Elastic fiber formation: a dynamic view of extracellular matrix assembly using timer reporters.
To study the dynamics of elastic fiber assembly, mammalian cells were transfected with a cDNA construct encoding bovine tropoelastin in frame with the Timer reporter. Timer is a derivative of the DsRed fluorescent protein that changes from green to red over time and, hence, can be used to distinguish new from old elastin. Using dynamic imaging microscopy, we found that the first step in elastic fiber formation is the appearance of small cell surface-associated elastin globules that increased in size with time (microassembly). The elastin globules are eventually transferred to pre-existing elastic fibers in the extracellular matrix where they coalesce into larger structures (macroassembly). Mechanical forces associated with cell movement help shape the forming, extracellular elastic fiber network. Time-lapse imaging combined with the use of Timer constructs provides unique tools for studying the temporal and spatial aspects of extracellular matrix formation by live cells.
Kozel
Beth A
BA
Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Rongish
Brenda J
BJ
Czirok
Andras
A
Zach
Julia
J
Little
Charles D
CD
Davis
Elaine C
EC
Knutsen
Russell H
RH
Wagenseil
Jessica E
JE
Levy
Marilyn A
MA
Mecham
Robert P
RP
D000818:Animals; D002417:Cattle; D002460:Cell Line; D002465:Cell Movement / Q000502:physiology; D019902:Chondrocytes / Q000648:ultrastructure; D004547:Elastic Tissue / Q000254:growth & development / Q000378:metabolism*; D004549:Elastin / Q000032:analysis / Q000378:metabolism / Q000648:ultrastructure; D005109:Extracellular Matrix / Q000378:metabolism* / Q000648:ultrastructure; D005347:Fibroblasts / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D032389:Myocytes, Smooth Muscle / Q000648:ultrastructure; D051381:Rats; D013997:Time Factors; D014162:Transfection; D014333:Tropoelastin / Q000235:genetics / Q000378:metabolism
D008164:Luminescent Proteins; D014333:Tropoelastin; C413663:fluorescent protein 583; D004549:Elastin