Modification of Tau by 8-Nitroguanosine 3′,5′-Cyclic Monophosphate (8-Nitro-cGMP): EFFECTS OF NITRIC OXIDE-LINKED CHEMICAL MODIFICATION ON TAU AGGREGATION

Jun Yoshitake,Yoshiyuki Soeda,Tomoaki Ida,Akio Sumioka,Misato Yoshikawa,Kenji Matsushita,Takaaki Akaike,Akihiko Takashima

doi:10.1074/jbc.m116.734350

Abstract

Neurofibrillar tangles caused by intracellular hyperphosphorylated tau inclusion and extracellular amyloid β peptide deposition are hallmarks of Alzheimer's disease. Tau contains one or two cysteine residues in three or four repeats of the microtubule binding region following alternative splicing of exon 10, and formation of intermolecular cysteine disulfide bonds accelerates tau aggregation. 8-Nitroguanosine 3′,5′-cyclic monophosphate (8-nitro-cGMP) acts as a novel second messenger of nitric oxide (NO) by covalently binding cGMP to cysteine residues by electrophilic properties, a process termed protein S-guanylation. Here we studied S-guanylation of tau and its effects on tau aggregation. 8-Nitro-cGMP exposure induced S-guanylation of tau both in vitro and in tau-overexpressed HEK293T cells. S-guanylated tau inhibited heparin-induced tau aggregation in a thioflavin T assay. Atomic force microscopy observations indicated that S-guanylated tau could not form tau granules and fibrils. Further biochemical analyses showed that S-guanylated tau was inhibited at the step of tau oligomer formation. In P301L tau-expressing Neuro2A cells, 8-nitro-cGMP treatment significantly reduced the amount of sarcosyl-insoluble tau. NO-linked chemical modification on cysteine residues of tau could block tau aggregation, and therefore, increasing 8-nitro-cGMP levels in the brain could become a potential therapeutic strategy for Alzheimer's disease.

Highlights

Neurofibrillar tangles caused by intracellular hyperphosphorylated tau inclusion and extracellular amyloid ␤ peptide deposition are hallmarks of Alzheimer’s disease
In polyanion-induced tau aggregation such as by heparin, cysteinedependent interactions occur before ␤ sheet aggregation or cysteine-independent interactions, suggesting that intermolecular disulfide bonds between cysteine residues trigger tau oligomer formation accompanied by cysteine-independent tau interaction (8 –10)
RS-cGMP showed a positive signal in 8-nitro-cGMP-treated wild-type 2N3R (Wild-3R) and 2N4R (Wild-4R) tau but not in vehicle-treated 2N3R and 2N4R tau or 8-nitro-cGMP-treated cysteine-to-alanine substitution mutant tau (Mutant-3R, Mutant-4R) (Fig. 1A)

Summary

Introduction

Neurofibrillar tangles caused by intracellular hyperphosphorylated tau inclusion and extracellular amyloid ␤ peptide deposition are hallmarks of Alzheimer’s disease. NO-linked chemical modification on cysteine residues of tau could block tau aggregation, and increasing 8-nitro-cGMP levels in the brain could become a potential therapeutic strategy for Alzheimer’s disease. These results suggest that 8-nitro-cGMP modulates the cysteine residues of tau and forms S-guanylated tau in vitro.

Results

Conclusion