Conserved Lysine Acetylation within the Microtubule-Binding Domain Regulates MAP2/Tau Family Members
Fig 4
MAPs are subject to auto-acetylation within the MTBR domain
A) Recombinant tau-K18 (residues 244–372) or MAP2c (residues 280–388) fragments were incubated in acetylation reactions containing [14C]-labeled acetyl-CoA followed by SDS-PAGE and phosphorimaging using Storm software to detect radiolabeled MAPs. B) Immunoprecipitated 2N4R tau (lane 1), 2N4R-2CA mutant tau (lane 2), or MAP2c proteins (lane 6) derived from QBI-293 lysates were immobilized on agarose beads, incubated with [14C]-Acetyl-CoA, and analyzed by SDS-PAGE and Coomassie staining followed by phosphorimaging analysis. Replicate IgG control samples (lanes 3–5) were used to clearly separate signal intensities from tau and MAP2c sample lanes. C) Purified MTBR fragments from wild-type MAP2c (280–388), a comparable MAP2c fragment containing a cysteine→alanine substitution (C348A), or MAP4 (925–1102) were incubated in acetylation reactions and direct incorporation of radiolabeled acetyl groups was quantified using a liquid scintillation analyzer. Shown are representative analysis using N = 3 technical replicates from N = 3 independent experiments.