The RGD motif in fibronectin is essential for development but dispensable for fibril assembly.
Fibronectin (FN) is secreted as a disulfide-bonded FN dimer. Each subunit contains three types of repeating modules: FN-I, FN-II, and FN-III. The interactions of alpha5beta1 or alphav integrins with the RGD motif of FN-III repeat 10 (FN-III10) are considered an essential step in the assembly of FN fibrils. To test this hypothesis in vivo, we replaced the RGD motif with the inactive RGE in mice. FN-RGE homozygous embryos die at embryonic day 10 with shortened posterior trunk, absent tail bud-derived somites, and severe vascular defects resembling the phenotype of alpha5 integrin-deficient mice. Surprisingly, the absence of a functional RGD motif in FN did not compromise assembly of an FN matrix in mutant embryos or on mutant cells. Matrix assembly assays and solid-phase binding assays reveal that alphavbeta3 integrin assembles FN-RGE by binding an isoDGR motif in FN-I5, which is generated by the nonenzymatic rearrangement of asparagines (N) into an iso-aspartate (iso-D). Our findings demonstrate that FN contains a novel motif for integrin binding and fibril formation whose activity is controlled by amino acid modification.
Takahashi
Seiichiro
S
Max Planck Institute of Biochemistry, Department of Molecular Medicine, 82152 Martinsried, Germany.
Leiss
Michael
M
Moser
Markus
M
Ohashi
Tomoo
T
Kitao
Tomoe
T
Heckmann
Dominik
D
Pfeifer
Alexander
A
Kessler
Horst
H
Takagi
Junichi
J
Erickson
Harold P
HP
Fässler
Reinhard
R
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001224:Aspartic Acid / Q000378:metabolism*; D001665:Binding Sites; D002461:Cell Line, Transformed; D019281:Dimerization; D004220:Disulfides / Q000737:chemistry; D004622:Embryo, Mammalian; D005109:Extracellular Matrix / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D005353:Fibronectins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006579:Heterozygote; D039302:Integrin alphaVbeta3 / Q000235:genetics / Q000378:metabolism; D051379:Mice; D009842:Oligopeptides / Q000737:chemistry*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000378:metabolism; D012155:Reticulin / Q000378:metabolism*; D012995:Solubility
D004220:Disulfides; D005353:Fibronectins; D039302:Integrin alphaVbeta3; D009842:Oligopeptides; D011994:Recombinant Proteins; D012155:Reticulin; C112658:arginyl-glycyl-glutamic acid; D001224:Aspartic Acid; C047981:arginyl-glycyl-aspartic acid