Role of DGCR8 in differentiation and function of murine macrophages

One-third of the world population is latently infected with Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB). Mtb is an intracellular bacterium that developed successful survival strategies to subvert innate and adaptive immune responses and to replicate in host macrophages. The most abundant glycolipid in the cell wall of virulent Mtb is the mycobacterial cord factor (TDM). The cord factor contributes to mycobacterial survival by delaying phagosomal maturation and by inhibiting expression of costimulatory molecules and cytokines. On the other hand, TDM is also recognized by host PRRs and induces important immune responses against mycobacterial infections, including granuloma formation, macrophage activation, and Th1/Th17 adjuvanticity. microRNAs (miRNAs), a class of small, regulatory RNAs, are critically involved in the regulation of innate immune responses against TB. However, there is also emerging evidence that Mtb exploits host miRNAs as immune modulators to avoid detection by the host immune system. In this project, the involvement of miRNA-based regulatory mechanisms in macrophage responses to the mycobacterial cord factor was elucidated. To this end, DGCR8-deficient bone marrow-derived macrophages (BMM) with a block in miRNA biogenesis were generated by conditional and inducible deletion of DGCR8 during in vitro macrophage differentiation of DGCR8fl/fl ROSA26-CreERT2+/- bone marrow progenitors. DGCR8 deficiency did not interfere with BMM differentiation or the capacity for phagocytosis and IL-6 production, yet it remarkably affected mature BMM numbers. Further, RNA sequencing analyses of gene expression profiles revealed a modest, though consistent type I IFN signature in resting DGCR8-/- BMM. TDM stimulation further increased expression of respective IFN-stimulated genes (ISGs) and induced additional ISGs as well as IFNb itself. As hyperinduction of ISGs could be largely reversed by neutralizing antibodies to type I interferons, overexpression of those genes can be, at least partially, explained by TDM-induced IFNb in DGCR8-/- BMM. In summary, this work shows that DGCR8 is dispensable for BMM maturation and the capacity for phagocytosis and IL-6 production in vitro, whereas it is required for BMM growth during differentiation. Moreover, DGCR8 and/ or miRNAs play a critical role in curbing macrophage responses to TDM and in avoiding an unintended expression of IFNb and ISGs.