title: Fibulin-5 mutations: mechanisms of impaired elastic fiber formation in recessive cutis laxa.
abstract: To elucidate the molecular mechanisms of impaired elastic fiber formation in recessive cutis laxa, we have investigated two disease-causing missense substitutions in fibulin-5, C217R and S227P. Pulse-chase immunoprecipitation experiments indicated that S227P mutant fibulin-5 was synthesized and secreted by skin fibroblasts at a reduced rate when compared with the wild-type protein. Both mutants failed to be incorporated into elastic fibers by transfected rat lung fibroblasts. Purified recombinant fibulin-5 with either mutation showed reduced affinity for tropoelastin in solid-phase binding assays. Furthermore, S227P mutant fibulin-5 also showed impaired association with fibrillin-1 microfibrils. The same mutation triggered an endoplasmic reticulum (ER) stress response, as indicated by the strong co-localization of this mutant protein with folding chaperones in the ER, including calreticulin, immunoglobulin-binding protein and protein disulfide isomerase, and by increased rates of apoptosis in patient fibroblasts. Histological analysis of skin sections from a cutis laxa patient with a homozygous S227P mutation showed a lack of fibulin-5 in the extracellular matrix and a concomitant disorganization of dermal elastic fibers. By electron microscopy, elastic fibers in the skin of this patient showed a failure of elastin globules to fuse into a continuous elastic fiber core. We conclude that recessive cutis laxa mutations in fibulin-5 result in misfolding, decreased secretion and a reduced interaction with elastin and fibrillin-1 leading to impaired elastic fiber development. These findings support the hypothesis that fibulin-5 is necessary for elastic fiber formation by facilitating the deposition of elastin onto a microfibrillar scaffold via direct molecular interactions.
authors.affiliation: Department of Pediatrics, Washington University School of Medicine, St Louis, MO 63110, USA.
authors.forename: Qirui
authors.initials: Q
authors.lastname: Hu
authors.forename: Bart L
authors.initials: BL
authors.lastname: Loeys
authors.forename: Paul J
authors.initials: PJ
authors.lastname: Coucke
authors.forename: Anne
authors.initials: A
authors.lastname: De Paepe
authors.forename: Robert P
authors.initials: RP
authors.lastname: Mecham
authors.forename: Jiwon
authors.initials: J
authors.lastname: Choi
authors.forename: Elaine C
authors.initials: EC
authors.lastname: Davis
authors.forename: Zsolt
authors.initials: Z
authors.lastname: Urban
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D037282:Calreticulin / Q000032:analysis / Q000378:metabolism; D003483:Cutis Laxa / Q000235:genetics* / Q000378:metabolism / Q000473:pathology*; D004547:Elastic Tissue / Q000254:growth & development / Q000378:metabolism* / Q000648:ultrastructure*; D004721:Endoplasmic Reticulum / Q000737:chemistry / Q000378:metabolism; D016326:Extracellular Matrix Proteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D000071838:Fibrillin-1; D000071837:Fibrillins; D005347:Fibroblasts / Q000378:metabolism / Q000473:pathology; D005808:Genes, Recessive; D006801:Humans; D047468:Immunoprecipitation; D008840:Microfilament Proteins / Q000032:analysis / Q000378:metabolism; D020125:Mutation, Missense; D029584:Prostatic Secretory Proteins / Q000378:metabolism; D019704:Protein Disulfide-Isomerases / Q000378:metabolism; D017510:Protein Folding; D051381:Rats; D012867:Skin / Q000378:metabolism / Q000473:pathology*; D014333:Tropoelastin / Q000378:metabolism; D054392:Adipokines
chemical_list: D037282:Calreticulin; D016326:Extracellular Matrix Proteins; D000071838:Fibrillin-1; D000071837:Fibrillins; D008840:Microfilament Proteins; D029584:Prostatic Secretory Proteins; D019704:Protein Disulfide-Isomerases; D014333:Tropoelastin; C492373:FBLN5 protein, human; C000605722:FBN1 protein, human; D054392:Adipokines; C000605725:Fbn1 protein, rat; C043076:beta-microseminoprotein