(A) HEK 293 cells overexpressing V5-tagged canine TRPV4 were transiently transfected with empty vector (none), an ineffective siRNA construct (control), or two different effective siRNA constructs

TRPV4 expression levels were monitored by Western blotting against V5 (with tubulin as a loading control). (B and C) MDCK cells stably expressing tetracycline-inducible TRPV4 II siRNA. siRNA expression in the absence (B) and presence of tetracycline (C) can be monitored by concomitant expression of GFP. (D) Flow-mediated Ca signaling was studied in ciliated MDCK cells using Fura-2. The time course of cytosolic Ca increase in response to laminar flow (ciliary bending) is shown in representative Fura-2 pseudocolor images reflecting the Ca concentration in MDCK cells (340/380 nm fluorescence ratio; blue/green: low [Ca]; yellow/red: high [Ca]; time is indicated in min:s). (E) Flow-induced Ca signals in cells expressing TRPV4 siRNA. Although the flow-induced response was abolished upon knockdown of TRPV4, 10 μM ATP still elicited a robust Ca increase. (F) Single-cell analysis of the Ca signals (ratio 340/380 nm) of the same experiment as shown in D (each trace represents an individual cell and the bold line depicts the mean; for better visibility, only 50% of cells in the visual field are represented). (G) Effect of TRPV4 siRNA on flow-induced Ca signals. Single-cell analysis of the same experiment as shown in E; again, 50% of cells in the visual field are represented. (H) Grouped data from three independent experiments ( = 3; −tet: 75 cells per ; +tet: 93 cells per ). All cells in the visual field were included to calculate the flow-induced Ca peak baseline. *, statistical significance. Error bars represent mean values ± SEM. Bars, 10 μm.<p><b>Copyright information:</b></p><p>Taken from "TRPP2 and TRPV4 form a polymodal sensory channel complex"</p><p></p><p>The Journal of Cell Biology 2008;182(3):437-447.</p><p>Published online 11 Aug 2008</p><p>PMCID:PMC2500130.</p><p>