerative diseases, which include Alzheimer disease (AD),3 frontotemporal dementia with Parkinson syndrome linked to chromosome 17 (FTDP17), and others [1, 2]. We found a 2–3-fold increase in labeling intensity for PHF-1, AD2 (not shown), and Ser422 antibodies in 3PO-tau-expressing cells (Fig. 4C).

Results

Conclusion