Microfibril-associated MAGP-2 stimulates elastic fiber assembly.
Elastic fibers are complex structures composed of a tropoelastin inner core and microfibril outer mantle guiding tropoelastin deposition. Microfibrillar proteins mainly include fibrillins and microfibril-associated glycoproteins (MAGPs). MAGP-2 exhibits developmental expression peaking at elastic fiber onset, suggesting that MAGP-2 mediates elastic fiber assembly. To determine whether MAGP-2 regulates elastic fiber assembly, we used an in vitro model featuring doxycycline-regulated cells conditionally overexpressing exogenous MAGP-2 and constitutively expressing enhanced green fluorescent protein-tagged tropoelastin. Analysis by immunofluorescent staining showed that MAGP-2 overexpression dramatically increased elastic fibers levels, independently of extracellular levels of soluble tropoelastin, indicating that MAGP-2 stimulates elastic fiber assembly. This was associated with increased levels of matrix-associated MAGP-2. Electron microscopy showed that MAGP-2 specifically associates with microfibrils and that elastin globules primarily colocalize with MAGP-2-associated microfibrils, suggesting that microfibril-associated MAGP-2 facilitates elastic fiber assembly. MAGP-2 overexpression did not change levels of matrix-associated fibrillin-1, MAGP-1, fibulin-2, fibulin-5, or emilin-1, suggesting that microfibrils and other elastic fiber-associated proteins known to regulate elastogenesis do not mediate MAGP-2-induced elastic fiber assembly. Moreover, mutation analysis showed that MAGP-2 does not stimulate elastic fiber assembly through its RGD motif, suggesting that integrin receptor binding does not mediate MAGP-2-induced elastic fiber assembly. Because MAGP-2 interacts with Jagged-1 that controls cell-matrix interaction and cell motility, two key factors in elastic fiber macroassembly, microfibril-associated MAGP-2 may stimulate elastic fiber macroassembly by targeting the release of elastin globules from the cell membrane onto developing elastic fibers.
Lemaire
Raphael
R
Department of Medicine, Arthritis Center, Boston University School of Medicine, Boston, Massachusetts 02118, USA. rlemaire@bu.edu
Bayle
Julie
J
Mecham
Robert P
RP
Lafyatis
Robert
R
D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003285:Contractile Proteins / Q000502:physiology*; D004318:Doxycycline / Q000494:pharmacology; D004548:Elasticity; D016326:Extracellular Matrix Proteins / Q000737:chemistry / Q000502:physiology*; D005347:Fibroblasts / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000378:metabolism; D051379:Mice; D020894:Microfibrils / Q000378:metabolism*; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D010957:Plasmids / Q000378:metabolism; D000072260:RNA Splicing Factors; D011994:Recombinant Proteins / Q000737:chemistry; D014162:Transfection
D003285:Contractile Proteins; D016326:Extracellular Matrix Proteins; C120769:Fbln5 protein, mouse; D000072260:RNA Splicing Factors; D011994:Recombinant Proteins; C016598:microfibrillar protein; D049452:Green Fluorescent Proteins; D004318:Doxycycline