Characteristics of TBS-Extractable Hyperphosphorylated Tau Species: Aggregation Intermediates in rTg4510 Mouse Brain

Article type: Research Article

Authors: Sahara, Naruhiko^{a; b; *} | DeTure, Michael^b | Ren, Yan^a | Ebrahim, Abdul-Shukkur^{b; 1} | Kang, Dongcheul^b | Knight, Joshua^{a; b} | Volbracht, Christiane^c | Pedersen, Jan Torleif^c | Dickson, Dennis W.^b | Yen, Shu-Hui^b | Lewis, Jada^{a; b}

Affiliations: [a] Center for Translational Research in Neurodegenerative Disease and Department of Neuroscience, University of Florida, Gainesville, FL, USA | [b] Department of Neuroscience, Mayo Clinic, Jacksonville, FL, USA | [c] H. Lundbeck A/S, Copenhagen, Denmark

Correspondence: [*] Correspondence to: Naruhiko Sahara, Department of Neuroscience, Center for Translational Research in Neurodegenerative Disease, University of Florida, 1275 Center Drive, Gainesville, FL 32610, USA. Tel.: +1 352 273 9661; Fax: +1 352 294 5060; E-mail: [email protected].

Note: [1] Current affiliation: Department of Internal Medicine, Wayne State University, Detroit, MI, USA.

Abstract: Conditional overexpression of four-repeat human tau containing the P301L missense mutation in the rTg4510 mouse model of tauopathy leads to progressive accumulation of neurofibrillary tangles and hyperphosphorylated, sarkosyl-insoluble tau species, which are biochemically comparable to abnormal tau characteristic of hereditary tauopathies termed FTDP-17. To fully understand the impact of tau species at different stages of self-assembly on neurodegeneration, we fractionated rTg4510 brain representing several stages of tauopathy to obtain TBS-extractable (S1), high salt/sarkosyl-extractable (S3), and sarkosyl-insoluble (P3) fractions. Under reducing condition, the S1 fraction was demonstrated by western blotting to contain both 50–60 kDa normally-sized and 64 kDa tau. Both are thermo-stable, but the 64 kDa tau showed a higher degree of phosphorylation. Under non-reducing condition, nearly all TBS-extractable 64 kDa tau were detected as ∼130 kDa species consistent with the size of dimer. Quantitative analysis showed ∼80 times more 64 kDa tau in S1 than P3 fraction. Immunoelectron microscopy revealed tau-positive granules/short filaments in S1 fraction. These structures displayed MC1 immunoreactivities indicative of conformational/pathological change of tau. MC1 immunoreactivity was detected by dot blotting in samples from 2.5 month-old mice, whereas Ab39 immunoreactivity indicative of late stages of tau assembly was detected only in P3 fraction. Quantitative analysis also demonstrated a significant inverse correlation between brain weight and 64 kDa tau, but the level of TBS-extractable 64 kDa tau reflects neurodegeneration better than that of sarkosyl-insoluble 64 kDa tau. Together, the findings suggest that TBS-extractable 64 kDa tau production is a potential target for therapeutic intervention of tauopathies.

Keywords: Dimer, FTDP-17, hyperphosphorylation, tau protein, tauopathy, transgenic mice

DOI: 10.3233/JAD-2012-121093

Journal: Journal of Alzheimer's Disease, vol. 33, no. 1, pp. 249-263, 2013