ROCK2 kinase activity is required for the effects of ROCK2 on tau. Representative immunoblots of kinase knockdown compared with mock or scramble (SCR) controls. Unless otherwise noted, n = 3 biological replicates per condition. A, ROCK1 shRNA (R1₁ and R1₂) reduced tau (***p = 0.0003) and S6K (***p = 0.0005). ROCK2 shRNA (R2₁ and R2₂) reduced tau, S6K (***p < 0.0001), and pMTOR (normalized to total mTOR; **p = 0.0016). B, R1₁ (**p = 0.005) and R2₂ (**p = 0.0026) reduced relative tau mRNA level. n = 6 biological replicates per condition. C, S6K knockdown reduced pmTOR and tau (***p < 0.0001). D, Tau accumulates in bafilomycin-treated samples. SCR, *p = 0.0228; R1₁, **p = 0.0013; R2₁, **p = 0.0057. E, ROCK2 expression reversed S6K and tau levels compared with R2₁ plus vector control (***p = 0.0002, ***p = 0.0004) or L121G (**p = 0.0039, ***p = 0.0001). All data are expressed as the percentage of the mean ± SEM.