N-glycosylation of mouse TRAIL-R and human TRAIL-R1 enhances TRAIL-induced death.

APO2L/TRAIL (TNF-related apoptosis-inducing ligand) induces death of tumor cells through two agonist receptors, TRAIL-R1 and TRAIL-R2. We demonstrate here that N-linked glycosylation (N-glyc) plays also an important regulatory role for TRAIL-R1-mediated and mouse TRAIL receptor (mTRAIL-R)-mediated apoptosis, but not for TRAIL-R2, which is devoid of N-glycans. Cells expressing N-glyc-defective mutants of TRAIL-R1 and mouse TRAIL-R were less sensitive to TRAIL than their wild-type counterparts. Defective apoptotic signaling by N-glyc-deficient TRAIL receptors was associated with lower TRAIL receptor aggregation and reduced DISC formation, but not with reduced TRAIL-binding affinity. Our results also indicate that TRAIL receptor N-glyc impacts immune evasion strategies. The cytomegalovirus (CMV) UL141 protein, which restricts cell-surface expression of human TRAIL death receptors, binds with significant higher affinity TRAIL-R1 lacking N-glyc, suggesting that this sugar modification may have evolved as a counterstrategy to prevent receptor inhibition by UL141. Altogether our findings demonstrate that N-glyc of TRAIL-R1 promotes TRAIL signaling and restricts virus-mediated inhibition

Targeted delivery of mycobacterial antigens to human dendritic cells via Siglec-7 induces robust T cell activation

Lipids from mycobacteria can be presented to human T cells by group 1 CD1 Ag-presenting molecules (CD1a, CD1b, and CD1c). Group 1 CD1-restricted T cells are activated by lipid Ags presented by myeloid dendritic cells (DCs), after which they generate antibacterial effector functions, including IFN-γ secretion and cytolysis. Thus, mycobacterial lipids are being investigated as components of novel vaccines for mycobacterial infections. In this study we show that the mycobacterial lipid Ag C80 glucose-6-monomycolate can be delivered to human CD1b(+) DCs via targeted liposomal nanoparticles, leading to robust group 1 CD1-restricted activation of T cells. Targeting was achieved by decorating the liposomes with a high-affinity glycan ligand of sialic acid-binding Ig-like lectin (Siglec)-7, a siglec receptor expressed on DCs that mediates rapid endocytosis and transport of its cargo to lysosomes. An Ab to Siglec-7 completely blocked the binding of targeted liposomes to human monocyte-derived DCs (Mo-DCs), demonstrating their targeting specificity. Mo-DCs pulsed with targeted liposomes containing C80 glucose-6-monomycolate more potently activated a CD1b-restricted T cell line relative to Mo-DCs pulsed with free lipid Ag or antigenic liposomes without Siglec-7 ligand. These data suggest that the endocytic function of Siglec-7 can be exploited to deliver glycolipid Ags to their target cell and increase the efficiency of display to T cells

Targeted delivery of mycobacterial antigens to human dendritic cells via Siglec-7 induces robust T cell activation

Lipids from mycobacteria can be presented to human T cells by group 1 CD1 Ag-presenting molecules (CD1a, CD1b, and CD1c). Group 1 CD1-restricted T cells are activated by lipid Ags presented by myeloid dendritic cells (DCs), after which they generate antibacterial effector functions, including IFN-γ secretion and cytolysis. Thus, mycobacterial lipids are being investigated as components of novel vaccines for mycobacterial infections. In this study we show that the mycobacterial lipid Ag C80 glucose-6-monomycolate can be delivered to human CD1b(+) DCs via targeted liposomal nanoparticles, leading to robust group 1 CD1-restricted activation of T cells. Targeting was achieved by decorating the liposomes with a high-affinity glycan ligand of sialic acid-binding Ig-like lectin (Siglec)-7, a siglec receptor expressed on DCs that mediates rapid endocytosis and transport of its cargo to lysosomes. An Ab to Siglec-7 completely blocked the binding of targeted liposomes to human monocyte-derived DCs (Mo-DCs), demonstrating their targeting specificity. Mo-DCs pulsed with targeted liposomes containing C80 glucose-6-monomycolate more potently activated a CD1b-restricted T cell line relative to Mo-DCs pulsed with free lipid Ag or antigenic liposomes without Siglec-7 ligand. These data suggest that the endocytic function of Siglec-7 can be exploited to deliver glycolipid Ags to their target cell and increase the efficiency of display to T cells

Cooperative loss of RAS feedback regulation drives myeloid leukemogenesis

RAS network activation is common in human cancers, and in acute myeloid leukemia (AML) this activation is achieved mainly through gain-of-function mutations in KRAS, NRAS or the receptor tyrosine kinase FLT3. We show that in mice, premalignant myeloid cells harboring a KrasG12D allele retained low levels of Ras signaling owing to negative feedback involving Spry4 that prevented transformation. In humans, SPRY4 is located on chromosome 5q, a region affected by large heterozygous deletions that are associated with aggressive disease in which gain-of-function mutations in the RAS pathway are rare. These 5q deletions often co-occur with chromosome 17 alterations involving the deletion of NF1 (another RAS negative regulator) and TP53. Accordingly, combined suppression of Spry4, Nf1 and p53 produces high levels of Ras signaling and drives AML in mice. Thus, SPRY4 is a tumor suppressor at 5q whose disruption contributes to a lethal AML subtype that appears to acquire RAS pathway activation through a loss of negative regulators