Human VPS34 is required for internal vesicle formation within multivesicular endosomes.

Area of Science:

• Cell Biology
• Molecular Biology
• Signal Transduction

Background:

• Activated epidermal growth factor receptors (EGFRs) internalize and traffic to multivesicular bodies (MVBs).
• Within MVBs, EGFRs are sorted into internal vesicles, distinct from recycling transferrin receptors.
• This sorting process is crucial for regulating EGFR signaling and degradation.

Purpose of the Study:

• To elucidate the molecular mechanisms governing EGFR sorting within MVBs.
• To determine the roles of phosphatidylinositol 3'-kinase (PI 3-kinase) and tyrosine kinase activity in EGFR trafficking.
• To investigate the impact of MVB inward vesiculation on signal transduction attenuation.

Main Methods:

• Utilized the PI 3-kinase inhibitor wortmannin to block internal vesicle formation.
• Employed microinjection of inhibitory antibodies to identify the specific PI 3-kinase (hVPS34).
• Investigated the trafficking of tyrosine kinase-negative EGFRs.

Main Results:

• Wortmannin treatment inhibits internal vesicle formation and causes EGFRs to accumulate on the MVB perimeter membrane.
• hVPS34 is identified as the PI 3-kinase responsible for MVB inward vesiculation.
• EGFRs are delivered to lysosomes independently of inward vesiculation, but selection for internal vesicles requires tyrosine kinase activity.
• EGFR retention on the MVB perimeter membrane leads to increased EGF-stimulated tyrosine phosphorylation.

Conclusions:

• EGFR selection for internal vesicles requires tyrosine kinase activity, while vesicle formation requires PI 3-kinase activity.
• Inward vesiculation within MVBs is directly involved in attenuating epidermal growth factor receptor signal transduction.