Abstract

Protein tau-3R/4R isoform ratio and phosphorylation regulates binding to microtubules and, when disturbed by aging or mutations, results in diverse tauopathies and in neurodegeneration. The underlying mechanisms were studied here in three transgenic mouse strains with identical genetic background, all expressing the tau-4R/2N isoform driven specifically in neurons by the thy1 gene promoter. Two strains, expressing human tau-4R/2N or mutant tau-4R/2N-P301L at similar, moderate levels, developed very different phenotypes. Tau-4R/2N mice became motor-impaired already around age 6-8 weeks, accompanied by axonopathy (dilatations, spheroids), but no tau aggregates, and surviving normally. In contrast, tau-P301L mice developed neurofibrillary tangles from age 6 months, without axonal dilatations and, despite only minor motor problems, all succumbing before the age of 13 months. The third strain, obtained by tau knock-out/knock-in (tau-KOKI), expressed normal levels of wild-type human tau-4R/2N replacing all mouse tau isoforms. Tau-KOKI mice survived normally with minor motor problems late in life and without any obvious pathology. Biochemically, a fraction of neuronal tau in aging tau-P301L mice was hyperphosphorylated concomitant with conformational changes and aggregation, but overall, tau-4R/2N was actually more phosphorylated than tau-P301L. Significantly, tau with changed conformation and with hyperphosphorylation colocalized in the same neurons in aging tau-P301L mice. Taken together, we conclude that excessive binding of tau-4R/2N as opposed to reduced binding of tau-P301L to microtubules is responsible for the development of axonopathy and tauopathy, respectively, in tau-4R/2N and tau-P301L mice and that the conformational change of tau-P301L is a major determinant in triggering the tauopathy.

Highlights

  • Protein tau-3R/4R isoform ratio and phosphorylation regulates binding to microtubules and, when disturbed by aging or mutations, results in diverse tauopathies and in neurodegeneration

  • We conclude that excessive binding of tau4R/2N as opposed to reduced binding of tau-P301L to microtubules is responsible for the development of axonopathy and tauopathy, respectively, in tau-4R/2N and tauP301L mice and that the conformational change of tauP301L is a major determinant in triggering the tauopathy

  • Generation of Tau-KOKI Mice and Tau-P301L Mice—To generate transgenic mice that lack the heterogeneity of the six tau isoforms due to alternative RNA splicing, we inactivated the endogenous tau gene by introducing into exon 1 a single copy of the human tau-4R/2N cDNA under control of the mouse thy1 promoter

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Summary

TABLE I Specifics of the primary antibodies against protein tau Epitope

Phosphorylation-independent Ser(P)202/Thr(P)205 Thr(P)212/Ser(P)214 Thr(P)231 Thr(P)181 Ser(P)396/Ser(P)404 Ser(P)422 aa 5–15/312–322, conformation-dependent Thr(P)231/Ser(P)235, conformation-dependent Ser(P)409. More confounding factors must act (i.e. tau at different concentrations in different cell types; immunological, hormonal, and nutritional status; cholesterol and lipids; ApoE genotype; and more) This results in different clinical phenotypes, including or not including parkinsonism and motor and behavioral problems, and is reflected at the molecular level in different phosphorylation patterns of tau and in different morphology of tau filaments All three strains express the same tau-4R/2N isoform, with the P301L mutation in the tauP301L mice at moderate levels from the same thy gene promoter and all in the same genetic background This strategy allowed us to separate the early, severe motor deficit in tau4R/2N mice due to axonal dilatations, but not tau aggregates, from the late minor motor impairment in tau-P301L mice, showing widespread intraneuronal tau aggregates and moribund neurodegeneration. This profound distinction is reflected at the molecular level by conformational changes of tau-P301L, not seen in wild-type tau-4R/2N

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