RESUMEN
Microfibril-associated glycoprotein 4 (MFAP4) is a 36-kDa extracellular matrix glycoprotein with critical roles in organ fibrosis, chronic obstructive pulmonary disease, and cardiovascular disorders, including aortic aneurysms. MFAP4 multimerises and interacts with elastogenic proteins, including fibrillin-1 and tropoelastin, and with cells via integrins. Structural details of MFAP4 and its potential interfaces for these interactions are unknown. Here, we present a cryo-electron microscopy structure of human MFAP4. In the presence of calcium, MFAP4 assembles as an octamer, where two sets of homodimers constitute the top and bottom halves of each octamer. Each homodimer is linked together by an intermolecular disulphide bond. A C34S missense mutation prevents disulphide-bond formation between monomers but does not prevent octamer assembly. The atomic model, built into the 3.55 Å cryo-EM map, suggests that salt-bridge interactions mediate homodimer assembly, while non-polar residues form the interface between octamer halves. In the absence of calcium, an MFAP4 octamer dissociates into two tetramers. Binding studies with fibrillin-1, tropoelastin, LTBP4, and small fibulins show that MFAP4 has multiple surfaces for protein-protein interactions, most of which depend upon MFAP4 octamer assembly. The C34S mutation does not affect these protein interactions or cell interactions. MFAP4 assemblies with fibrillin-1 abrogate MFAP4 interactions with cells.
Asunto(s)
Microscopía por Crioelectrón , Proteínas de la Matriz Extracelular , Fibrilina-1 , Microfibrillas , Tropoelastina , Humanos , Adipoquinas , Calcio/metabolismo , Proteínas de la Matriz Extracelular/metabolismo , Proteínas de la Matriz Extracelular/química , Proteínas de la Matriz Extracelular/genética , Fibrilina-1/metabolismo , Fibrilina-1/genética , Fibrilina-1/química , Glicoproteínas , Células HEK293 , Microfibrillas/metabolismo , Microfibrillas/química , Microfibrillas/ultraestructura , Modelos Moleculares , Mutación Missense , Unión Proteica , Multimerización de Proteína , Tropoelastina/metabolismo , Tropoelastina/química , Tropoelastina/genéticaRESUMEN
The microfibril-elastin fiber system, an important constituent of the extracellular matrix, was studied in the rat left atrioventricular valve to investigate the interrelationship of oxytalan, elaunin and elastic fibers in left atrioventricular valve morphology. The elastin fibers forms continuous bundles observed along the length of the valve in atrial and ventricular layers and oriented parallel to endothelium. The elaunin and oxytalan fibers are distributed in the thickest fiber bundles along the length of the valve. The thinner fibers which radiated towards both the atrial and spongiosa layers, either as isolated or arborescent fiber bundles were identified as oxytalan fibers. With transmission electron microscopy elastic fibers were seen mainly in the atrial layer. The spongiosa layer was composed of elaunin and oxytalan fibers and ventricular layer showed elaunin fibers arranged in continuous bundles parallel to the endothelium. Both fibrillin and elastin were seen and identified by immunocytochemistry with colloidal gold in the left atrioventricular valve spongiosa and atrial layers. These observations allow us to suggest that the microfibril-elastin fiber system plays a role in the mechanical protection and maintenance of the integrity of the rat left atrioventricular valve.
Fue estudiado el sistema de fibras microfibrillas-elastina, un componente importante de la matriz extracelular, en la valva atrioventricular izquierda de rata, con la finalidad de investigar la interrelación de oxitalán, elaunin y fibras elásticas en la morfología de dicha valva. Las fibras de elastina forman paquetes continuos a lo largo de la valva en las capas atriales y ventriculares, orientadas paralelamente al endotelio. Las fibras de elaunin y oxitalán se distribuyen en haces de fibras más gruesas a lo largo de la valva. Las fibras más delgadas, las cuales se irradiaban hacia las capas atrial y esponjosa, ya sea como haces de fibras aisladas o arborescentes, fueron identificadas como fibras oxitalán. En la capa atrial a través de microscopía electrónica de transmisión se observaron principalmente fibras elásticas. La capa esponjosa estaba compuesta por fibras de elaunin y oxitalán; la capa ventricular mostró fibras de elaunin dispuestas en haces continuos paralelos al endotelio. Tanto fibrilina y elastina se observaron e identificaron por inmunocitoquímica con oro coloidal en las capas esponjosa y atrial de la valva atrioventricular izquierda. Estas observaciones nos permiten sugerir que el sistema de fibras de elastina-microfibrillas tienen participación en la protección mecánica y la mantención de la integridad de la valva atrioventricular izquierda en la rata.