Macropinocytosis and TAK1 mediate anti-inflammatory to pro-inflammatory macrophage differentiation by HIV-1 Nef

Area of Science:

• Immunology
• Virology
• Cell Biology

Background:

• Macrophages (MΦ) are key immune cells with distinct M1 (pro-inflammatory) and M2 (anti-inflammatory) phenotypes.
• HIV-1 infection is associated with sustained immune activation, the mechanisms of which are not fully understood.
• Soluble HIV-1 proteins, particularly Nef, have been implicated in modulating macrophage function.

Purpose of the Study:

• To elucidate the mechanisms by which HIV-1 Nef protein preferentially activates M2 macrophages.
• To investigate the role of TAK1 (TGF-β-activated kinase 1) and macropinocytosis in Nef-induced M2 macrophage polarization.
• To understand how Nef-mediated M2 macrophage activation contributes to immune dysregulation in HIV-1.

Main Methods:

• Comparative analysis of M-CSF-derived M2 and GM-CSF-derived M1 macrophages.
• Assessment of TAK1 activation pathways, including MAP kinases and NF-κB.
• Utilizing TAK1 inhibitors (5Z-7-oxozeaenol) and dominant-negative mutants of TAK1 and TRAF2.
• Investigating Nef protein internalization via macropinocytosis using inhibitors (EIPA).

Main Results:

• Nef preferentially activated TAK1 in M2 macrophages but not in M1 macrophages.
• Nef mutants deficient in MAP kinase and NF-κB activation also failed to activate TAK1.
• Inhibition of TAK1 or its upstream regulator TRAF2 blocked Nef-induced signaling and M2 to M1-like polarization.
• Nef internalization was significantly higher in M2 macrophages due to their elevated macropinocytosis activity.
• Macropinocytosis inhibition blocked Nef entry into M2 macrophages.

Conclusions:

• HIV-1 Nef protein exploits the higher macropinocytosis activity of M2 macrophages for efficient entry.
• Nef activates TAK1 signaling in M2 macrophages, driving their differentiation into M1-like cells.
• This Nef-mediated M2 to M1 polarization contributes to the sustained immune activation observed in HIV-1 infection.