This project focuses on the formation of the fibrillar pathology in AD and on its chief component, tau protein. We hypothesize that progressive modification and polymerization of tau proteins into straight and paired helical filaments (PHFs) result from conformational changes that can be identified and quantified using specific monoclonal antibodies. Conformation-selective antibodies, most of which were produced in the laboratory of Peter Davies at Albert Einstein College of medicine, can be used to determine th evolution of cytoskeletal abnormalities as a function of AD progression in vulnerable brain regions, and to correlate these with structural features of tau. Using these antibodies, the stability of the PHF- or AD-selective tau conformations can be assessed and quantified through affinity measurements against recombinant monomeric tau, synthetic polymeric tau, and authentic PHF tau. We propose that early changes in tau phosphorylation and /or conformation lead to the formation of fibrillar pathology in AD, and that these changes are stabilized and can be observed very early in the disease process using early marker monoclonal antibodies prior to the onset of PHF formation. This hypothesis will be tested as follows; 1. We will model the structural features of tau in vitro that underlie binding of PHF-selective antibodies in situ. This will be accomplished through oligonucleotide-directed mutagenesis and affinity measurements. Both phosphorylation-dependent and conformation-dependent antibodies will be characterized. This information will suggest a progression for tau conformation-dependent antibodies will be characterized. This information will suggest a progression for tau conformational changes associated with the formation of the fibrillar pathology; 2. We propose to select novel monoclonal conformation-sensitive tau antibodies for analysis and use in Aims 1 and 3, respectively; 3. We will probe for early conformational changes in tau protein that occur during disease progression using quantitative immunohisto- and cytochemistry at the light and E.M. level; and, 4. We propose to confirm that the staining documented in situ is due to conformationally altered tau by using two site capture ELISAs in vulnerable brain regions in aged control populations, mild, moderate and severe AD. These studies will be performed in conjunction with projects to correlate the formation of the fibrillar pathology with galanin hyperinervation of the basal forebrain cholinergic neurons and expression of specific NGF receptors. Data will be further correlated with structural atrophy as measured by MRI and behavioral symptomatology as well as electrophysiological measures of function.
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