Figure 4. Overexpression of 14-3-3ζ inhibits tau binding to microtubules and reduces the amount of polymerized microtubules in rat hippocampal primary neurons in culture.

Primary neurons infected with Ln-14-3-3ζ or Ln-vector were subjected to a microtubule sedimentation assay. The resulting microtubule pellet (P) and the supernatant (S) were analyzed by Western blotting and the relative distribution of each protein in its respective fraction was determined. The relative distribution value was determined by dividing the band intensity of a protein in the fraction by the total (sum of the intensity value of that protein, both S and P fractions), and is expressed as a % of the total. Values with S.E. are an average of three determinations from three cultures. *p< 0.05 with respect to the P fraction of the Ln-vector control. (B) Relative amount. The relative amounts of polymerized microtubules are relative distribution values from the microtubule pellet in panel (A) and are expressed as the % of Ln-vector control. Likewise, the relative amounts of microtubule-bound tau are the values of total tau in microtubule pellet in panel (A), and are expressed as a % of Ln-vector control. The relative amount of Ser²⁶² phosphorylated tau was determined by normalizing the Ser²⁶² blot by the corresponding tau blot, as in Figure 1. Values are average of three determinations from three cultures. *p<0.05 with respect to the Ln-vector control.