Studies of mechanotransduction mediated by stress-sensitive ion channels generally focus on the site of force application to the cell. Here we show that global, cell-wide changes in cytoskeletal structure and mechanics can regulate mechanotransduction previously shown to be triggered by activation of the mechanosensitive calcium channel, polycystin-2, in the apical primary cilium of renal epithelial cells [S.M. Nauli, F.J. Alenghat, Y. Luo, E. Williams, P. Vassilev, X. Li, A.E. Elia, W. Lu, E.M. Brown, S.J. Quinn, D.E. Ingber, J. Zhou, Polycystins 1 and 2 mediate mechanosensation in the primary cilium of kidney cells. Nat. Genet. 33 (2003) 129–37]. Disrupting cytoplasmic microfilaments or microtubules in these cells eliminated fluid shear stress-induced increase of intracellular calcium. Altering the cytoskeletal force balance by inhibiting actomyosin-based tension generation (using 2,3-butanedione monoxime), interfering with microtubule polymerization (using nocodazole, cochicine, or taxol), or disrupting basal integrin-dependent extracellular matrix adhesions (using soluble GRGDSP peptide or anti-β1 integrin antibody), also inhibited the calcium spike in response to fluid stress. These data indicate that although fluid stress-induced displacement of the primary cilium may be transduced into a calcium spike through activation of polycystin-2 and associated calcium-induced calcium release from intracellular stores, this mechanotransduction response is governed by global mechanical cues, including isometric tension (prestress) within the entire cytoskeleton and intact adhesions to extracellular matrix.