| Co-Investigator(Kenkyū-buntansha) |
SANO Hitomi Sapporo Medical University, School of medicine, Department of Biochemistry, Inst, 医学部, 助手 (80295344)
KUROKI Yoshio Sapporo Medical University, School of medicine, Department of Biochemistry, Prof, 医学部, 教授 (70161784)
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| Research Abstract |
Pulmonary surfactant, a mixture of lipids and proteins, is synthesized by alveolar type II cells and secreted to alveolar space. Surfactant proteins A and D (SP-A and SP-D) have been shown to play roles in phospholipid homeostasis and host defense in lung. In the present study, we focused on the functional roles of SP-A and SP-D, and on the SP-A binding protein (annexin IV)
1. SP-A and SP-D are homologous proteins and belong to the C-type lectin family. Both proteins bind to phospoholipids and glycolipids : SP-A to diparmitoylphosphatidylcholine (DPPC) and GalCer, and SP-D to phosphtidylinositol (PI) and GluCer. SP-A also inhibits phospholipid secretion by alveolar type II cells and augments uptake of phospohlipids via SP-A receptors, whereas SP-D does not have these properties. Using chimeric proteins with SP-A and SP-D, it was revealed that Glu^<195> to C-terminus of SP-A is important for the functional properties of SP-A and that Glu^<321> to C-terminus and Cys^<261> to C-terminus of
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SP-D are required for PI binding and GluCer binding, respectively.
2. Annexin IV is a member of the annexin family that consists of a unique N-terminal and a C-terminal core region. The core region consists of four consensus 70 amino acid-repeats (domains 1-4), in which Ca^<2+> and phospholipid binding sites are located. Using truncation mutants of annexin IV, SP-A binding site was shown to locate in domain 3 of annexin IV.Moreover, the Ca^<2+>-binding site in domain 3 of annexin IV and Arg^<197> of SP-A were also shown to be important for Ca^<2+>-dependent binding by site-directed mutagenesis study.
3. SP-A has been shown to interact with the Gram-negative bacteria. As LPS is the main constituent of the bacterial membrane, we investigate possible interaction between SP-A and LPS.It was revealed that SP-A bound to both rough LPS and lipid A but not to smooth LPS.LPS-induced augmentation of expression of TNF-alpha was attenuated by SP-A.SP-A was also shown to interact with CD14, which is the smooth LPS receptor. Therefore, SP-A inhibited LPS-induced TNF-alpha expression by competing with LPS for CD14. Less
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