Different domains in nidogen-1 and nidogen-2 drive basement membrane formation in skin organotypic cocultures

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Different domains in nidogen-1 and nidogen-2 drive basement membrane formation in skin organotypic cocultures. / Bechtel, Manuela; Keller, Manuel V; Bloch, Wilhelm; Sasaki, Takako; Boukamp, Petra; Zaucke, Frank; Paulsson, Mats; Nischt, Roswitha.

in: FASEB journal : official publication of the Federation of American Societies for Experimental Biology, Jahrgang 26, Nr. 9, 01.09.2012, S. 3637-3648.

Publikationen: Beitrag in FachzeitschriftZeitschriftenaufsätzeForschung

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@article{54c010d3f8a147b7bab25513621b77fc,
title = "Different domains in nidogen-1 and nidogen-2 drive basement membrane formation in skin organotypic cocultures",
abstract = "Nidogen-1 and nidogen-2 are homologous proteins found in all basement membranes (BMs). They show comparable binding activities in vitro and partially redundant functions in vivo. Previously, we showed that in skin organotypic cocultures, BM formation was prevented in the absence of nidogens and that either nidogen was able to rescue this failure. We now dissected the two nidogens to identify the domains required for BM deposition. For that purpose, HaCaT cells were grown on collagen matrices containing nidogen-deficient, murine fibroblasts. After addition of nidogen-1 or nidogen-2 protein fragments comprising different binding domains, BM deposition was analyzed by immunofluorescence and electron microscopy. We could demonstrate that the rod-G3 domain of nidogen-2 was sufficient to achieve deposition of BM components at the epidermal-collagen interface. In contrast, for nidogen-1, both the G2 and G3 domains were required. Immunoblot analysis confirmed that all BM components were present in comparable amounts under all culture conditions. This finding demonstrates that nidogens, although homologous proteins, exert their effect on BM assembly through different binding domains, which may in turn result in alterations of BM structure and functions, thus providing an explanation for the phenotypical differences observed between nidogen-1 and -2 deficient mice.",
keywords = "Base Sequence, Basement Membrane, Binding Sites, Cell Line, Coculture Techniques, Culture Media, Conditioned, DNA Primers, Fluorescent Antibody Technique, Indirect, Laminin, Membrane Glycoproteins, Microscopy, Electron, Transmission, Polymerase Chain Reaction, Skin",
author = "Manuela Bechtel and Keller, {Manuel V} and Wilhelm Bloch and Takako Sasaki and Petra Boukamp and Frank Zaucke and Mats Paulsson and Roswitha Nischt",
year = "2012",
month = "9",
day = "1",
doi = "10.1096/fj.11-194597",
language = "English",
volume = "26",
pages = "3637--3648",
journal = "FASEB journal : official publication of the Federation of American Societies for Experimental Biology",
issn = "1530-6860",
publisher = "FASEB",
number = "9",

}

RIS

TY - JOUR

T1 - Different domains in nidogen-1 and nidogen-2 drive basement membrane formation in skin organotypic cocultures

AU - Bechtel, Manuela

AU - Keller, Manuel V

AU - Bloch, Wilhelm

AU - Sasaki, Takako

AU - Boukamp, Petra

AU - Zaucke, Frank

AU - Paulsson, Mats

AU - Nischt, Roswitha

PY - 2012/9/1

Y1 - 2012/9/1

N2 - Nidogen-1 and nidogen-2 are homologous proteins found in all basement membranes (BMs). They show comparable binding activities in vitro and partially redundant functions in vivo. Previously, we showed that in skin organotypic cocultures, BM formation was prevented in the absence of nidogens and that either nidogen was able to rescue this failure. We now dissected the two nidogens to identify the domains required for BM deposition. For that purpose, HaCaT cells were grown on collagen matrices containing nidogen-deficient, murine fibroblasts. After addition of nidogen-1 or nidogen-2 protein fragments comprising different binding domains, BM deposition was analyzed by immunofluorescence and electron microscopy. We could demonstrate that the rod-G3 domain of nidogen-2 was sufficient to achieve deposition of BM components at the epidermal-collagen interface. In contrast, for nidogen-1, both the G2 and G3 domains were required. Immunoblot analysis confirmed that all BM components were present in comparable amounts under all culture conditions. This finding demonstrates that nidogens, although homologous proteins, exert their effect on BM assembly through different binding domains, which may in turn result in alterations of BM structure and functions, thus providing an explanation for the phenotypical differences observed between nidogen-1 and -2 deficient mice.

AB - Nidogen-1 and nidogen-2 are homologous proteins found in all basement membranes (BMs). They show comparable binding activities in vitro and partially redundant functions in vivo. Previously, we showed that in skin organotypic cocultures, BM formation was prevented in the absence of nidogens and that either nidogen was able to rescue this failure. We now dissected the two nidogens to identify the domains required for BM deposition. For that purpose, HaCaT cells were grown on collagen matrices containing nidogen-deficient, murine fibroblasts. After addition of nidogen-1 or nidogen-2 protein fragments comprising different binding domains, BM deposition was analyzed by immunofluorescence and electron microscopy. We could demonstrate that the rod-G3 domain of nidogen-2 was sufficient to achieve deposition of BM components at the epidermal-collagen interface. In contrast, for nidogen-1, both the G2 and G3 domains were required. Immunoblot analysis confirmed that all BM components were present in comparable amounts under all culture conditions. This finding demonstrates that nidogens, although homologous proteins, exert their effect on BM assembly through different binding domains, which may in turn result in alterations of BM structure and functions, thus providing an explanation for the phenotypical differences observed between nidogen-1 and -2 deficient mice.

KW - Base Sequence

KW - Basement Membrane

KW - Binding Sites

KW - Cell Line

KW - Coculture Techniques

KW - Culture Media, Conditioned

KW - DNA Primers

KW - Fluorescent Antibody Technique, Indirect

KW - Laminin

KW - Membrane Glycoproteins

KW - Microscopy, Electron, Transmission

KW - Polymerase Chain Reaction

KW - Skin

U2 - 10.1096/fj.11-194597

DO - 10.1096/fj.11-194597

M3 - Journal articles

VL - 26

SP - 3637

EP - 3648

JO - FASEB journal : official publication of the Federation of American Societies for Experimental Biology

T2 - FASEB journal : official publication of the Federation of American Societies for Experimental Biology

JF - FASEB journal : official publication of the Federation of American Societies for Experimental Biology

SN - 1530-6860

SN - 0892-6638

IS - 9

ER -

ID: 66957