80K-H as a new Ca2+ sensor regulating the activity of the epithelial Ca2+ channel transient receptor potential cation channel V5 (TRPV5).
The epithelial Ca(2+) channel transient receptor potential cation channel V5 (TRPV5) constitutes the apical Ca(2+) entry pathway in the process of active Ca(2+) reabsorption. Ca(2+) influx through TRPV5 is tightly controlled by modulators of Ca(2+) homeostasis, including 1,25-dihydroxyvitamin D(3) and dietary Ca(2+). However, little is known about intracellular proteins that interact with TRPV5 and directly regulate the activation of this channel. By the use of cDNA microarrays, the present study identified 80K-H as the first protein involved in the Ca(2+)-dependent control of the epithelial Ca(2+) channel TRPV5. 80K-H was initially identified as a protein kinase C substrate, but its biological function remains to be established. We demonstrated a specific interaction between 80K-H and TRPV5, co-localization of both proteins in the kidney, and similar transcriptional regulation by 1,25-dihydroxyvitamin D(3) and dietary Ca(2+). Furthermore, 80K-H directly bound Ca(2+), and inactivation of its two EF-hand structures totally abolished Ca(2+) binding. Electrophysiological studies using 80K-H mutants showed that three domains of 80K-H (the two EF-hand structures, the highly acidic glutamic stretch, and the His-Asp-Glu-Leu sequence) are critical determinants for TRPV5 activity. Importantly, inactivation of the EF-hand pair reduced the TRPV5-mediated Ca(2+) current and increased the TRPV5 sensitivity to intracellular Ca(2+), accelerating the feedback inhibition of the channel. None of the 80K-H mutants altered the TRPV5 plasma membrane localization nor the association of 80K-H with TRPV5, suggesting that 80K-H has a direct effect on TRPV5 activity. In conclusion, we report a novel function for 80K-H as a Ca(2+) sensor controlling TRPV5 channel activity.
Gkika
Dimitra
D
Department of Physiology, Nijmegen Centre for Molecular Life Sciences, University Medical Centre Nijmegen, NL-6500 HB Nijmegen, The Netherlands.
Mahieu
Frank
F
Nilius
Bernd
B
Hoenderop
Joost G J
JG
Bindels
René J M
RJ
D000042:Absorption; D000818:Animals; D019921:Biotinylation; D002117:Calcitriol / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004594:Electrophysiology; D005959:Glucosidases; D005982:Glutathione Transferase / Q000378:metabolism; D007150:Immunohistochemistry; D047908:Intracellular Signaling Peptides and Proteins; D007668:Kidney / Q000378:metabolism; D008565:Membrane Proteins; D051379:Mice; D008810:Mice, Inbred C57BL; D008954:Models, Biological; D009154:Mutation; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D020411:Oligonucleotide Array Sequence Analysis; D009865:Oocytes / Q000378:metabolism; D010750:Phosphoproteins / Q000502:physiology*; D011233:Precipitin Tests; D011493:Protein Kinase C / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D050916:TRPV Cation Channels; D014018:Tissue Distribution; D014981:Xenopus
D015220:Calcium Channels; D018076:DNA, Complementary; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D010750:Phosphoproteins; C512949:Prkcsh protein, mouse; D012333:RNA, Messenger; D050916:TRPV Cation Channels; C492240:Trpv5 protein, mouse; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D005982:Glutathione Transferase; D011493:Protein Kinase C; D005959:Glucosidases; D002117:Calcitriol; D002118:Calcium