Endocytic trafficking from the small intestinal brush border probed with FM dye

Area of Science:

• Cell Biology
• Gastroenterology
• Membrane Trafficking

Background:

• The small intestinal brush border is crucial for nutrient absorption and pathogen defense.
• Brush border enzymes are organized in lipid rafts, but their dynamic properties are poorly understood.
• The role of the terminal web cytoskeleton in membrane trafficking is unclear.

Purpose of the Study:

• To investigate the dynamic properties of endocytic membrane trafficking from the small intestinal brush border.
• To determine the role of the terminal web cytoskeleton in regulating apical endocytosis.
• To understand how brush border enzyme localization affects endocytosis.

Main Methods:

• Organ-cultured pig intestinal mucosal explants.
• Fluorescent lipophilic dye (FM dye) for live imaging of membrane trafficking.
• Cholera toxin B subunit as a ligand for receptor-mediated endocytosis.

Main Results:

• Constitutive endocytosis occurs at the brush border, forming early endosomes within the terminal web region.
• Apical endocytosis is enhanced by cholera toxin B subunit, leading to deeper cytoplasmic trafficking.
• Key brush border enzymes (alkaline phosphatase, aminopeptidase N) are excluded from endocytic vesicles.

Conclusions:

• The terminal web cytoskeleton restricts the further trafficking of apical early endosomes.
• This restriction contributes to the intestinal permeability barrier function.
• Brush border enzyme organization and endocytic pathways are distinct.