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Many innate immune receptors function collaboratively to detect and elicit immune responses to pathogens, but the physical mechanisms that govern the interaction and signaling crosstalk between the receptors are unclear. In this study, we report that the signaling crosstalk between Fc gamma receptor (FcγR) and Toll-like receptor (TLR)2/1 can be overall synergistic or inhibitory depending on the spatial proximity between the receptor pair on phagosome membranes. Using a geometric manipulation strategy, we physically altered the spatial distribution of FcγR and TLR2 on single phagosomes. We demonstrate that the signaling synergy between FcγR and TLR2/1 depends on the proximity of the receptors and decreases as spatial separation between them increases. However, the inhibitory effect from FcγRIIb on TLR2-dependent signaling is always present and independent of receptor proximity. The overall cell responses are an integration from these two mechanisms. This study presents quantitative evidence that the nanoscale proximity between FcγR and TLR2 functions as a key regulatory mechanism in their signaling crosstalk.

The dengue virus (DENV) circulates between humans and mosquitoes and requires no other mammals or birds for its maintenance in nature. The virus is well-adapted to humans, as reflected by high-level viraemia in patients. To investigate its high adaptability, the DENV induction of host type-I interferon (IFN) was assessed in vitro in human-derived HeLa cells and compared with that induced by the Japanese encephalitis virus (JEV), a closely related arbovirus that generally exhibits low viraemia in humans. A sustained viral spread with a poor IFN induction was observed in the DENV-infected cells, whereas the JEV infection resulted in a self-limiting and abortive infection with a high IFN induction. There was no difference between DENV and JEV double-stranded RNA (dsRNA) as IFN inducers. Instead, the dsRNA was poorly exposed in the cytosol as late as 48 h post-infection (p.i.), despite the high level of DENV replication in the infected cells. In contrast, the JEV-derived dsRNA appeared in the cytosol as early as 24 h p.i. Our results provided evidence for the first time in DENV, that concealing dsRNA in the intracellular membrane diminishes the effect of the host defence mechanism, a strategy that differs from an active suppression of IFN activity.

T LYMPHOCYTES reactive to pancreatic beta-cells are thought to have a central role in the autoimmune process leading to type 1 (insulin-dependent) diabetes, but the molecular targets of these T cells have not yet been defined. As identification of such antigens may enable measures to be developed to prevent the disease, we have characterized an antigen that is recognized by insulinoma membrane-reactive T-cell clones established from a newly diagnosed type-1 diabetes patient. Subcellular fractionation studies using rat insulinoma indicate that the antigenic determinant recognized by one of these clones is an integral membrane component of the insulin secretory granule. After a 5,000-fold purification, we have defined the antigen as a monomer of relative molecular mass 38,000. As granular membrane proteins are transiently exposed on the cell surface during exocytosis, their accessibility to components of the immune system may be a function of the secretory activity of beta-cells.

Import of several precursor proteins into isolated yeast mitochondria is inhibited by rabbit antiserum raised against the total mitochondrial outer membrane or against electrophoretically purified 45-kd outer membrane proteins. Antisera against other outer membrane proteins are only marginally active or inactive. Inhibition by the antiserum against 45-kd proteins is only weak with untreated mitochondria, but reaches 80-90% with mitochondria that had been pretreated with 0.1 mg/ml trypsin. This trypsin pretreatment by itself inhibits precursor import only slightly (30-50%). Selective inhibition of import does not correlate with binding of the various IgGs to the mitochondrial surface and is also observed with the corresponding Fab fragments. Inhibition by antibodies against 45-kd outer membrane proteins strongly suggests the existence of a mitochondrial surface protein mediating protein import and offers a means of isolating this protein.