SNAP-23 in rat kidney: colocalization with aquaporin-2 in collecting duct vesicles.
Vesicle targeting proteins ("SNAREs") have been proposed to direct vasopressin-induced trafficking of aquaporin-2 water channels in kidney collecting ducts. A newly identified SNARE protein, SNAP-23, is proposed to mediate vesicle targeting to the plasma membrane in diverse tissues. The current studies were done to determine whether SNAP-23 is expressed in collecting ducts with an intracellular distribution compatible with a role in aquaporin-2 trafficking. RT-PCR demonstrated SNAP-23 mRNA in microdissected collecting ducts and other tubular segments including the proximal tubule and thick ascending limb. Immunoblotting using a polyclonal antibody raised against a COOH-terminal peptide revealed a solitary band at an apparent molecular mass of 30 kDa in renal medullary membrane fractions and inner medullary collecting duct suspensions. Differential centrifugation revealed that SNAP-23 is present in membrane fractions including the low-density fraction enriched in intracellular vesicles. Immunocytochemistry revealed SNAP-23 labeling at both the apex and the cytoplasm of collecting duct principal cells. Immunoblotting of intracellular vesicles immunoisolated using an aquaporin-2 antibody revealed the presence of both SNAP-23 and synaptobrevin-2 (VAMP-2) in aquaporin-2-bearing vesicles. We conclude that SNAP-23 is strongly expressed in collecting duct principal cells, consistent with a role in vasopressin-regulated trafficking of aquaporin-2. However, localization of SNAP-23 in both intracytoplasmic vesicles and plasma membranes suggests a function different from that originally proposed for SNAP-25 in synaptic vesicle targeting.
Inoue
T
T
Laboratory of Kidney and Electrolyte Metabolism, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892-1603, USA.
Nielsen
S
S
Mandon
B
B
Terris
J
J
Kishore
B K
BK
Knepper
M A
MA
D000818:Animals; D051399:Aquaporin 2; D051403:Aquaporin 6; D020346:Aquaporins / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D007685:Kidney Tubules, Collecting / Q000378:metabolism*; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism; D016133:Polymerase Chain Reaction; D050766:Qb-SNARE Proteins; D050767:Qc-SNARE Proteins; D050683:R-SNARE Proteins; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D017207:Rats, Sprague-Dawley
C496025:Aqp2 protein, rat; D051399:Aquaporin 2; D051403:Aquaporin 6; D020346:Aquaporins; D002352:Carrier Proteins; D008565:Membrane Proteins; D050766:Qb-SNARE Proteins; D050767:Qc-SNARE Proteins; D050683:R-SNARE Proteins; D012333:RNA, Messenger; C100493:SNAP23 protein, human