|Budget Amount *help
¥6,600,000 (Direct Cost: ¥6,600,000)
Fiscal Year 1997: ¥1,800,000 (Direct Cost: ¥1,800,000)
Fiscal Year 1996: ¥4,800,000 (Direct Cost: ¥4,800,000)
Activin belonging to the TGF-beta superfamily binds to and signals through a receptor complex comprising two transmembrane serine/threonine kinases, called type I and type II.So far, two type II (ActR-IIA,-IIB) and two type I (ActR-IA,IB) activin receptors have been cloned from mammalian sources. In this study, we attempted to elucidate the role of type I receptors in activin signaling for growth, differentiation and apoptosis, and obtained following results.
(1) We established stable HS-72 transfectants overexpressing ActR-IA or ActR-IB by the electroporation procedure. Overexpression of ActR-IA suppressed activin-induced cell-cycle arrest in the G1 phase caused by inhibition of retinoblastoma protein phosphorylation through induction of p21, a cyclin-dependent kinase inhibitor, and subsequent apoptosis. In contrast, HS-72 clones that overexpressed ACtR-IB significantly facilitated activin induced apoptosis. These results indicate that ActR-IA and ActR-IB are distinct from each other in activin signal transduction.
(2) Significant degradation of a cytoskeleton, alpha-fodrin was observed upon activin-induced apoptosis in HS-72 cells. Overexpression of ActR-IA in HS-72 cells caused prevention of the alpha-fodrin degradation, suggesting that the apoptotic signal of activin in relevant to the proteolytic degradation of alpha-fodrin.
(3) Follistatin (activin-binding protein) was found to promote the binding of activin to cell surface heparan sulfate. When the cells were incubated with ^<125>I-activin in the presence of follistatin, significant degradation of activin was observed. This activin degradation was abolished by heparan sulfate, chloroquine, and lysosomal protease inhibitors. These results indicate that cell-associated follistatin accelerates the uptake of activin into cells, leading to increased degradation by lysosomal enzymes, and thus plays a role in the activin clearance system.