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Title: Cloning and expression of ligand-gated ion-channel receptor L2 in central nervous system

Abstract

An orphan receptor of ligand-gated ion-channel type (L2, also termed ZAC according to the presence of zinc ion for channel activation) was identified by computer-assisted search programs on human genome database. The L2 protein shares partial homology with serotonin receptors 5HT3A and 5HT3B. We have cloned L2 cDNA derived from human caudate nucleus and characterized the exon-intron structure as follows: (1) The L2 protein has four transmembrane regions (M1-M4) and a long cytoplasmic loop between M3 and M4. (2) The sequence is conserved in species including chimpanzee, dog, cow, and opossum. (3) Nine exons form its protein-coding region and especially exon 5 corresponds to a disulfide bond region on the amino-terminal side. Our analysis using multiple tissue cDNA panels revealed that at least two splicing variants of L2 mRNA are present. The cDNA PCR amplification study revealed that L2 mRNA is expressed in tissues including brain, pancreas, liver, lung, heart, kidney, and skeletal muscle while 5HT3A mRNA could be detected in brain, heart, placenta, lung, kidney, pancreas, and skeletal muscle, and 5HT3B mRNA in brain, kidney, and skeletal muscle, suggesting different significance in tissue expression of these receptors. Regional expression of L2 mRNA and protein was examined in brain. Themore » RT-PCR studies confirmed L2 mRNA expression in hippocampus, striatum, amygdala, and thalamus in adult brain. The L2 protein was immunolocalized by using antipeptide antibodies. Immunostained tissue sections revealed that L2-like immunoreactivity was dominantly expressed in the hippocampal CA3 pyramidal cells and in the polymorphic layer of the dentate gyrus. We analyzed the expression of L2 protein in HEK293 cells using GFP fusion protein reporter system. Western blots revealed that L2 protein confers sugar chains on the extracellular side. In transfected HEK293 cells, cellular membranes and intracellular puncta were densely labeled with GFP, suggesting selective dispatch to the final destination.« less

Authors:
 [1];  [1];  [1];  [1];  [1];  [1]
  1. Department of Anatomy and Brain Science, Kansai Medical University, Moriguchi, Osaka 570-8506 (Japan)
Publication Date:
OSTI Identifier:
20710971
Resource Type:
Journal Article
Journal Name:
Biochemical and Biophysical Research Communications
Additional Journal Information:
Journal Volume: 335; Journal Issue: 2; Other Information: DOI: 10.1016/j.bbrc.2005.07.079; PII: S0006-291X(05)01521-4; Copyright (c) 2005 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved; Country of input: International Atomic Energy Agency (IAEA); Journal ID: ISSN 0006-291X
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; CLONING; DOGS; HEART; HIPPOCAMPUS; KIDNEYS; LIGANDS; LIVER; LUNGS; MARSUPIALS; MUSCLES; NERVE CELLS; PANCREAS; PLACENTA; POLYMERASE CHAIN REACTION; RECEPTORS; SACCHAROSE; SEROTONIN; SPLICING; THALAMUS; ZINC IONS

Citation Formats

Houtani, Takeshi, Munemoto, Yumi, Kase, Masahiko, Sakuma, Satoru, Tsutsumi, Toshiyuki, and Sugimoto, Tetsuo. Cloning and expression of ligand-gated ion-channel receptor L2 in central nervous system. United States: N. p., 2005. Web. doi:10.1016/j.bbrc.2005.07.079.
Houtani, Takeshi, Munemoto, Yumi, Kase, Masahiko, Sakuma, Satoru, Tsutsumi, Toshiyuki, & Sugimoto, Tetsuo. Cloning and expression of ligand-gated ion-channel receptor L2 in central nervous system. United States. https://doi.org/10.1016/j.bbrc.2005.07.079
Houtani, Takeshi, Munemoto, Yumi, Kase, Masahiko, Sakuma, Satoru, Tsutsumi, Toshiyuki, and Sugimoto, Tetsuo. 2005. "Cloning and expression of ligand-gated ion-channel receptor L2 in central nervous system". United States. https://doi.org/10.1016/j.bbrc.2005.07.079.
@article{osti_20710971,
title = {Cloning and expression of ligand-gated ion-channel receptor L2 in central nervous system},
author = {Houtani, Takeshi and Munemoto, Yumi and Kase, Masahiko and Sakuma, Satoru and Tsutsumi, Toshiyuki and Sugimoto, Tetsuo},
abstractNote = {An orphan receptor of ligand-gated ion-channel type (L2, also termed ZAC according to the presence of zinc ion for channel activation) was identified by computer-assisted search programs on human genome database. The L2 protein shares partial homology with serotonin receptors 5HT3A and 5HT3B. We have cloned L2 cDNA derived from human caudate nucleus and characterized the exon-intron structure as follows: (1) The L2 protein has four transmembrane regions (M1-M4) and a long cytoplasmic loop between M3 and M4. (2) The sequence is conserved in species including chimpanzee, dog, cow, and opossum. (3) Nine exons form its protein-coding region and especially exon 5 corresponds to a disulfide bond region on the amino-terminal side. Our analysis using multiple tissue cDNA panels revealed that at least two splicing variants of L2 mRNA are present. The cDNA PCR amplification study revealed that L2 mRNA is expressed in tissues including brain, pancreas, liver, lung, heart, kidney, and skeletal muscle while 5HT3A mRNA could be detected in brain, heart, placenta, lung, kidney, pancreas, and skeletal muscle, and 5HT3B mRNA in brain, kidney, and skeletal muscle, suggesting different significance in tissue expression of these receptors. Regional expression of L2 mRNA and protein was examined in brain. The RT-PCR studies confirmed L2 mRNA expression in hippocampus, striatum, amygdala, and thalamus in adult brain. The L2 protein was immunolocalized by using antipeptide antibodies. Immunostained tissue sections revealed that L2-like immunoreactivity was dominantly expressed in the hippocampal CA3 pyramidal cells and in the polymorphic layer of the dentate gyrus. We analyzed the expression of L2 protein in HEK293 cells using GFP fusion protein reporter system. Western blots revealed that L2 protein confers sugar chains on the extracellular side. In transfected HEK293 cells, cellular membranes and intracellular puncta were densely labeled with GFP, suggesting selective dispatch to the final destination.},
doi = {10.1016/j.bbrc.2005.07.079},
url = {https://www.osti.gov/biblio/20710971}, journal = {Biochemical and Biophysical Research Communications},
issn = {0006-291X},
number = 2,
volume = 335,
place = {United States},
year = {Fri Sep 23 00:00:00 EDT 2005},
month = {Fri Sep 23 00:00:00 EDT 2005}
}