Measuring Endocytosis and Endosomal Uptake at Single Cell Resolution

Area of Science:

• Cell Biology
• Developmental Biology
• Stem Cell Biology

Background:

• Endocytosis plays a vital role in embryonic stem cell (ESC) functions.
• It is essential for FGF-signaling and ERK activation during the ESC to epiblast-like cell (EpiLC) transition.
• Current quantitative methods lack single-cell resolution for studying endocytosis in these dynamic cell populations.

Purpose of the Study:

• To develop and present an easily adaptable, multiplexable platform for monitoring and quantifying endocytosis.
• To enable single-cell resolution analysis of endosomal uptake in live cells.
• To investigate the roles and regulation of endocytosis in pluripotent cell transitions.

Main Methods:

• Development of a quantitative, live-cell imaging platform.
• Utilizing multiplexable assays for monitoring endosomal uptake.
• Application to receptor-mediated, non-receptor-mediated, and pinocytosis mechanisms.

Main Results:

• The presented methods allow for the quantification of endocytosis at single-cell resolution.
• The platform is adaptable for various endocytic pathways, including nonspecific pinocytosis.
• Demonstrated feasibility for studying endocytosis in transitioning pluripotent cells.

Conclusions:

• The developed platform provides a powerful tool for dissecting endocytosis in stem cell biology.
• Enables detailed investigation of endocytosis regulation and function in ESCs and EpiLCs.
• Facilitates a deeper understanding of pluripotency maintenance and cell fate decisions.