Direct visualization of protease activity on cells migrating in three-dimensions

Area of Science:

• Cell Biology
• Biochemistry
• Biotechnology

Background:

• Proteases play a critical role in cell migration and invasion.
• Understanding protease activity at the site of action is crucial for studying these processes.
• Previous methods lacked the resolution to pinpoint protease activity during live-cell migration.

Purpose of the Study:

• To develop a novel fluorescent biosensor for detecting localized extracellular protease activity.
• To investigate the role of matrix metalloproteases (MMPs) in cell migration and invasion through three-dimensional collagen matrices.
• To clarify discrepancies in previous studies regarding protease contributions to metastasis.

Main Methods:

• Designed a fluorescent biosensor that fluoresces upon cleavage of a peptide site in interstitial collagen by proteases.
• Utilized the biosensor in three-dimensional collagen matrices to image protease activity during live-cell migration.
• Tested the necessity of protease activity for cell migration in native and pepsin-treated collagen matrices.

Main Results:

• Protease activity was localized at the polarized leading edge of migrating tumor cells.
• Protease activity was found to be essential for cell migration in native cross-linked collagen matrices.
• The biosensor functions without requiring cell transfection or modification.

Conclusions:

• The novel biosensor provides high-resolution, site-specific reporting of extracellular protease activity.
• Protease activity at the leading edge is essential for tumor cell migration in complex extracellular matrices.
• This technology offers insights into cell migration mechanisms and resolves previous study discrepancies.