Phospholipase C (PLC) is involved in bradykinin (BK) induced focal contact (FC) restructuration

MÁRQUEZ G,; GAGLIANO L; SERRANO D; STERIN-SPEZIALE N

Iguazu, Misiones

Congreso; XL REUNIÓN DE LA SOCIEDAD ARGENTINA DE INVESTIGACIONES BIOQUÍMICAS Y BIOLOGÍA MOLECULAR (SAIB); 2004

SOCIEDAD ARGENTINA DE INVESTIGACIONES BIOQUÍMICAS Y BIOLOGÍA MOLECULAR (SAIB)

FC are structures of cell attachment to extracelular matrix. We previously found that FC proteins vinculin (V) and and talin are associated with DRMs- detergent-resistant membrane domains, that DRMs-phosphatidylinositol 4,5 biphosphate (PIP2) binding to V is essential for FC formation, and that BK modulates FC assembly. Since BK stimulates PIP2 hydrolysis by activating PLC, we tried to determine whether BK can modulate FC assembly by this mechanism. We treated cultured rat renal papillary collecting duct cell with U73122 -an inhibitor of PLC- before BK stimulation and analyzed FC and actin cytoskeleton by confocal microscopy analysis. In untreated cells, intense V staining was localized to FC and colocalized with PIP2, and actin stress fibers were observed throughout the cell body terminating at the FC. After 1 and 5 minutes of BK treatment, V and PIP2 immunostaining was removed from FC, and actin accumulated at the cell periphery with a concomitant loss of stress fibers. BK did not affect at any time talin staining in FC. Pretreatment of cells with U73122 inhibited the BK-induced loss of FC-V staining and provoked a reorganization of the actin stress fibers into a more dense filamentous actin network. An intense FC-PIP2 staining was also observed, accompanied by an accumulation of PIP2 with a granular aspect throughout the cell body. Since talin, but not V, remain localized to FC after BK stimulation, BK could be inducing a restructuration rather than a dissipation of FC by a mechanism that involves DRMs-PIP2 hydrolysis by activation of PLC.