The biochemical parameters of tau pathology

1) the quantitative aspects, using western blots

Native tau proteins are normally phosphorylated on numerous serine or threonine sites. In a living cell, there are different pools of tau proteins with different states of phosphorylation. The more tau are phosphorylated, the less they bind to microtubules. Also, the state of phosphorylation of tau proteins is likely different according to the cell compartments. Tau are less phosphorylated in axons, as demonstrated by monoclonal antibody tau1. Also, phosphorylation of tau proteins is developmentally regulated. The antibodies such as AT8, AD2, PHF-1, that are well known in the Alzheimer field to label tau pathology, also strongly label native tau proteins (reviewed in (Buée et al. 2000)).

A) Quantification of pathological tau proteins:

Alzheimer biochemistry is performed on post-mortem human brains. From the work of (Matsuo et al. 1994), we know that native tau proteins are almost totally dephosphorylated during post-mortem delays. Dephosphorylation results from the strong phosphatasic activity which is released after cell death. But in parallel, aggregated tau proteins that constitute brain lesions are not dephosphorylated, because the phosphatase enzymes are unable to access phosphorylated sites that are buried deep in brain lesions. Therefore, aggregated tau proteins remain phosphorylated after death. Antibodies such as AT8 or AD2 (Buee-Scherrer et al. 1996) specifically detect tau pathology on post-mortem brain tissues (figure 1), due to artefactual dephosphorylation of native tau proteins.

(see 4) the different states of phosphorylation).

As shown in this figure,  a characteristic triplet of pathological tau proteins is exclusively detected in Alzheimer brain extracts from polymodal association brain areas while no trace of tau pathology is observed in the same brain areas from non-demented aged-matched controls. Pathological tau proteins from AD, named PHF-tau, are composed of three main electrophoretic variants designed as Tau 60, 64, 69, as a function of their molecular mass. A minor fourth band named Tau 74 is also detected at 74 kDa (Mulot et al. 1994), (Sergeant et al. 1999). Furthermore the intensity of the detection is proportional to the intensity of NFD and tauopathy. Therefore, a semi-quantification by western blot is able to easily detect and quantify NFD, in good agreement with immunohistochemical observations.

Delacourte A. The molecular parameters of tau pathology. Tau as a killer and a witness. Adv Exp Med Biol 2001;487:5-19.

B) Quantification of normal tau proteins

Normal tau proteins are detected using antibodies that are not phospho-dependent. Best antibdies to detect tau proteins are against the amino or carboxy terminal part of tau.

Using this approach, it is possible to detect different profiles that characterize different tauopathies:

1) increase of isoforms are observed in FTDP-17 with mutations in intron 10 of tau gene (due to an abnormal splicing of exon 10)

2) decrease of isoforms with exon 2 or 2+3 is observed in myotonic dystrophy (DM1)

3) a general decrease of all tau isoforms is observed in FTD (DLDH or tauopathy type 0).