title: Urinary β-galactosidase stimulates Ca2+ transport by stabilizing TRPV5 at the plasma membrane.
abstract: Transcellular Ca(2+)transport in the late distal convoluted tubule and connecting tubule (DCT2/CNT) of the kidney is a finely controlled process mediated by the transient receptor potential vanilloid type 5 (TRPV5) channel. A complex-type-N-glycan bound at the extracellular residue Asn358 of TRPV5 through post-translational glycosylation has been postulated to regulate the activity of TRPV5 channels. Using in vitro Ca(2+)transport assays, immunoblot analysis, immunohistochemistry, patch clamp electrophysiology and total internal reflection fluorescence microscopy, it is demonstrated that the glycosidase β-galactosidase (β-gal), an enzyme that hydrolyzes galactose, stimulates TRPV5 channel activity. However, the activity of the non-glycosylated TRPV(N358Q)mutant was not altered in the presence of β-gal, showing that the stimulation is dependent on the presence of the TRPV5N-glycan. In addition, β-gal was found to stimulate transcellular Ca(2+)transport in isolated mouse primary DCT2/CNT cells. β-gal expression was detected in the apical membrane of the proximal tubules, and the protein was found in mouse urine. In summary, β-gal is present in the pro-urine from where it is thought to stimulate TRPV5 activity.
authors.affiliation: Department of Physiology, Radboud University Medical Center, Radboud Institute for Molecular Life Sciences, Nijmegen 6500 HB, The Netherlands.
authors.forename: Elizabeth H P
authors.initials: EH
authors.lastname: Leunissen
authors.affiliation: Department of Physiology, Radboud University Medical Center, Radboud Institute for Molecular Life Sciences, Nijmegen 6500 HB, The Netherlands.
authors.forename: Maxime G
authors.initials: MG
authors.lastname: Blanchard
authors.affiliation: Department of Physiology, Radboud University Medical Center, Radboud Institute for Molecular Life Sciences, Nijmegen 6500 HB, The Netherlands.
authors.forename: Fareeba
authors.initials: F
authors.lastname: Sheedfar
authors.affiliation: Department of Physiology, Radboud University Medical Center, Radboud Institute for Molecular Life Sciences, Nijmegen 6500 HB, The Netherlands.
authors.forename: Marla
authors.initials: M
authors.lastname: Lavrijsen
authors.affiliation: Department of Physiology, Radboud University Medical Center, Radboud Institute for Molecular Life Sciences, Nijmegen 6500 HB, The Netherlands.
authors.forename: Jenny
authors.initials: J
authors.lastname: van der Wijst
authors.affiliation: Department of Physiology, Radboud University Medical Center, Radboud Institute for Molecular Life Sciences, Nijmegen 6500 HB, The Netherlands.
authors.forename: René J M
authors.initials: RJ
authors.lastname: Bindels
authors.affiliation: Department of Physiology, Radboud University Medical Center, Radboud Institute for Molecular Life Sciences, Nijmegen 6500 HB, The Netherlands joost.hoenderop@radboudumc.nl.
authors.forename: Joost G J
authors.initials: JG
authors.lastname: Hoenderop
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D017136:Ion Transport / Q000235:genetics; D007686:Kidney Tubules, Distal / Q000378:metabolism*; D051379:Mice; D008822:Mice, Transgenic; D055550:Protein Stability; D050916:TRPV Cation Channels / Q000235:genetics / Q000378:metabolism*; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism* / Q000652:urine
chemical_list: D015220:Calcium Channels; D050916:TRPV Cation Channels; C492239:TRPV5 protein, human; C492240:Trpv5 protein, mouse; D001616:beta-Galactosidase; D002118:Calcium
keywords: TRPV5; Urine; calcium channel; glycoprotein; post-transcriptional regulation; β-galactosidase