Department of Pediatrics, Peking University First Hospital, Beijing 100034, China
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History+
Received
Revised
Published
2005-09-20
1900-01-01
2005-11-15
Issue Date
2005-11-15
Abstract
ObjectiveTo investigate the effect of NPHS2 gene mutation of both V165X(467-468insT) and R168H (503G>A) on the expression and distribution of podocins in HEK293 cells. MethodsThe wild-type, V165X and R168H mutant cDNAs in the expression plasmids were transfected into HEK293 cells, and the subcellular localization of the wild-type, mutant podocins was studied by immunofluorescence staining with a specific podocin N-terminal antibody (P21) and a specific podocin C-terminal antibody (P35), immunolabeling and confocal microscopy. ResultsThe fluorescence stainings of the wild-type, V165X and R168H mutant podocins with antibody P21 were positive. The fluorescence stainings of the wild-type and R168H mutant podocins with antibody P35 were also positive, whereas the staining of V165X mutant podocin with P35 was negative. The staining for wild-type podocin was distributed around nuclei and mainly on the cell membrane surface in a filamentous pattern, whereas V165X and R168H mutant podocins staining localized predominantly around nuclei with a loss of surface expression. ConclusionsBoth the molecular structure and the subcellular localization of V165X mutant podocin are changed evidently, so is the subcellular localization of R168H mutant podocin, whereas the molecular structure of R168H mutant podocin is little changed. The normal biological functions of the wild podocinrely onits normal molecular structure and subcellular localization as well.
YU Zi-hua;DING Jie;FAN Qing-feng;GUAN Na;WANG Yun-feng;BU Ding-fang.
Expression of V165X and R168H mutant podocins in mammalian cells[J]. Chinese Journal of Nephrology, 2005, 21(11): 659-663.
