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Chloride channel ClC‐5 linked to Dent's disease decreases protein level of the intestinal calcium entry channel TRPV6 in Xenopus oocytes
Author(s) -
Na Tao,
Zhang Wei,
Peng JiBin
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.998.15
Subject(s) - trpv6 , chemistry , xenopus , nitrendipine , calcium , calcium channel , biophysics , voltage dependent calcium channel , endocrinology , medicine , biochemistry , biology , gene , organic chemistry
TRPV6, a member of the transient receptor potential superfamily of ion channels, mediates transcellular Ca 2+ absorption in the intestine. Diet‐dependent hypercalciuria in patients with Dent's disease and in ClC‐5 knock‐down mice promoted us to investigate whether ClC‐5 modulates TRPV6‐mediated Ca 2+ transport. When ClC‐5 was expressed in Xenopus laevis oocytes for 2 days, it induced a Gd 3+ ‐sensitive Ca 2+ uptake, which was insensitive to L‐type voltage‐gated Ca 2+ channel blocker verapamil and nitrendipine. Thus, TRPV6‐mediated Na + current was used as a measurement of TRPV6 activity instead of radiotracer Ca 2+ influx assay. When ClC‐5 was co‐expressed with TRPV6 in oocytes, TRPV6‐induced Na + current was almost completely suppressed using voltage clamp. Consistent with these results, TRPV6 protein level was greatly diminished by co‐expression with ClC‐5. The decrease in TRPV6 protein level by ClC‐5 was dose‐dependent and correlated with the decrease in TRPV6 Na + current. To locate the key fragment in ClC‐5 accounting for TRPV6 regulation, amino‐terminal region (amino acids 1‐53), carboxy‐terminal region (amino acids 546‐746) or both of them in ClC‐5 were truncated. Although the three ClC‐5 constructs didn't exhibit outward Cl − current as the wild‐type ClC‐5 did, they retained the ability to inhibit TRPV6‐induced Na + current by decreasing TRPV6 protein level. Co‐expression of ClC‐5 with TRPV6 in mouse duodenum and in human intestinal cell line Caco‐2 was confirmed by RT‐PCR. Therefore, ClC‐5 may play an important role in regulating TRPV6 protein level, and in turn in modulating intestinal Ca 2+ absorption. Further studies are being directed toward the mechanisms by which TRPV6 protein is decreased by ClC‐5.