In this study, we aimed to determine the role of EVs derived from Lm-infected DCs in the innate response, and the potential role of HDAC6 in DEV loading under these conditions. Our results show that Lm infection induced an elevated production of DEVs. Lm infection also altered protein sorting. Interestingly, HDAC6 deletion altered DEV protein sorting, although it did not affect acetylation of DEV-loaded proteins. However, protein ubiquitination was severely altered. Here, we show that the protein content of dendritic cells (DCs) and their secreted EVs (DEVs) vary dramatically during Lm infection. The proteomic profiles of DCs and DEV differ depending on whether HDAC6 is expressed or not.