Impact of Cyclin-Dependent Kinase Inhibition with Roscovitine on Actin Cytoskeleton and Intracellular Calcium Signaling in Renal Cells: Dual Effect of the S and R Enantiomer

Background: Cyclin-dependent kinases (CDK) inhibitors represent interesting therapeutic candidates due to their ability to target cell cycle proteins. Of these, roscovitine is currently entering phase II clinical trials against cancers and phase I clinical tests against glomerulonephritis because reduces the abnormal cell proliferation. The two roscovitine enantiomers (R and S) are both very promising therapeutic tools due to their ability to regulate cell proliferation, however they might exert distinct actions at tissue levels. Here we evaluated roscovitine effect on actin cytoskeleton and intracellular calcium signaling in MDCK cells. Methods: Intracellular calcium was evaluated by Fura-2AM microfluorimetry. Actin filaments were visualized by phalloidin-TRITC. Results: The two enantiomers had opposite effects on actin organization as R-roscovitine caused actin depolymerization whereas S-roscovitine stabilized actin filaments. Long term R-roscovitine treatment significantly reduced basal cytosolic calcium compared to control cells. In contrast, S-roscovitine treated cells showed a significant increase in basal intracellular calcium. Short term exposure to S-roscovitine induced a cytosolic calcium peak, which was abolished after store depletion with cyclopiazonic acid (CPA). Instead R-roscovitine caused cytosolic calcium oscillations followed by a small calcium plateau. Calcium oscillations were prevented after store depletion with CPA or treatment with the PLC inhibitor U73122. Bafilomycin, a selective vacuolar H+-ATPase inhibitor abolished the small calcium plateau. Conclusions: To our knowledge this is the first study revealing the differential effect of S- and R-roscovitine on cytoskeleton and intracellular calcium signaling in renal cells. Since calcium and CDKs are pleiotropic cellular regulators and both exert powerful effects on cell proliferation and regulation of membrane transporter trafficking through actin dynamics, the use of S- and R-roscovitine as therapeutic tools has to be carefully evaluated.