The human homologue of Caenorhabditis elegans CED-6 specifically promotes phagocytosis of apoptotic cells

AbstractA key feature of the process of programmed cell death (apoptosis) is the efficiency with which the dying cells are recognized and engulfed by phagocytes [1]. Apoptotic cells are rapidly cleared either by neighbouring cells acting as semi-professional phagocytes or by experts of the macrophage line, so that an inflammatory response is avoided [2]. The Caenorhabditis elegans gene ced-6 is required for efficient engulfment of apoptotic cells [3] and is one of a group of genes that define two partially redundant parallel pathways for the engulfment process [4,5]. These pathways may be conserved across evolution, as two other engulfment genes have human homologues. A CED-5 homologue is part of a human CrkII–DOCK180–Rac signaling pathway proposed to mediate cytoskeletal reorganization [6–8] and a CED-7 homologue is similar to the ABC transporters [9,10]. Here, we report the cloning and characterization of human CED-6, a human homologue of C. elegans CED-6. The 34 kDa hCED-6 protein is expressed in most tissues, some human cancer cells, and in primary human macrophages. We developed an assay that quantitates the phagocytic activity of mammalian macrophages: the number of apoptotic cells that have been internalized is measured by the uptake of lacZ-positive apoptotic cells by adherent transgenic macrophages. The results of this assay demonstrate that overexpression of hCED-6 promotes phagocytosis only of apoptotic cells and suggest that hCED-6 is the mammalian orthologue of C. elegans CED-6 and is a part of a highly conserved pathway that specifically mediates the phagocytosis of apoptotic cells

Uptake and degradation of dsRNA in the gut lumen of insects in relation to environmental RNAi efficiency

RNA interference is a highly conserved post transcriptional gene silencing mechanism that is triggered by dsRNA molecules when they are present in the cell. It is obvious that the RNAi technology, by downregulating essential genes, thereby causing mortality, gained significant interest in pest management research over the past years. Several studies have demonstrated that this technique shows great potential for the development of novel strategies for selectively controlling agricultural pests. Nevertheless, several important issues remain to be solved in order to optimize the use of RNAi in insect pest control and to extend its use. The degree of sensitivity towards RNAi varies strongly, with many economically important species being refractory to environmentally delivered dsRNA (feeding and/or injection). For oral delivery of dsRNA, the insect midgut forms the first barrier. dsRNase activity has been demonstrated in gut juice of several insects, suggesting that dsRNA degradation in the alimentary tract could contribute to lower RNAi efficiencies. In addition, the cellular uptake of dsRNA from the gut lumen could also constitute an important rate-limiting step for environmental RNAi. However, most studies regarding the mechanisms of dsRNA uptake are not focusing on uptake mechanisms directly from the gut lumen, but rather from the body cavity after injection. Therefore, the general aim of this research was to study the environmental RNAi response with a focus on the oral uptake route in the desert locust Schistocerca gregaria, and the Colorado potato beetle, Leptinotarsa decemlineata. A definitive role for dsRNA degradation in the RNAi response was shown.status: publishe

Characterization of the murine IL-5 receptor complex with the use of a panel of monoclonal antibodies: relationship to the murine IL-3 receptor

To obtain mAb against the murine IL-5R (mIL-5R), Wistar rats were immunized with B13 cells, a murine Ly-1+ (CD5+) pre-B cell line which is dependent on IL-3 or IL-5 for its growth. A first group of six mAb could immunoprecipitate, from detergent-lysed B13 cells, a 60-kDa polypeptide (p60) corresponding to the recently cloned mIL-5R alpha-chain. A second group of three mAb was able to immunoprecipitate a protein doublet of 130 to 140 kDa (p130 and p140) corresponding to the previously characterized mIL-3R and mIL-3R-like polypeptide, respectively. One mAb (25C9) specifically bound the p130 polypeptide only. Here we show that: 1) mAb directed against the mIL-5R p60 component completely block IL-5 binding; 2) mAb recognizing the p130-p140 doublet interfere with both IL-3 and IL-5 binding; 3) mAb recognizing p130-p140 block the high affinity binding of IL-5 and hence the high affinity mIL-5R consists of the association of the p60 and p130 and/or p140 component; 4) one particular mAb, 25C9, which binds only to the p130 polypeptide, interferes with only IL-3 binding, and has no effect on the binding of IL-5. These results on binding were corroborated by a biologic assay based on the cytokine-dependent proliferation of B13 cells. The results presented here support a model for the mIL-5R consisting of the alpha-chain (p60) associated with the p140 (IL-3R-like), whereas the p130 (IL-3R) is not involved in the IL-5R complex