A novel role for reciprocal CD30-CD30L signaling in the cross-talk between natural killer and dendritic cells

The interplay between dendritic cells (DCs) and natural killer (NK) cells directs adaptive immune responses. The molecular basis of the cross-talk is largely undefined. Here, we provide evidence for a contribution of CD30 (TNFRSF8) and its ligand CD30L (TNFSF8) expressed on NK cells and DCs, respectively. We demonstrate that CD30-mediated engagement of CD30L induced cytokine secretion from immature DCs via the mitogen-activated protein kinase pathway. Moreover, CD30L engagement promoted differentiation to mature DCs. On the contrary, the engagement of CD30 on NK cells resulted in an NF-κB-dependent release of TNF-α/IFN-γ. These data uncover a novel and unexpected role for CD30/CD30L that contributes to proinflammatory immune responses.     

Synthesis of (3,4-dimethoxyphenoxy)alkylamino acetamides as orexin-2 receptor antagonists

The discovery and synthesis of a series of (dimethoxyphenoxy)alkylamino acetamides as orexin-2 receptor antagonists from a small-molecule combinatorial library using a high-throughput calcium mobilization functional assay (HEK293-human OX2-R cell line) is described. Active compounds show a good correlation between high-throughput single concentration screening data and measured IC(50)s. Specific examples exhibit IC(50) values of approximately 20 nM using human orexin A as the peptide agonist for the orexin-2 receptor.     

Benzo[a]pyrene diol epoxide-deoxyguanosine adducts are accurately bypassed by yeast DNA polymerase zeta in vitro

The possible role of bypass DNA polymerase zeta in mutagenic translesion synthesis past benzo[a]pyrene (BP) 7,8-diol-9,10-epoxide (DE) N(2)-deoxyguanosine (dG) adducts has been examined. We prepared 59-mer DNA templates containing dG adducts derived from trans opening of enantiomers of BP DE-2, in which the 7-hydroxyl group and epoxide oxygen are trans. The 10S-BP DE-dG and 10R-BP DE-dG adducts derive from the (+)- and (-)-DE-2 enantiomers, respectively. The adducted dG is located at a site identified as a G-->T mutational hotspot in random mutagenesis studies of (+)-BP DE-2 in Chinese hamster V-79 cells. Yeast pol zeta (complex of Gst-Rev3p and Rev7p) formed extension products (total of all lengths) of 71, 74 and 88% of a primer annealed to the 10S-BP DE-dG, 10R-BP DE-dG and non-adducted 59-mer templates, respectively. However, only 18 and 19% of the primer was extended to the full-length product on 10S-BP DE-dG and 10R-BP DE-dG adducted templates compared to 55% of the primer on the non-adducted template. A major 34-mer product corresponding to primer elongation up to and including the base before the adduct indicated that nucleotide incorporation opposite both adducts was strongly blocked. Full-length products were isolated from gels and subjected to PCR amplification and cloning. Sequence analysis of more than 300 clones of these full-length products on each template showed that only the correct dCMP was incorporated opposite both the adducted and non-adducted G-hotspot in the template. This corresponds to a probability of mutation lower than 0.3%, the limit of detection, and demonstrates the remarkable fidelity of yeast pol zeta in translesion synthesis past these BP DB-dG lesions in vitro.     

Dynamin interacts with members of the sumoylation machinery

Dynamin is a GTP-binding protein whose oligomerization-dependent assembly around the necks of lipid vesicles mediates their scission from parent membranes. Dynamin is thus directly involved in the regulation of endocytosis. Sumoylation is a post-translational protein modification whereby the ubiquitin-like modifier Sumo is covalently attached to lysine residues on target proteins by a process requiring the concerted action of an activating enzyme (ubiquitin-activating enzyme), a conjugating enzyme (ubiquitin carrier protein), and a ligating enzyme (ubiquitin-protein isopeptide ligase). Here, we show that dynamin interacts with Sumo-1, Ubc9, and PIAS-1, all of which are members of the sumoylation machinery. Ubc9 and PIAS-1 are known ubiquitin carrier protein and ubiquitin-protein isopeptide ligase enzymes, respectively, for the process of sumoylation. We have identified the coiled-coil GTPase effector domain (GED) of dynamin as the site on dynamin that interacts with Sumo-1, Ubc9, and PIAS-1. Although we saw no evidence of covalent Sumo-1 attachment to dynamin, Sumo-1 and Ubc9 are shown here to inhibit the lipid-dependent oligomerization of dynamin. Expression of Sumo-1 and Ubc9 in mammalian cells down-regulated the dynamin-mediated endocytosis of transferrin, whereas dynamin-independent fluid-phase uptake was not affected. Furthermore, using high resolution NMR spectroscopy, we have identified amino acid residues on Sumo-1 that directly interact with the GED of dynamin. The results suggest that the GED of dynamin may serve as a scaffold that concentrates the sumoylation machinery in the vicinity of potential acceptor proteins.     

Reconstructing Indian-Australian phylogenetic link

Background  

An early dispersal of biologically and behaviorally modern humans from their African origins to Australia, by at least 45 thousand years via southern Asia has been suggested by studies based on morphology, archaeology and genetics. However, mtDNA lineages sampled so far from south Asia, eastern Asia and Australasia show non-overlapping distributions of haplogroups within pan Eurasian M and N macrohaplogroups. Likewise, support from the archaeology is still ambiguous.     

Dendritic cells release HLA-B-associated transcript-3 positive exosomes to regulate natural killer function

NKp30, a natural cytotoxicity receptor expressed on NK cells is critically involved in direct cytotoxicity against various tumor cells and directs both maturation and selective killing of dendritic cells. Recently the intracellular protein BAT3, which is involved in DNA damage induced apoptosis, was identified as a ligand for NKp30. However, the mechanisms underlying the exposure of the intracellular ligand BAT3 to surface NKp30 and its role in NK-DC cross talk remained elusive. Electron microscopy and flow cytometry demonstrate that exosomes released from 293T cells and iDCs express BAT3 on the surface and are recognized by NKp30-Ig. Overexpression and depletion of BAT3 in 293T cells directly correlates with the exosomal expression level and the activation of NK cell-mediated cytokine release. Furthermore, the NKp30-mediated NK/DC cross talk resulting either in iDC killing or maturation was BAT3-dependent. Taken together this puts forward a new model for the activation of NK cells through intracellular signals that are released via exosomes from accessory cells. The manipulation of the exosomal regulation may offer a novel strategy to induce tumor immunity or inhibit autoimmune diseases caused by NK cell-activation.