Mg132

Macrophages were seeded at 0.2 × 10⁶ per well in a six-well plate. The next day, cells were infected with TB40-delUL16-eGFP for 2 h. Twenty-four hours later, cells were inoculated with dimethyl sulfoxide (DMSO), 1 μM MG132 (AdipoGen Life Sciences), 500 nM bafilomycin A1 (AdipoGen), or 0.1 μM MLN4924 (Chemgood) for 6 h. Afterwards, macrophages were collected and stained for FACS analysis.

Rabbit polyclonal anti-MuLV p30 antiserum was described previously [13 (link)]. Mouse monoclonal anti-HIV-1 p24^CA antibody (YDHIVgp24) was purchased from MyBioSource (San Diego, CA, USA). β-Tubulin was used for the loading control in western blots and was detected by rabbit anti-β-Tubulin antibody (Cell Signaling Technology, Danvers, MA, USA). For western blots, we used an anti-mouse IgG antibody conjugated with horseradish peroxidase (Thermo Fisher Scientific, Waltham, MA, USA) and an anti-rabbit IgG antibody conjugated with horseradish peroxidase (Thermo Fisher Scientific, Waltham, MA, USA). The antibodies for LC3 (Cell Signaling Technology) and p62/SQSTM1 (Proteintech Group, Inc., Rosemont, IL, USA) were used to check autophagic flux in the cells expressing Arf6Q67L. To examine the role of Arf6 in MuLV release, the following chemical inhibitors were used: UNC3230 (MedChemExpress, Monmouth Junction, NJ, USA), ML299 (Aobious Inc., Gloucester, MA, USA), FIPI (Tocris, Minneapolis, MN, USA), LY294002, wortmannin, rapamycin, and bafilomycin A1 (Cayman Chemical, Ann Arbor, MI, USA). MG132 and lactacystin were purchased from AdipoGen Life Sciences (San Diego, CA, USA).