BLaER1 macrophage model for the investigation of RNA stimulation and viral infections
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Abstract
During infections, the endolysosomal Toll-like receptor (TLR8) in macrophages of the innate immune response recognizes foreign RNA and triggers a signaling cascade leading to the release of cytokines and immune system activation. However, TLR8 stimulation is inhibited by 2’-O-methylation (2’-O-me), which is a known marker of eukaryotic RNA, through an unknown mechanism. To elucidate the molecular processes upstream of TLR8, we identified 2’-O-me-dependent RNA-binding proteins using the BLaER1 macrophage model and mass spectrometry-based proteomics. In this context, TRMT61A demonstrated increased binding to unmodified RNA, thereby suggesting a potential role in RNA processing prior to TLR8 activation. Moreover, studies of the subcellular localization of RNases and work by our collaborators confirmed the role of RNase6 in 2’-O-me-dependent processing of bacterial RNA as prerequisite for TLR8 activation. Furthermore, we characterized the BLaER1 macrophage model by analyzing proteomes during transdifferentiation and established it as model for orthoebolavirus infections. We demonstrated the susceptibility and permissiveness of BLaER1 to the virus and, using time-resolved proteome and secretome data, identified host factors that could explain orthoebolavirus pathogenicity and modulation of the immune response. Overall, our results establish BLaER1 macrophages as versatile model to studying RNA recognition and to expand our understanding of orthoebolavirus pathogenicity.