HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

J. Biol. Chem., Vol. 283, Issue 4, 1911-1920, January 25, 2008

This Article Full Text Full Text (PDF) Supplemental Data All Versions of this Article: 283/4/1911    most recent M708213200v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Hamamoto, S. Articles by Uozumi, N. Search for Related Content PubMed PubMed Citation Articles by Hamamoto, S. Articles by Uozumi, N. Social Bookmarking                      What's this?

Characterization of a Tobacco TPK-type K⁺ Channel as a Novel Tonoplast K⁺ Channel Using Yeast Tonoplasts^*

From the ^aDepartment of Biomolecular Engineering, Graduate School of Engineering, Tohoku University, Aobayama 6-6-07, Sendai 980-8579, Japan, the ^bNew Industry Creation Hatchery Center, Tohoku University, Aobayama 6-6-10, Sendai 980-8579, Japan, the ^cLaboratory of Plant Nutrition, Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan, the ^dGraduate School of Pharmaceutical Sciences, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8675, Japan, the ^eDepartment of Molecular and Functional Genomics, Center for Integrated Research in Science, Shimane University, 1060 Nishikawatsu, Matsue 690-8504, Japan, the ^fDepartment of Genome Applied Microbiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University, Tsushima, Okayama 700-8530, Japan, the ^gDepartment of Synthetic Chemistry and Biological Chemistry, Graduate School of Engineering, Kyoto University, Katsura Campus, Kyoto 615-8510, Japan, the ^hGraduate School of Natural Science and Technology, Okayama University, 3-1-1 Tsushima-naka, Okayama 700-8530, Japan, the ⁱLaboratory of Cell Dynamics, Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya 464-8601, Japan, and the ^jInstitute of Molecular and Cellular Biosciences, The University of Tokyo, Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan

The tonoplast K⁺ membrane transport system plays a crucial role in maintaining K⁺ homeostasis in plant cells. Here, we isolated cDNAs encoding a two-pore K⁺ channel (NtTPK1) from Nicotiana tabacum cv. SR1 and cultured BY-2 tobacco cells. Two of the four variants of NtTPK1 contained VHG and GHG instead of the GYG signature sequence in the second pore region. All four products were functional when expressed in the Escherichia coli cell membrane, and NtTPK1 was targeted to the tonoplast in tobacco cells. Two of the three promoter sequences isolated from N. tabacum cv. SR1 were active, and expression from these was increased 2-fold by salt stress or high osmotic shock. To determine the properties of NtTPK1, we enlarged mutant yeast cells with inactivated endogenous tonoplast channels and prepared tonoplasts suitable for patch clamp recording allowing the NtTPK1-related channel conductance to be distinguished from the small endogenous currents. NtTPK1 exhibited strong selectivity for K⁺ over Na⁺. NtTPK1 activity was sensitive to spermidine and spermine, which were shown to be present in tobacco cells. NtTPK1 was active in the absence of Ca²⁺, but a cytosolic concentration of 45 µM Ca²⁺ resulted in a 2-fold increase in the amplitude of the K⁺ current. Acidification of the cytosol to pH 5.5 also markedly increased NtTPK1-mediated K⁺ currents. These results show that NtTPK1 is a novel tonoplast K⁺ channel belonging to a different group from the previously characterized vacuolar channels SV, FV, and VK.

Received for publication, October 3, 2007 , and in revised form, November 19, 2007.

^* This work was supported by grants-in-aid for scientific research (17078005 and 19380058) (to N. U.) from MEXT and JSPS. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains supplemental Figs. 1 and 2. The nucleotide sequence(s) reported in this paper has been submitted to the GenBank^TM/EBI Data Bank with accession number(s) EU161633 [GenBank] .

¹ To whom correspondence should be addressed. Fax: 81-22-795-7293; E-mail: uozumi{at}biophy.che.tohoku.ac.jp.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?