Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
Proc Natl Acad Sci U S A
2001 May 22;9811:6488-93. doi: 10.1073/pnas.101556598.
Show Gene links
Show Anatomy links
Evidence in support of a four transmembrane-pore-transmembrane topology model for the Arabidopsis thaliana Na+/K+ translocating AtHKT1 protein, a member of the superfamily of K+ transporters.
Kato Y
,
Sakaguchi M
,
Mori Y
,
Saito K
,
Nakamura T
,
Bakker EP
,
Sato Y
,
Goshima S
,
Uozumi N
.
???displayArticle.abstract???
The Arabidopsis thaliana AtHKT1 protein, a Na(+)/K(+) transporter, is capable of mediating inward Na(+) currents in Xenopus laevis oocytes and K(+) uptake in Escherichia coli. HKT1 proteins are members of a superfamily of K(+) transporters. These proteins have been proposed to contain eight transmembrane segments and four pore-forming regions arranged in a mode similar to that of a K(+) channel tetramer. However, computer analysis of the AtHKT1 sequence identified eleven potential transmembrane segments. We have investigated the membrane topology of AtHKT1 with three different techniques. First, a gene fusion alkaline phosphatase study in E. coli clearly defined the topology of the N-terminal and middle region of AtHKT1, but the model for membrane folding of the C-terminal region had to be refined. Second, with a reticulocyte-lysate supplemented with dog-pancreas microsomes, we demonstrated that N-glycosylation occurs at position 429 of AtHKT1. An engineered unglycosylated protein variant, N429Q, mediated Na(+) currents in X. laevis oocytes with the same characteristics as the wild-type protein, indicating that N-glycosylation is not essential for the functional expression and membrane targeting of AtHKT1. Five potential glycosylation sites were introduced into the N429Q. Their pattern of glycosylation supported the model based on the E. coli-alkaline phosphatase data. Third, immunocytochemical experiments with FLAG-tagged AtHKT1 in HEK293 cells revealed that the N and C termini of AtHKT1, and the regions containing residues 135-142 and 377-384, face the cytosol, whereas the region of residues 55-62 is exposed to the outside. Taken together, our results show that AtHKT1 contains eight transmembrane-spanning segments.
Akiyama,
Export of Escherichia coli alkaline phosphatase attached to an integral membrane protein, SecY.
1989, Pubmed
Akiyama,
Export of Escherichia coli alkaline phosphatase attached to an integral membrane protein, SecY.
1989,
Pubmed
Bibi,
Functional expression of mouse mdr1 in Escherichia coli.
1993,
Pubmed
Blount,
Membrane topology and multimeric structure of a mechanosensitive channel protein of Escherichia coli.
1996,
Pubmed
Buurman,
Genetic evidence for two sequentially occupied K+ binding sites in the Kdp transport ATPase.
1995,
Pubmed
Derman,
Escherichia coli alkaline phosphatase fails to acquire disulfide bonds when retained in the cytoplasm.
1991,
Pubmed
Diatloff,
Site directed mutagenesis reduces the Na+ affinity of HKT1, an Na+ energized high affinity K+ transporter.
1998,
Pubmed
Doyle,
The structure of the potassium channel: molecular basis of K+ conduction and selectivity.
1998,
Pubmed
Durell,
Evolutionary relationship between K(+) channels and symporters.
1999,
Pubmed
Durell,
Structural models of the KtrB, TrkH, and Trk1,2 symporters based on the structure of the KcsA K(+) channel.
1999,
Pubmed
Durell,
Does the KdpA subunit from the high affinity K(+)-translocating P-type KDP-ATPase have a structure similar to that of K(+) channels?
2000,
Pubmed
Enomoto,
Topological study of Vibrio alginolyticus NhaB Na+/H+ antiporter using gene fusions in Escherichia coli cells.
1998,
Pubmed
Gaber,
TRK1 encodes a plasma membrane protein required for high-affinity potassium transport in Saccharomyces cerevisiae.
1988,
Pubmed
Gassman,
Alkali cation selectivity of the wheat root high-affinity potassium transporter HKT1.
1996,
Pubmed
,
Xenbase
Goder,
Glycosylation can influence topogenesis of membrane proteins and reveals dynamic reorientation of nascent polypeptides within the translocon.
1999,
Pubmed
Henn,
Probing the transmembrane topology of cyclic nucleotide-gated ion channels with a gene fusion approach.
1995,
Pubmed
Jan,
Potassium channels and their evolving gates.
