Transient expression of the class II heterodimer, HLA-DR, induces intracellular accumulation of Gag and a marked reduction in extracellular virus release from producer cells [8] . This previous study focused on HLA-DR in the absence of Ii and HLA-DM, critical components of the antigen processing and presentation pathway which influence MHC class II trafficking. To determine if expression of the complete MHC class II pathway could overcome Gag and virus retention we used CIITA to coordinately express these genes. Cells were either transiently (pcCIITA) or stably (A293T-CIITA) transfected with CIITA ( Figure S1 ) and HIV-1 NL4-3 DNA, and Gag localization was monitored by immunofluorescence and compared to cells transiently expressing the HLA-DR a and b heterodimers (pcDRab1b5), or vector-only (pcDNA3.1). As expected, a weak but uniform Gag signal was present in vector-only transfected cells (Figure 1Aa ) and transient HLA-DR expression induced a marked redistribution of Gag as indicated by a dense, punctuate Gag staining pattern (Figure 1Ad ). Conversely, Gag accumulation was not observed when CIITA was either transiently or stably expressed (Figure 1Ab or c, respectively). Therefore, transient expression of HLA-DR in producer cells does indeed lead to Gag retention, which is not observed in the presence of CIITA. However, recapitulation of the MHC class II pathway in trans (pcDRab1b5+Ii+HLA-DMab), also resulted in dense accumulation of Gag signal (Figure 1Ae ), suggesting that CIITA-mediated coordinate activation of HLA-DR, -DM and Ii expression is insufficient to overcome Gag retention. Flow cytometric analysis confirmed these findings, as cells transfected with HLA-DR heterodimers and/or co-transfected with some or all of the components of the class II antigen presentation pathway stained as Gag hi , indicating Gag accumulation ( Figure 1B and C). However, this population was absent in cells transiently or stably expressing CIITA ( Figure 1B and C) . Therefore, absence of Gag accumulation in CIITA expressing cells is likely not due to its transactivation of the MHC class II antigen presentation pathway.