Expression of synaptotagmin VIII in rat kidney.
The synaptotagmins are a family of integral membrane proteins proposed to function as regulators of both exocytosis and endocytosis. Here, we have used immunochemical techniques and RT-PCR to assess sites of renal expression of synaptotagmin VIII. A polyclonal antibody was raised to a synthetic peptide corresponding to the carboxy-terminal 21 amino acids of mouse synaptotagmin VIII. On immunoblots of membrane fractions from renal cortex and medulla (and in several other tissues), the antibody labeled a 52-kDa band (absent with preimmune IgG). Immunofluorescence localization was carried out in tissue sections from rat kidney. The synaptotagmin VIII antibody labeled early proximal tubules, thin ascending limbs, thick ascending limbs, connecting tubules, and collecting ducts. In collecting ducts, both type A and B intercalated cells exhibited basolateral labeling, whereas principal cells were labeled chiefly in the apical and subapical portion of the cells. Thick ascending limbs were labeled in both the basolateral and apical regions. RT-PCR experiments using total RNA extracted from cortex and medulla or microdissected inner medullary collecting ducts gave a single band of appropriate size. Sequencing of the PCR product confirmed that the amplified target is synaptotagmin VIII. We conclude that synaptotagmin VIII is broadly expressed among renal tubule epithelia, raising the possibility that it is involved in regulation of transport and/or cell remodeling at several sites in the nephron and collecting duct.
Kishore
B K
BK
Laboratory of Kidney and Electrolyte Metabolism, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda 20892-0951, Maryland.
Wade
J B
JB
Schorr
K
K
Inoue
T
T
Mandon
B
B
Knepper
M A
MA
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D002135:Calcium-Binding Proteins; D005455:Fluorescent Antibody Technique; D007668:Kidney / Q000166:cytology / Q000378:metabolism*; D007672:Kidney Cortex / Q000166:cytology / Q000378:metabolism; D007679:Kidney Medulla / Q000166:cytology / Q000378:metabolism; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000378:metabolism; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000032:analysis / Q000096:biosynthesis* / Q000737:chemistry; D008565:Membrane Proteins / Q000096:biosynthesis*; D051379:Mice; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000032:analysis / Q000096:biosynthesis* / Q000737:chemistry; D009928:Organ Specificity; D010446:Peptide Fragments / Q000737:chemistry / Q000276:immunology; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000096:biosynthesis; D011817:Rabbits; D051381:Rats; D050857:Synaptotagmins; D014158:Transcription, Genetic
D000906:Antibodies; D002135:Calcium-Binding Proteins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D010446:Peptide Fragments; D012333:RNA, Messenger; C494306:Syt8 protein, mouse; C494307:Syt8 protein, rat; D050857:Synaptotagmins