1994,
Pubmed
Kim,
AtKUP1: an Arabidopsis gene encoding high-affinity potassium transport activity.
1998,
Pubmed
,
Xenbase
Ko,
TRK1 and TRK2 encode structurally related K+ transporters in Saccharomyces cerevisiae.
1991,
Pubmed
Koizumi,
Overexpression of a gene that encodes the first enzyme in the biosynthesis of asparagine-linked glycans makes plants resistant to tunicamycin and obviates the tunicamycin-induced unfolded protein response.
1999,
Pubmed
Kubo,
Primary structure and functional expression of a mouse inward rectifier potassium channel.
1993,
Pubmed
,
Xenbase
Kyte,
A simple method for displaying the hydropathic character of a protein.
1982,
Pubmed
Lerouge,
N-glycoprotein biosynthesis in plants: recent developments and future trends.
1998,
Pubmed
Liu,
Partial deletion of a loop region in the high affinity K+ transporter HKT1 changes ionic permeability leading to increased salt tolerance.
2000,
Pubmed
,
Xenbase
Manoil,
A genetic approach to analyzing membrane protein topology.
1986,
Pubmed
Nakamura,
KtrAB, a new type of bacterial K(+)-uptake system from Vibrio alginolyticus.
1998,
Pubmed
Nilsson,
Proline-induced disruption of a transmembrane alpha-helix in its natural environment.
1998,
Pubmed
Ota,
Assessment of topogenic functions of anticipated transmembrane segments of human band 3.
1998,
Pubmed
Ota,
Forced transmembrane orientation of hydrophilic polypeptide segments in multispanning membrane proteins.
1998,
Pubmed
Popov,
Mapping the ends of transmembrane segments in a polytopic membrane protein. Scanning N-glycosylation mutagenesis of extracytosolic loops in the anion exchanger, band 3.
1997,
Pubmed
Rubio,
Sodium-driven potassium uptake by the plant potassium transporter HKT1 and mutations conferring salt tolerance.
1995,
Pubmed
,
Xenbase
Rubio,
Genetic selection of mutations in the high affinity K+ transporter HKT1 that define functions of a loop site for reduced Na+ permeability and increased Na+ tolerance.
1999,
Pubmed
,
Xenbase
Sakaguchi,
Functions of signal and signal-anchor sequences are determined by the balance between the hydrophobic segment and the N-terminal charge.
1992,
Pubmed
Sato,
The amino-terminal structures that determine topological orientation of cytochrome P-450 in microsomal membrane.
1990,
Pubmed
Schachtman,
Structure and transport mechanism of a high-affinity potassium uptake transporter from higher plants.
1994,
Pubmed
,
Xenbase
Schachtman,
Expression of an inward-rectifying potassium channel by the Arabidopsis KAT1 cDNA.
1992,
Pubmed
,
Xenbase
Schlösser,
TrkH and its homolog, TrkG, determine the specificity and kinetics of cation transport by the Trk system of Escherichia coli.
1995,
Pubmed
Takase,
Sequencing and characterization of the ntp gene cluster for vacuolar-type Na(+)-translocating ATPase of Enterococcus hirae.
1994,
Pubmed
Tholema,
Change to alanine of one out of four selectivity filter glycines in KtrB causes a two orders of magnitude decrease in the affinities for both K+ and Na+ of the Na+ dependent K+ uptake system KtrAB from Vibrio alginolyticus.
1999,
Pubmed
Tjaden,
Expression of a plastidic ATP/ADP transporter gene in Escherichia coli leads to a functional adenine nucleotide transport system in the bacterial cytoplasmic membrane.
1998,
Pubmed
Uozumi,
Identification of strong modifications in cation selectivity in an Arabidopsis inward rectifying potassium channel by mutant selection in yeast.
1995,
Pubmed
,
Xenbase
Uozumi,
Determination of transmembrane topology of an inward-rectifying potassium channel from Arabidopsis thaliana based on functional expression in Escherichia coli.
1998,
Pubmed
Uozumi,
The Arabidopsis HKT1 gene homolog mediates inward Na(+) currents in xenopus laevis oocytes and Na(+) uptake in Saccharomyces cerevisiae.
2000,
Pubmed
,
Xenbase
von Heijne,
Membrane protein structure prediction. Hydrophobicity analysis and the positive-inside rule.
1992,
Pubmed
von Heijne,
Mitochondrial targeting sequences may form amphiphilic helices.
1986,
Pubmed