Claudins form a large family of TJ (tight junction) proteins featuring four transmembrane segments (TM1–TM4), two extracellular loops, one intracellular loop and intracellular N- and C-termini. They form continuous and branched TJ strands by homo- or heterophilic interaction within the same membrane (cis-interaction) and with claudins of the opposing lateral cell membrane (trans-interaction). In order to clarify the molecular organization of TJ strand formation, we investigated the cis-interaction of two abundant prototypic claudins. Human claudin-1 and claudin-3, fused to ECFP or EYFP at the N- or C-terminus, were expressed in the TJ-free cell line HEK (human embryonic kidney)-293. Using FRET analysis, the proximity of claudin N- and C-termini integrated in homopolymeric strands composed of claudin-3 or of heteropolymeric strands composed of claudin-1 and claudin-3 were determined. The main results are that (i) within homo- and heteropolymers, the average distance between the cytoplasmic ends of the TM1s of cis-interacting claudin molecules is shorter than the average distance between their TM4s, and (ii) TM1 segments of neighbouring claudins are oriented towards each other as the cytoplasmic end of TM1 is in close proximity to more other TM1 segments than TM4 is to other TM4 segments. The results indicate at least two different cis-interaction interfaces within claudin-3 homopolymers as well as within claudin-1/claudin-3 heteropolymers. The data provide novel insight into the molecular TJ architecture consistent with a model with an antiparallel double-row cis-arrangement of classic claudin protomers within strands.
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Research Article|
June 15 2015
Probing the cis-arrangement of prototype tight junction proteins claudin-1 and claudin-3
Susanne Milatz;
Susanne Milatz
*Institute of Clinical Physiology, Campus Benjamin Franklin, Charité–Universitätsmedizin Berlin, 12200 Berlin, Germany
†Institute of Physiology, Christian-Albrechts-University Kiel, Hermann-Rodewald-Straße 5, 24118 Kiel, Germany
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Jörg Piontek;
Jörg Piontek
*Institute of Clinical Physiology, Campus Benjamin Franklin, Charité–Universitätsmedizin Berlin, 12200 Berlin, Germany
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Jörg-Dieter Schulzke;
Jörg-Dieter Schulzke
*Institute of Clinical Physiology, Campus Benjamin Franklin, Charité–Universitätsmedizin Berlin, 12200 Berlin, Germany
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Ingolf E. Blasig;
Ingolf E. Blasig
‡Leibniz-Institut für Molekulare Pharmakologie, Robert-Rössle-Straße 10, 13125 Berlin, Germany
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Michael Fromm;
Michael Fromm
*Institute of Clinical Physiology, Campus Benjamin Franklin, Charité–Universitätsmedizin Berlin, 12200 Berlin, Germany
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Dorothee Günzel
Dorothee Günzel
1
*Institute of Clinical Physiology, Campus Benjamin Franklin, Charité–Universitätsmedizin Berlin, 12200 Berlin, Germany
1To whom correspondence should be addressed (email dorothee.guenzel@charite.de).
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Publisher: Portland Press Ltd
Received:
February 04 2015
Revision Received:
April 02 2015
Accepted:
April 07 2015
Accepted Manuscript online:
April 07 2015
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2015 Biochemical Society
2015
Biochem J (2015) 468 (3): 449–458.
Article history
Received:
February 04 2015
Revision Received:
April 02 2015
Accepted:
April 07 2015
Accepted Manuscript online:
April 07 2015
Citation
Susanne Milatz, Jörg Piontek, Jörg-Dieter Schulzke, Ingolf E. Blasig, Michael Fromm, Dorothee Günzel; Probing the cis-arrangement of prototype tight junction proteins claudin-1 and claudin-3. Biochem J 15 June 2015; 468 (3): 449–458. doi: https://doi.org/10.1042/BJ20150148
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