Human Molecular Genetics Advance Access originally published online on October 11, 2006
Human Molecular Genetics 2006 15(23):3379-3386; doi:10.1093/hmg/ddl414
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Fibulin-5 mutations: mechanisms of impaired elastic fiber formation in recessive cutis laxa
1 Department of Pediatrics, 2 Department of Cell Biology and Physiology and 3 Department of Genetics, Washington University School of Medicine, St Louis, MO 63110, USA, 4 Center for Medical Genetics, Ghent University, Ghent 9000, Belgium and 5 Department of Anatomy and Cell Biology, McGill University, Montreal, Canada H3A 2B2
* To whom correspondence should be addressed at: Department of Pediatrics and Department of Genetics, Washington University School of Medicine, 660 South Euclid Avenue, Campus Box 8208, St Louis, MO 63110, USA. Tel: +1 3142862973; Fax: +1 3142862893; Email: urban_z{at}kids.wustl.edu
Received August 26, 2006; Accepted October 9, 2006
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.
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