We compared the pathogenic effects of three variants of human tau with point mutations in the microtubule-binding domain that control the propensity of the protein to acquire a β-sheet conformation. Following ICV injection of AAV-tau in the mouse forebrain, we showed motor deficits in the rotarod task caused by tau overexpression. The pro-aggregant P301S tau mutant led to motor impairments that worsened over time. The WT form of tau also induced behavioral deficits, which however did not significantly progress. In contrast, tau overexpression had no effects on motor behavior when β-sheet breaking proline residues were introduced in the microtubule-binding region.