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Published December 2016 | Published
Journal Article Open

Visualizing enzymatic and cellular activities during tissue morphogenesis using ex vivo twophoton FRET microscopy

Abstract

Morphogenesis is a critical step in embryogenesis during which tissue/organ transforms into its functional shape. The drastic change in tissue morphology signifies active cellular reorganization and extra‐cellular matrix remodeling. Enzymatic activity vital to this process, especially that of matrix metalloproteinase (MMP), attracts a lot of research interests but proves difficult to investigate using traditional methods. Förster resonance energy transfer (FRET) has emerged as a powerful technique for investigating biochemical interaction, but its usefulness in live tissues has not been well established. Here we use a genetically encoded FRET biosensor with ex vivo two‐photon microscopy to demonstrate a clear increase in MT1‐MMP activity at the tip of a growing feather bud, which coincides with increased cellular motion, sometimes across the epidermal‐dermal border, and weakened laminin structure in the basement membrane. Laminin and collagen III are newly synthesized at the tip, strengthening the newly formed structure. These events also exhibit spatio‐temporal correlation with increased Src activity observed using another genetically encoded FRET biosensor. Our ex vivo biochemistry approach provides insights into the spatial and temporal profile of enzyme activities, and together with traditional approaches, offers a comprehensive understanding of the morphogenetic process. Furthermore, it can readily lend itself to the studying of other biological processes, such as cancer metastasis, where enzymatic activity plays a central role.

Additional Information

© 2016 American Society for Cell Biology.

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