1998 Fiscal Year Final Research Report Summary
Cellular and Subcellular Immunolocalization of CIC-5 Chloride Channel in the mouse Kidney : Colocalization with H^+-ATPase
| Project/Area Number |
09671153
|
|---|
| Research Category |
Grant-in-Aid for Scientific Research (C)
|
| Allocation Type | Single-year Grants |
|---|
| Section | 一般 |
|---|
| Research Field |
Kidney internal medicine
|
|---|
| Research Institution | Tokyo Medical and Dental University |
|---|
Principal Investigator |
SAKAMOTO Hisato Tokyo Medical and Dental University, School of Medicine, Instructor, 医学部, 助手 (80187046)
|
|---|
| Co-Investigator(Kenkyū-buntansha) |
UCHIDA Shinichi Tokyo Medical and Dental University, School of Medicine, Instructor, 医学部, 助手 (50262184)
|
|---|
| Project Period (FY) |
1997 – 1998
|
| Keywords | CIC-5 chloride channel / cytoplasmic vesicle / endocytosis / H^+-ATPase / low molecular V proteinuria weight / monoclonal antibody / Dent's disease |
|---|
| Research Abstract |
CIC-S chloride channel has attracted special interest in relation to three different inherited renal disease associated with nephrolithiasis, but cellular and subcellular localization of this protein has yet to be detennined. To analyze localization of CIC-5 in the kidney, a CIC-5-specific- rat monoclonal antibody (MAb) was developed to an authentic peptide containing amino acids near the. Carboxyl terminal of CIG-5 using rat lymph node method. Specificity of MAb was established by Western blotting and immunohistochemistry of the stably-CIC-5 transfected mammalian cultured cells. Furthermore the epitope recognizing 5- ammino acid residues was determined by multipin-peptide scanning. With the use of high-resolution immunohistochemical technique using confocal microscope, CIC-5 was found in the apical cytoplasm of proximal tubule cell (PT), as well as in the apical plasma membrane of PT, and in the apical plasma membrane of partial collecting duct cells in mouse kidney. These subcellular localizations were confirmed by transmission electron microscopy. In addition, the cellular localization of CIC-5 in the collecting duct was determined by double staining of the same section with antibodies against CIC-5 and aquaporin-2 (AQP-2), by which an exclusive staining of CIC-5 was found only in the cells without staining of AQP-2.
These findings suggest that CIC-5 might be involved in the endocytosis and/or the H^+-secretion in the proximal tubule cells and the cortical collecting duct type A intercalated cells in mouse kidney. The establishment of CIC-5-specific MAb would provide insight into the pathogenesis of inherited kidney diseases associated with mutations of the CIC-5 gene.
|
