SNAP-25-associated Hrs-2 protein colocalizes with AQP2 in rat kidney collecting duct principal cells

Alok Shukla, Henrik Hager, Thomas Juhl Corydon, Andrew J. Bean, Ronald Dahl, Z. Vajda, Hui Li, Hans Jürgen Hoffmann, Søren Nielsen

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

The vasopressin-induced trafficking of aquaporin-2 (AQP2) water channels in kidney collecting duct is likely mediated by vesicle-targeting proteins (N-ethylmaleimide-sensitive factor attachment protein receptors). Hrs-2 is an ATPase believed to have a modulatory role in regulated exocytosis. To examine whether Hrs-2 is expressed in rat kidney, we carried out RT-PCR combined with DNA sequence analysis and Northern blotting using a digoxigenin-labeled Hrs-2 RNA probe. RT-PCR and Northern blotting revealed that Hrs-2 mRNA is localized in all zones of rat kidney. The presence of Hrs-2 protein in rat kidney was confirmed by immunoblotting, revealing a 115-kDa protein in kidney and brain membrane fractions corresponding to the expected molecular size of Hrs-2. Immunostaining and confocal laser scanning microscopy of LLC-PK1 cells (a porcine proximal tubule cell line) transfected with Hrs-2 DNA confirmed the specificity of the antibody and revealed that Hrs-2 is mainly localized in intracellular compartments, including cathepsin D-containing lysosomal/endosomal compartments. The cellular and subcellular localization of Hrs-2 in rat kidney was examined by immunocytochemistry and confocal laser scanning microscopy. Hrs-2 immunoreactivity was observed in collecting duct principal cells, and weaker labeling was detected in other nephron segments. The labeling was predominantly present in intracellular vesicles, but labeling was also observed in the apical plasma membrane domains of some cells. Colabeling with AQP2 revealed colocalization in vesicles and apical plasma membrane domains, suggesting a role for Hrs-2 in regulated AQP2 trafficking.

Original languageEnglish
JournalAmerican Journal of Physiology - Renal Physiology
Volume281
Issue number3 50-3
Publication statusPublished - 2001

Fingerprint

Collecting Kidney Tubules
Aquaporin 2
Kidney
Proteins
Confocal Microscopy
Northern Blotting
N-Ethylmaleimide-Sensitive Proteins
Cell Membrane
LLC-PK1 Cells
RNA Probes
Digoxigenin
Polymerase Chain Reaction
Cathepsin D
Aquaporins
Antibody Specificity
Nephrons
Exocytosis
Protein Transport
Vasopressins
DNA Sequence Analysis

Keywords

  • Aquaporin-2
  • Collecting duct
  • N-ethylmaleimide-sensitive factor attachment protein receptors
  • Vesicle-targeting receptors

ASJC Scopus subject areas

  • Physiology
  • Physiology (medical)

Cite this

SNAP-25-associated Hrs-2 protein colocalizes with AQP2 in rat kidney collecting duct principal cells. / Shukla, Alok; Hager, Henrik; Corydon, Thomas Juhl; Bean, Andrew J.; Dahl, Ronald; Vajda, Z.; Li, Hui; Hoffmann, Hans Jürgen; Nielsen, Søren.

In: American Journal of Physiology - Renal Physiology, Vol. 281, No. 3 50-3, 2001.

Research output: Contribution to journalArticle

Shukla, A, Hager, H, Corydon, TJ, Bean, AJ, Dahl, R, Vajda, Z, Li, H, Hoffmann, HJ & Nielsen, S 2001, 'SNAP-25-associated Hrs-2 protein colocalizes with AQP2 in rat kidney collecting duct principal cells', American Journal of Physiology - Renal Physiology, vol. 281, no. 3 50-3.
Shukla, Alok ; Hager, Henrik ; Corydon, Thomas Juhl ; Bean, Andrew J. ; Dahl, Ronald ; Vajda, Z. ; Li, Hui ; Hoffmann, Hans Jürgen ; Nielsen, Søren. / SNAP-25-associated Hrs-2 protein colocalizes with AQP2 in rat kidney collecting duct principal cells. In: American Journal of Physiology - Renal Physiology. 2001 ; Vol. 281, No. 3 50-3.
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AU - Hager, Henrik

AU - Corydon, Thomas Juhl

AU - Bean, Andrew J.

AU - Dahl, Ronald

AU - Vajda, Z.

AU - Li, Hui

AU - Hoffmann, Hans Jürgen

AU - Nielsen, Søren

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