Antigen-presenting cells (APCs) provide essential signals for Th cell differentiation. We used skin-relevant, fluorescently-labeled bacterial, helminth or fungal pathogens to track the skin APC populations that drive the development of IFNγ, IL-4 or IL-17-producing CD4+ T cells in vivo. By coupling single-cell RNA sequencing and flow cytometry technologies, we identified that immunization-induced transcriptional changes were predominantly restricted to monocytes and conventional migratory IRF4+ dendritic cells (migDC2s) positive for the fluorescent pathogen (Ag+). Depletion of migDC2s significantly blunted CD4+ T cell responses to all tested pathogens, indicating that migDC2s are necessary for directing the appropriate Th differentiation program. Indeed, Ag+ migDC2s displayed condition-specific expression of Th cell polarizing molecules including Il12a, Il12b, Il6 and Il23a. Interestingly, depletion of monocytes specifically impaired the differentiation of IFNγ+Tbet+ cells following immunization with Mycobacteria (Ms). Temporal reconstruction of single-cell RNA sequencing data highlighted a migDC2-NK-IFNγ-monocyte communication pathway in Ms-treated mice, which was required for optimal effector Th1 differentiation. Our study highlights the importance of innate immune cell-pathogen interactions in driving the transcriptional programs required for directing appropriate Th cell differentiation in vivo.