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Dephosphorylation of Alzheimer paired helical filaments by protein phosphatase-2A and -2B
1New York State Institute for Basic Research in Developmental Disabilities, Staten Island 10314.
Abstract:
Microtubule-associated protein tau is abnormally hyperphosphorylated in the brain of patients with Alzheimer disease and in this form is the major protein subunit of the paired helical filaments (PHF), the most prominent lesion of the disease. In this study the dephosphorylation of sparingly soluble PHF, PHF II-tau by brain protein phosphatase (PP)-2A1 and PP-2B, and the resulting biochemical, biological, and structural alterations were investigated. Both of the phosphatases dephosphorylated PHF II-tau at the sites of Ser-199/Ser-202 and partially dephosphorylated it at Ser-396/Ser-404; in addition, PHF II-tau was dephosphorylated at Ser-46 by PP-2A1 and Ser-235 by PP-2B. The relative electrophoretic mobility of PHF II-tau increased after dephosphorylation by either enzyme. Divalent cations, manganese, and magnesium increased the activities of PP-2A1 and PP-2B toward PHF II-tau. Dephosphorylation both by PP-2B and PP-2A1 decreased the resistance of PHF II-tau to proteolysis by the brain calcium-activated neutral proteases (CANP). The ability of PHF II-tau to promote the in vitro microtubule assembly was restored after dephosphorylation by PP-2A1 and PP-2B. Microtubules assembled by the dephosphorylated PHF II-tau were structurally identical to those assembled by bovine tau used as a control. The dephosphorylation both by PP-2A1 and PP-2B caused dissociation of the tangles and the PHF; some of the PHF dissociated into straight protofilaments/subfilaments. Approximately 25% of the total tau was released from PHF on dephosphorylation by PP-2A1. These observations demonstrate that PHF II-tau is accessible to dephosphorylation by PP-2A1 and PP-2B, and dephosphorylation makes PHF dissociate, accessible to proteolysis by CANP, and biologically active in promoting the assembly of tubulin into microtubules.
Insights
Dephosphorylating abnormal tau protein (PHF II-tau) with phosphatases PP-2A1 and PP-2B reverses its disease-associated changes. This process restores tau
Area of Science:
- Neuroscience
- Biochemistry
- Cell Biology
Background:
- Abnormal hyperphosphorylation of tau protein is a hallmark of Alzheimer disease, forming paired helical filaments (PHF).
- PHF tau is resistant to degradation and impairs microtubule assembly, contributing to neurodegeneration.
- Protein phosphatases play a crucial role in regulating protein phosphorylation states.
Purpose of the Study:
- To investigate the dephosphorylation of sparingly soluble PHF tau (PHF II-tau) by brain protein phosphatases PP-2A1 and PP-2B.
- To analyze the biochemical, biological, and structural consequences of PHF II-tau dephosphorylation.
Main Methods:
- Dephosphorylation assays using purified PHF II-tau and recombinant PP-2A1 and PP-2B.
- Analysis of phosphorylation sites using electrophoretic mobility.
- Assessing susceptibility to proteolysis by calcium-activated neutral proteases (CANP).
- Evaluating the ability to promote microtubule assembly in vitro.
- Structural analysis of PHF and assembled microtubules.
Main Results:
- PP-2A1 and PP-2B dephosphorylated PHF II-tau at specific serine residues, increasing its electrophoretic mobility.
- Dephosphorylation reduced PHF II-tau's resistance to CANP proteolysis and caused dissociation of PHF and tangles.
- Microtubule assembly activity was restored in tau dephosphorylated by PP-2A1 and PP-2B.
Conclusions:
- PHF II-tau is a substrate for PP-2A1 and PP-2B, indicating these phosphatases can reverse pathological tau modifications.
- Dephosphorylation renders PHF tau prone to degradation and restores its physiological function in microtubule assembly.
- Targeting tau dephosphorylation may offer a therapeutic strategy for Alzheimer disease.