Blocking anti-PD-1 monoclonal antibodies (mAb) have shown unprecedented success in cancer immune therapies by overcoming activation brakes induced by the immune checkpoint receptor PD-1 expressed on T cells, thus improving anti-tumour T cell immune responses in patients. While immune checkpoint receptors are commonly associated with T cells, we have shown that mouse dendritic cell subsets upregulate PD-1 in response to activation and in the tumour environment. DC lacking PD-1 develop normally and express normal levels of activation markers. However, in the absence of PD-1 or following anti-PD-1 mAb therapies, the cDC1 subset produces higher levels of the cytokine interferon (IFN)-λ, a type III IFN with anti-viral and anti-tumour properties. Furthermore, PD-1-KO cDC1 are hyperstimulatory towards T cells. In humans, blood DC express PD-1 at variable levels between individuals and PD-1 blocking in cDC1 also increases their capacity to produce IFN-λ, similarly to their mouse counterparts. Altogether, these data show not only T cells, but also DC, are regulated by the immune checkpoint PD-1 and that PD-1 blocking enhances their functions. These findings provide new insights into the mechanism of action of cancer anti-PD-1 immunotherapies and may aid improving their efficacy and their overall safety.