title: Tissue transglutaminase inhibits the TRPV5-dependent calcium transport in an N-glycosylation-dependent manner.
abstract: Tissue transglutaminase (tTG) is a multifunctional Ca(2+)-dependent enzyme, catalyzing protein crosslinking. The transient receptor potential vanilloid (TRPV) family of cation channels was recently shown to contribute to the regulation of TG activities in keratinocytes and hence skin barrier formation. In kidney, where active transcellular Ca(2+) transport via TRPV5 predominates, the potential effect of tTG remains unknown. A multitude of factors regulate TRPV5, many secreted into the pro-urine and acting from the extracellular side. We detected tTG in mouse urine and in the apical medium of polarized cultures of rabbit connecting tubule and cortical collecting duct (CNT/CCD) cells. Extracellular application of tTG significantly reduced TRPV5 activity in human embryonic kidney cells transiently expressing the channel. Similarly, a strong inhibition of transepithelial Ca(2+) transport was observed after apical application of purified tTG to polarized rabbit CNT/CCD cells. Furthermore, tTG promoted the aggregation of the plasma membrane-associated fraction of TRPV5. Using patch clamp analysis, we observed a reduction in the pore diameter after tTG treatment, suggesting distinct structural changes in TRPV5 upon crosslinking by tTG. As N-linked glycosylation of TRPV5 is a key step in regulating channel function, we determined the effect of tTG in the N-glycosylation-deficient TRPV5 mutant. In the absence of N-linked glycosylation, TRPV5 was insensitive to tTG. Taken together, these observations imply that tTG is a novel extracellular enzyme inhibiting the activity of TRPV5. The inhibition of TRPV5 occurs in an N-glycosylation-dependent manner, signifying a common final pathway by which distinct extracellular factors regulate channel activity.
authors.affiliation: 286 Department of Physiology, Radboud University Nijmegen Medical Centre, 6500 HB Nijmegen, The Netherlands.
authors.forename: Sandor
authors.initials: S
authors.lastname: Boros
authors.forename: Qi
authors.initials: Q
authors.lastname: Xi
authors.forename: Henrik
authors.initials: H
authors.lastname: Dimke
authors.forename: Annemiete W
authors.initials: AW
authors.lastname: van der Kemp
authors.forename: Kukiat
authors.initials: K
authors.lastname: Tudpor
authors.forename: Sjoerd
authors.initials: S
authors.lastname: Verkaart
authors.forename: Kyu Pil
authors.initials: KP
authors.lastname: Lee
authors.forename: René J
authors.initials: RJ
authors.lastname: Bindels
authors.forename: Joost G
authors.initials: JG
authors.lastname: Hoenderop
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D006031:Glycosylation; D057809:HEK293 Cells; D006801:Humans; D017136:Ion Transport; D011817:Rabbits; D050916:TRPV Cation Channels / Q000037:antagonists & inhibitors / Q000502:physiology*; D011503:Transglutaminases / Q000502:physiology*
chemical_list: D050916:TRPV Cation Channels; C492239:TRPV5 protein, human; D011503:Transglutaminases; D002118:Calcium