Measuring abscission spatiotemporal dynamics using quantitative high-resolution microscopy

Area of Science:

• Cell Biology
• Molecular Biology
• Microscopy

Background:

• Endosomal Sorting Complex Required for Transport (ESCRT) machinery mediates crucial cellular processes, including cell division.
• Cytokinetic abscission, the final step in cell division, is a complex process involving ESCRT proteins.
• Understanding the spatiotemporal dynamics of ESCRT during abscission is vital for cell biology.

Purpose of the Study:

• To describe the application of spinning disk live cell imaging and structured illumination microscopy (SIM) for analyzing ESCRT-mediated abscission.
• To quantitatively define the dynamics and structural organization of abscission.
• To correlate structural data from SIM with dynamic information from live cell imaging.

Main Methods:

• Quantitative live cell imaging using spinning disk microscopy.
• High-resolution imaging with structured illumination microscopy (SIM).
• Development of a protocol to integrate dynamic and structural data.

Main Results:

• The study provides a detailed methodology for observing ESCRT-mediated abscission.
• It enables quantitative analysis of protein dynamics and structural organization during the process.
• A protocol is presented to correlate structural and dynamic data.

Conclusions:

• Spinning disk microscopy and SIM are powerful tools for studying the spatiotemporal characteristics of ESCRT-mediated abscission.
• The presented protocol allows for a comprehensive quantitative analysis of this critical cell division event.
• This approach enhances our understanding of the molecular mechanisms governing cell abscission.