Abstract
BackgroundA defining pathological hallmark of the progressive neurodegenerative disorder Alzheimer’s disease (AD) is the accumulation of misfolded tau with abnormal post-translational modifications (PTMs). These include phosphorylation at Threonine 231 (T231) and acetylation at Lysine 274 (K274) and at Lysine 281 (K281). Although tau is recognized to play a central role in pathogenesis of AD, the precise mechanisms by which these abnormal PTMs contribute to the neural toxicity of tau is unclear.MethodsHuman 0N4R tau (wild type) was expressed in touch receptor neurons of the genetic model organism C. elegans through single-copy gene insertion. Defined mutations were then introduced into the single-copy tau transgene through CRISPR-Cas9 genome editing. These mutations included T231E, to mimic phosphorylation of a commonly observed pathological epitope, and K274/281Q, to mimic disease-associated lysine acetylation – collectively referred as “PTM-mimetics” – as well as a T231A phosphoablation mutant. Stereotypical touch response assays were used to assess behavioral defects in the transgenic strains as a function of age. Genetically-encoded fluorescent biosensors were expressed in touch neurons and used to measure neuronal morphology, mitochondrial morphology, mitophagy, and macro autophagy.ResultsUnlike existing tau overexpression models, C. elegans single-copy expression of tau did not elicit overt pathological phenotypes at baseline. However, strains expressing disease associated PTM-mimetics (T231E and K274/281Q) exhibited reduced touch sensation and neuronal morphological abnormalities that increased with age. In addition, the PTM-mimetic mutants lacked the ability to engage neuronal mitophagy in response to mitochondrial stress.ConclusionsLimiting the expression of tau results in a genetic model where modifications that mimic pathologic tauopathy-associated PTMs contribute to cryptic, stress-inducible phenotypes that evolve with age. These findings and their relationship to mitochondrial stress provides a new perspective into the pathogenic mechanisms underlying AD.
Highlights
A defining pathological hallmark of the progressive neurodegenerative disorder Alzheimer’s disease (AD) is the accumulation of misfolded tau with abnormal post-translational modifications (PTMs)
Plasmid construction Briefly, pBJ1 codes for the fluorescent photo-convertible protein Dendra2 [45], cloned downstream of the mec-7 promoter in a pFH6.II C. elegans expression vector [46]. pBJ2 adds the coding sequence for tau (0N4R) inserted downstream and in-frame with Dendra2. pBJ5 and pBJ6 are derivatives of pBJ1 and pBJ2, respectively, with the tau expression cassette sub-cloned into pCFJ151 (Addgene) to generate Mos-mediated single copy insertion (MosSCI) inserts at the ttTi5605 loci in C. elegans chromosome II [47]. pSKG1 contains a mec-4 promoter driving the expression of C. elegans codon-optimized mito-mKeima
In an attempt to circumvent potential caveats related to overexpression, novel transgenic AD models were engineered using single-copy Mos-transposon mediated insertion of a tau expression cassette into the worm genome [41, 47]
Summary
A defining pathological hallmark of the progressive neurodegenerative disorder Alzheimer’s disease (AD) is the accumulation of misfolded tau with abnormal post-translational modifications (PTMs). These include phosphorylation at Threonine 231 (T231) and acetylation at Lysine 274 (K274) and at Lysine 281 (K281). Alzheimer’s disease (AD) is the most common degenerative brain disease in the aged population It is characterized by the progressive decline of cognition and memory, as well as changes in behavior and personality [1]. Tau isolated from AD brain exhibits a number of post-translational modifications (PTMs); including increases in phosphorylation and acetylation at specific residues [3, 4]. Toxicity appears to result from soluble or oligomeric forms of tau that exhibit increased, disease-associated phosphorylation and acetylation at specific residues altering its turnover and function [7, 8]
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