Cytoskeletal regulation of calcium-permeable cation channels in the human syncytiotrophoblast: role of gelsolin

NICOLAS MONTALBETTI, QIANG LI, GUSTAVO A. TIMPANARO, SILVIA GONZALEZ-PERRETT, XIAO-QING DAI,XING-ZHEN CHEN AND HORACIO F. CANTIELLO

THE JOURNAL OF PHYSIOLOGY

Año: 2005 p. 309 - 325

0022-3751

The human syncytiotrophoblast (hST) is the most apical epithelial barrier that covers the villous tree of the human placenta. An intricate and highly organized network of cytoskeletal structures supports the hST. Recently, polycystin-2 (PC2), a TRP-type nonselective cation channel, was functionally observed in hST, where it may be an important player to Ca2+ transport. Little is known, however, about channel regulation in hST. In this report, the regulatory role of actin dynamics on PC2 channels reconstituted fromhST apical membranes was explored. Acute addition of cytochalasin D (CD, 5 ìgml−1) to reconstituted hST apical membranes transiently increased K+-permeable channel activity. The actin-binding proteins á-actinin and gelsolin, as well as PC2, were observed by Western blot and immunofluorescence analyses in hST vesicles. CD treatment of hST vesicles resulted in a re-distribution of actin filaments, in agreement with the effect ofCDon K+ channel activity. In contrast, addition of exogenous monomeric actin, but not prepolymerized actin, induced a rapid inhibition of channel function in hST. This inhibition was obliterated by the presence of CD in the medium. The acute (<15 min) CD stimulation of K+ channel activity was mimicked by addition of the actin-severing protein gelsolin in the presence, but not in the absence, of micromolar Ca2+. Ca2+ transport through PC2 triggers a regulatory feedback mechanism, which is based on the severing and re-formation of filamentous actin near the channels. Cytoskeletal structures may thus be relevant to ion transport regulationin the human placenta.2+ transport. Little is known, however, about channel regulation in hST. In this report, the regulatory role of actin dynamics on PC2 channels reconstituted fromhST apical membranes was explored. Acute addition of cytochalasin D (CD, 5 ìgml−1) to reconstituted hST apical membranes transiently increased K+-permeable channel activity. The actin-binding proteins á-actinin and gelsolin, as well as PC2, were observed by Western blot and immunofluorescence analyses in hST vesicles. CD treatment of hST vesicles resulted in a re-distribution of actin filaments, in agreement with the effect ofCDon K+ channel activity. In contrast, addition of exogenous monomeric actin, but not prepolymerized actin, induced a rapid inhibition of channel function in hST. This inhibition was obliterated by the presence of CD in the medium. The acute (<15 min) CD stimulation of K+ channel activity was mimicked by addition of the actin-severing protein gelsolin in the presence, but not in the absence, of micromolar Ca2+. Ca2+ transport through PC2 triggers a regulatory feedback mechanism, which is based on the severing and re-formation of filamentous actin near the channels. Cytoskeletal structures may thus be relevant to ion transport regulationin the human placenta.