Browsing by Author "Narasaraju, Telugu A."

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Characterization of -soluble n-ethylmaleimide–sensitive fusion attachment protein in alveolar type ii cells implications in lung surfactant secretion
(2002) Abonyo, B. O.; Wang, Pengcheng; Narasaraju, Telugu A.; Rowan III, William H.; Zimmerman, Un-Jin; Lin Liu
N-ethylmaleimide–sensitive fusion protein (NSF) and soluble NSF attachment protein (α-SNAP) are thought to be soluble factors that transiently bind and disassemble SNAP receptor complex during exocytosis in neuronal and endocrine cells. Lung surfactant is secreted via exocytosis of lamellar bodies from alveolar epithelial type II cells. However, the secretion of lung surfactant is a relatively slow process, and involvement of SNAP receptor and its cofactors (NSF and α-SNAP) in this process has not been demonstrated. In this study, we investigated a possible role of α-SNAP in surfactant secretion. α-SNAP was predominantly associated with the membranes in alveolar type II cells as determined by Western blot and immunocytochemical analysis using confocal microscope. Membrane-associated α-SNAP was not released from the membrane fraction when the cells were lyzed in the presence of Ca2+ or Mg2+ATP. The alkaline condition (0.1 M Na2CO3, pH 12), known to extract peripheral membrane proteins also failed to release it from the membrane. Phase separation using Triton X-114 showed that α-SNAP partitioned into both aqueous and detergent phases. NSF had membrane-bound characteristics similar to α-SNAP in type II cells. Permeabilization of type II cells with β-escin resulted in a partial loss of α-SNAP from the cells, but cellular NSF was relatively unchanged. Addition of exogenous α-SNAP to the permeabilized cells increased surfactant secretion in a dose-dependent manner, whereas exogenous NSF has much less effects. An α-SNAP antisense oligonucleotide decreased its protein level and inhibited surfactant secretion. Our results suggest a role of α-SNAP in lung surfactant secretion.
Reorganization of cytoskeleton during surfactant secretion in lung type II cells: a role of annexin II
(Elsevier, 2003) Singh, Taran K.; Abonyo, B. O.; Narasaraju, Telugu A.
The secretion of lung surfactant requires the movement of lamellar bodies to the plasma membrane through cytoskeletal barrier at the cell cortex. We hypothesized that the cortical cytoskeleton undergoes a transient disassembly/reassembly in the stimulated type II cells, therefore allowing lamellar bodies access to the plasma membrane. Stabilization of cytoskeleton with Jasplakinolinde (JAS), a cell permeable actin microfilament stabilizer, caused a dose-dependent inhibition of lung surfactant secretion stimulated by terbutaline. This inhibition was also observed in ATP-, phorbol 12-myristate 13-acetate (PMA)- or Ca2 + ionophore A23187-stimulated surfactant secretion. Stimulation of type II cells with terbutaline exhibited a transient disassembly of filamentous actin (F-actin) as determined by staining with Oregon Green 488 Phalloidin. The protein kinase A inhibitor, H89, abolished the terbutaline-induced F-actin disassembly. Western blot analysis using anti-actin and anti-annexin II antibodies showed a transient increase of G-actin and annexin II in the Triton X-100 soluble fraction of terbutaline-stimulated type II cells. Furthermore, introduction of exogenous annexin II tetramer (AIIt) into permeabilized type II cells caused a disruption in the cortical actin. Treatment of type II cells with N-ethylmaleimide (NEM) resulted in a disruption of the cortical actin. NEM also inhibited annexin II’s abilities to bundle F-actin. The results suggest that cytoskeleton undergoes reorganization in the stimulated type II cells, and annexin II tetramer plays a role in this process. D 2003 Elsevier Inc. All rights reserved.