Adenovirus triggers macropinocytosis and endosomal leakage together with its clathrin-mediated uptake

Area of Science:

• Cell Biology
• Virology
• Molecular Biology

Background:

• Adenovirus (Ad) entry into epithelial cells is initiated by binding to the coxsackie B virus Ad receptor.
• Ad uptake is further facilitated by the activation of alphav integrin coreceptors.

Purpose of the Study:

• To investigate the role of clathrin-mediated endocytosis and macropinocytosis in Adenovirus type 2 (Ad2) infection.
• To elucidate the mechanisms by which Ad2 utilizes cellular endocytic pathways for entry and infection.

Main Methods:

• Expression of dominant-negative inhibitors (clathrin hub, eps15, K44A-dynamin) to block specific endocytic pathways.
• Monitoring fluid phase tracer uptake and transferrin uptake to assess endocytic activity.
• Investigating the dependence of macropinocytosis on specific cellular components (alphav integrins, PKC, F-actin, Na+/H+ exchanger).

Main Results:

• Ad2 uptake was inhibited by blocking clathrin-dependent endocytosis.
• Ad2 significantly stimulated macropinocytosis, a fluid uptake process, independent of transferrin uptake.
• Macropinocytosis stimulation and subsequent macropinosomal lysis, releasing contents into the cytosol, were dependent on alphav integrins and other signaling molecules.
• Macropinosomal lysis occurred even when viral escape was blocked, indicating Ad-induced signaling controls this process.

Conclusions:

• Adenovirus type 2 utilizes clathrin-dependent endocytosis for cellular entry.
• Ad2 actively induces and exploits macropinocytosis, leading to macropinosome leakage and release of contents into the cytosol.
• Cell surface signaling by Ad controls macropinosome formation and lysis, correlating with viral escape and infection.