Forkhead box O3 (FOXO3) transcription factor mediates apoptosis in BCG-infected macrophages

Enhanced apoptosis of BCG-infected macrophages has been shown to induce stronger dendritic cell-mediated cross-priming of T cells, leading to higher protection against tuberculosis (TB). Uncovering host effectors underlying BCG-induced apoptosis may then prove useful to improve BCG efficacy through priming macrophage apoptosis. Her we report that BCG-mediated apoptosis of human macrophages relies on FOXO3 transcription factor activation. BCG induced a significant apoptosis of THP1 (TDMs) and human monocytes (MDMs)-derived macrophages when a high moi was used, as shown by annexin V/7-AAD staining. BCG-induced apoptosis was associated with dephosphorylation of the prosurvival activated threonine kinase (Akt) and its target FOXO3. Cell fractionation and immunofluorescence microscopy showed translocation of FOXO3 to the nucleus in BCG-infected cells, concomitantly with an increase of FOXO3 transcriptional activity. Moreover, FOXO3 expression knock-down by small interfering RNA (siRNA) partially inhibited the BCG-induced apoptosis. Finally, real-time quantitative PCR (qRT-PCR) analysis of the expression profile of BCG-infected macrophages showed an upregulation of two pro-apoptotic targets of FOXO3, NOXA and p53 upregulated modulator of apoptosis (PUMA). Our results thus indicate that FOXO3 plays an important role in BCG-induced apoptosis of human macrophages and may represent a potential target to improve vaccine efficacy through enhanced apoptosis-mediated cross-priming of T cells.