We examined the function of GFP-IP3R3 (green fluorescent protein-tagged inositol 1,4,5-trisphosphate receptor type 3) in Ca2+ release and entry using a mutant DT40 cell line (IP3R-KO) in which all three IP3R genes had been disrupted. GFP-IP3R3 fluorescence largely overlapped with the distribution of endoplasmic reticulum, whereas a portion of GFP-IP3R3 apparently co-localized with the plasma membrane. The application of IP3 to permeabilized WT (wild-type) DT40 cells induced Ca2+ release from internal stores. Although this did not occur in IP3R-KO cells it was restored by expression of GFP-IP3R3. In intact cells, application of anti-IgM, an activator of the BCR (B-cell receptor), or trypsin, a protease-activated receptor 2 agonist, did not cause any Ca2+ response in IP3R-KO cells, whereas these treatments induced oscillatory or transient Ca2+ responses in GFP-IP3R3-expressing IP3R-KO cells, as well as in WT cells. In addition, BCR activation elicited Ca2+ entry in WT and GFP-IP3R3-expressing IP3R-KO cells but not in IP3R-KO cells. This BCR-mediated Ca2+ entry was observed in the presence of La3+, which blocks capacitative Ca2+ entry. Thapsigargin depleted Ca2+ stores and led to Ca2+ entry in IP3R-KO cells irrespective of GFP-IP3R3 expression. In contrast with BCR stimulation, thapsigargin-induced Ca2+ entry was completely blocked by La3+, suggesting that the BCR-mediated Ca2+ entry pathway is distinct from the capacitative Ca2+ entry pathway. The present study demonstrates that GFP-IP3R3 could compensate for native IP3R in both IP3-induced Ca2+ release and BCR-mediated Ca2+ entry.
Skip Nav Destination
Article navigation
September 2004
-
Cover Image
Cover Image
- PDF Icon PDF LinkFront Matter
- PDF Icon PDF LinkTable of Contents
- PDF Icon PDF LinkEditorial Board
Research Article|
September 07 2004
Functional analysis of the green fluorescent protein-tagged inositol 1,4,5-trisphosphate receptor type 3 in Ca2+ release and entry in DT40 B lymphocytes
Takao MORITA;
Takao MORITA
*Department of Dental Pharmacology, School of Dentistry, Health Sciences University of Hokkaido, Ishikari-Tobetsu, Hokkaido 061-0293, Japan
Search for other works by this author on:
Akihiko TANIMURA;
Akihiko TANIMURA
1
*Department of Dental Pharmacology, School of Dentistry, Health Sciences University of Hokkaido, Ishikari-Tobetsu, Hokkaido 061-0293, Japan
1To whom correspondence should be addressed (email tanimura@hoku-iryo-u.ac.jp).
Search for other works by this author on:
Akihiro NEZU;
Akihiro NEZU
*Department of Dental Pharmacology, School of Dentistry, Health Sciences University of Hokkaido, Ishikari-Tobetsu, Hokkaido 061-0293, Japan
Search for other works by this author on:
Tomohiro KUROSAKI;
Tomohiro KUROSAKI
†Department of Molecular Genetics, Institute for Liver Research, Kansai Medical University, Moriguchi 570-8506, Japan
‡Laboratory for Lymphocyte Differentiation, RIKEN Research Center for Allergy and Immunology, Turumi-ku, Yokohama 230-0045, Japan
Search for other works by this author on:
Yosuke TOJYO
Yosuke TOJYO
*Department of Dental Pharmacology, School of Dentistry, Health Sciences University of Hokkaido, Ishikari-Tobetsu, Hokkaido 061-0293, Japan
Search for other works by this author on:
Publisher: Portland Press Ltd
Received:
December 23 2003
Revision Received:
May 14 2004
Accepted:
June 03 2004
Accepted Manuscript online:
June 03 2004
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London
2004
Biochem J (2004) 382 (3): 793–801.
Article history
Received:
December 23 2003
Revision Received:
May 14 2004
Accepted:
June 03 2004
Accepted Manuscript online:
June 03 2004
Citation
Takao MORITA, Akihiko TANIMURA, Akihiro NEZU, Tomohiro KUROSAKI, Yosuke TOJYO; Functional analysis of the green fluorescent protein-tagged inositol 1,4,5-trisphosphate receptor type 3 in Ca2+ release and entry in DT40 B lymphocytes. Biochem J 15 September 2004; 382 (3): 793–801. doi: https://doi.org/10.1042/BJ20031970
Download citation file:
Sign in
Don't already have an account? Register
Sign in to your personal account
You could not be signed in. Please check your email address / username and password and try again.
Could not validate captcha. Please try again.