A gene silencing pipeline to interrogate human cDC1 and pDC development and functions

Type 1 conventional dendritic cells (cDC1s) and plasmacytoid dendritic cells (pDCs) are thought to be critical for anti-tumor or antiviral immunity. In vitro differentiation systems have unlocked the ability to produce large numbers of these cells. However, a method is lacking to systematically identify the cell-intrinsic factors controlling their differentiation and functions that remain therefore poorly understood, in contrast to the situation in mice. Here, we developed a workflow for efficient gene silencing and its tracing in human cDC1s/pDCs generated in vitro. As proof of concept, we confirmed the key role of IRF8 in their development, and of IRF7/MyD88 in human pDC production of interferons-α/λ. We found that SAMHD1 and RAB7B promote human cDC1 differentiation, while SEPT3 promotes human pDC differentiation. We also found that PPT1 and RAB5 are required for optimal differentiation of pDCs and cDC1s. Finally, we identified BCL11A, PPT1 and RAB7 as novel HIV-1 restriction factors in cDC1s/pDCs. This approach will enable broader genetic screens to advance our understanding of human cDC1s/pDCs and harness them against viral infections or cancer. ### Competing Interest Statement X Luo, N.M. and M.D. are inventors on a patent application filed on the described method. The remaining authors declare no competing interests.