Ribosomal protein RPL20B modulates cytokine production by macrophage in reponse to yeast

Phagocytosis is the process by which myeloid phagocytes bind and internalize potentially dangerous microbes. During phagocytosis, innate immune receptors and the associated signaling adapters are localized to the maturing phagosome compartment. We used proximity labeling of phagosomal contents (PhagoPL) to identify proteins localizing to phagosomes containing model yeast and bacteria and identified programmed death-ligand 1 (PD-L1) as a protein that specifically enriches in phagosomes containing yeast. We found that PD-L1 directly binds to yeast upon processing in phagosomes. By surface display library screening, we identified the ribosomal protein RPL20B as a fungal protein ligand for PD-L1. Using an auxin-inducible depletion system, we found that detection of RPL20B by macrophages influences production of a subset of inflammatory cytokines and chemokines produced when macrophages encounter yeast. To investigate whether host detection of RPL20B during phagocytosis has any influence on inflammatory responses of macrophages to yeast, GM-CSF primed bone marrow-derived macrophages (BMDMs) were stimulated with either RPL20B sufficient-yeast (w/oIAA yeast) or the PRL20B depleted-yeast (induced yeast) for 4h. Comparative gene expression profiling analysis were performed using data obtained from RNA-seq of w/oIAA or induced yeast-stimulated BMDMs. 3 biological replicates per condition were independently prepared for RNA-sequencing (different batches of BMDMs stimulated with different batches of yeast preparations).