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  • Title: Beta1-integrins in the primary cilium of MDCK cells potentiate fibronectin-induced Ca2+ signaling.
    Author: Praetorius HA, Praetorius J, Nielsen S, Frokiaer J, Spring KR.
    Journal: Am J Physiol Renal Physiol; 2004 Nov; 287(5):F969-78. PubMed ID: 15226154.
    Abstract:
    Because beta(1)-integrin is involved in sensing of fluid flow rate in endothelial cells, a function that in Madin-Darby canine kidney (MDCK) cells is confined to the primary cilium, we hypothesized beta(1)-integrin to be an important part of the primary ciliary mechanosensory apparatus in MDCK cells. We observed that beta(1)-integrin, alpha(3)-integrin, and perhaps alpha(5)-integrin were localized to the primary cilium of MDCK cells by combining lectin and immunofluorescence confocal microscopy. beta(1)-Integrin was also colocalized with tubulin to the primary cilia of the rat renal collecting ducts, as well as to the cilia of proximal tubules and thick ascending limbs. Immunogold-electron microscopy confirmed the presence of beta(1)-integrin on primary cilia of MDCK cells and rat collecting ducts. Intracellular Ca(2+) levels, monitored by fluorescence microscopy on fluo 4-loaded MDCK cells, significantly increased on addition of fibronectin, a beta(1)-integrin ligand, to mature MDCK cells with an IC(50) of 0.02 mg/l. In immature, nonciliated cells or in deciliated mature cells, the IC(50) was 0.40 mg/l. Blocking the fibronectin-binding sites of beta(1)-integrin with RGD peptide prevented the Ca(2+) signal. Cross-linking of beta(1)-integrins by Sambucus nigra agglutinin produced a Ca(2+) response similar to the addition of fibronectin. Furthermore, the fibronectin-induced response was not dependent on flow or a flow-induced Ca(2+) response. Finally, the flow-induced Ca(2+) response was not prevented by the fibronectin-induced signal. Although beta(1)-integrin on the primary cilium greatly potentiates the fibronectin-induced Ca(2+) signaling in MDCK cells, the flow-dependent Ca(2+) signal is not mediated through activation of beta(1)-integrin.
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