By crossing mice expressing Cre recombinase under the control of the Clec9a locus to Rosa26-STOPflox-yellow fluorescent protein (YFP) reporter mice, we have recently generated a genetic model with which to fate map the progeny of DNGR-1+ CDP and preDC [ 21••]. Although Clec9a-Cre reporter mice suffer from limitations, as DNGR-1

is also expressed on CD8α+/CD103+ cDCs and to a lower extent on pDCs, we were able to demonstrate YFP expression in DCs but not monocytes or macrophages even after intestinal inflammation and Listeria monocytogenes infection [ 21••]. Importantly, using Clec9a-Cre reporter mice, we identified CDP-derived cells within CD64+ cell populations I-BET-762 mw previously thought to represent monocytes/macrophages [ 21••, 85 and 86]. CD64+ CDP-derived cells are especially

frequent in kidneys, where they resemble yolk sac-derived F4/80hi tissue-resident MØs, appear to lack Zbtb46 expression [ 73•] and where their affiliation as DCs or macrophages Venetoclax in vitro has been debated [ 87]. The presence of a few YFP+ cells in the CD64 component of lung and small intestine indicates the existence of these atypical CDP-derived cells also in other tissues [ 21••]. Notably, CD64+ kidney DCs stimulated naïve T cells in vitro, although less efficiently than CD11b+ cDCs [ 21••]. Thus, fate mapping of DC precursors reveals previously unappreciated heterogeneity among mononuclear phagocytes, raising the question of why cells of distinct ontogeny but overlapping phenotype exist in the same tissue. More detailed analyses of such atypical CDP-derived cells,

for instance by transcriptome profiling, will contribute Exoribonuclease to elucidating their function and help determine how it is shaped by the local environment. The classification of DCs based on phenotypic and functional properties that are often shared with other cell types has led to difficulties in cell identification and even debate over the existence of DCs as a discrete leukocyte lineage. Phenotype-based and function-based definitions are inherently problematic as functional roles and phenotypic markers often change under the influence of environmental cues. While mononuclear phagocyte classification may be considered a semantic issue, it is key to scientific communication. It needs to be robust enough to withstand arguments pertaining to levels of surface marker expression or degree of T cell stimulatory capacity to make it clearly accessible to all researchers inside and outside the field. The identification of distinct developmental precursors underscores that cDCs, pDCs and monocytes constitute separate cell types and enables a move towards cell classification based on ontogeny. Importantly, ontogenetic definitions are independent of functional or phenotypic properties, allowing the investigation of the full spectrum of phagocyte activity in an unbiased manner.