Crystal structure of <Emphasis Type="Italic">Escherichia coli</Emphasis> protein ybgI,a toroidal structure with a dinuclear metal site

Background

The protein encoded by the gene ybgI was chosen as a target for a structural genomics project emphasizing the relation of protein structure to function.

Results

The structure of the ybgI protein is a toroid composed of six polypeptide chains forming a trimer of dimers. Each polypeptide chain binds two metal ions on the inside of the toroid.

Conclusion

The toroidal structure is comparable to that of some proteins that are involved in DNA metabolism. The di-nuclear metal site could imply that the specific function of this protein is as a hydrolase-oxidase enzyme.

     

IL-18 induces monocyte chemotactic protein-1 production in macrophages through the phosphatidylinositol 3-kinase/Akt and MEK/ERK1/2 pathways

Macrophages are activated during an inflammatory response and produce multiple inflammatory cytokines. IL-18 is one of the most important innate cytokines produced from macrophages in the early stages of the inflammatory immune response. Monocyte chemoattractant protein (MCP-1) is expressed in many inflammatory diseases such as multiple sclerosis and rheumatoid arthritis, and its expression is correlated with the severity of the disease. Both IL-18 and MCP-1 have been shown to be involved in inflammatory immune responses. However, it has been unclear whether IL-18 is involved in the induction of MCP-1. This investigation was initiated to determine whether IL-18 can induce MCP-1 production, and if so, by which signal transduction pathways. We found that IL-18 induced the production of MCP-1 in macrophages, which was IL-12-independent and was not mediated by autocrine cytokines such as IFN-gamma or TNF-alpha. We then examined signal transduction pathways involved in IL-18-induced MCP-1 production. We found that IL-18 did not activate the IkappaB kinase/NF-kappaB pathway, evidenced by no degradation of IkappaBalpha and no translocation of NF-kappaB p65 to the nucleus in IL-18-stimulated macrophages. Instead, IL-18 activated the PI3K/Akt and MEK/ERK1/2 pathways. Inhibition of either of these pathways attenuated MCP-1 production in macrophages, and inhibition of both signaling pathways resulted in the complete inhibition of MCP-1 production. On the basis of these observations, we conclude that IL-18 induces MCP-1 production through the PI3K/Akt and MEK/ERK1/2 pathways in macrophages.     

Detecting sequence polymorphisms associated with meiotic recombination hotspots in the human genome

Background  

Meiotic recombination events tend to cluster into narrow spans of a few kilobases long, called recombination hotspots. Such hotspots are not conserved between human and chimpanzee and vary between different human ethnic groups. At the same time, recombination hotspots are heritable. Previous studies showed instances where differences in recombination rate could be associated with sequence polymorphisms.     

Regulation of insulin response in skeletal muscle cell by caveolin status

Recent studies on the role of caveolin-1 in adipocytes showed that caveolin has emerged as an important regulatory element in insulin signaling but little is known on its role in skeletal muscle cells. In this study, we demonstrate for the first time that caveolin-1 plays a crucial role in insulin dependent glucose uptake in skeletal muscle cells. Differentiation of L6 skeletal muscle cells induce the expression of caveolin-1 and caveolin-3 with partial colocalization. However in contrast to adipocytes, phosphorylation of insulin receptor beta (IRbeta) and Akt/Erk was not affected by the respective downregulation of caveolin-1 or caveolin-3 in the muscle cells. Moreover, the phosphorylation of IRbeta was detected not only in the caveolae but also in the non-caveolae fractions of the muscle cells despite the interaction of IRbeta with caveolin-1 and caveolin-3. These data implicate the lack of relationship between caveolins and IRbeta pathway in the muscle cells, different from the adipocytes. However, glucose uptake was reduced specifically by downregulation of caveolin-1, but not that of caveolin-3. Taken together, these observations suggest that caveolin-1 plays a crucial role in glucose uptake in differentiated muscle cells and that the regulation of caveolin-1 expression may be an important mechanism for insulin sensitivity, implying the role of muscle cells for type 2 diabetes.     

Role of CTLA-4 in the activation of single- and double-positive thymocytes

CTLA-4, a homologue of CD28, is a negative regulator of T cell activation in the periphery and is transiently expressed on the cell surface after T cell activation. However, the role of CTLA-4 in T cell activation in the thymus is not clear. This investigation was initiated to determine the role of CTLA-4 in the activation of CD4(+)CD8(+) double-positive (DP) and CD4(+)CD8(-) and CD4(-)CD8(+) single-positive (SP) thymocytes using fetal thymic organ cultures (FTOC) of MHC class II-restricted, OVA(323-339)-restricted TCR transgenic mice (DO11.10). We found that treatment of the FTOC with anti-CTLA-4-blocking Ab during activation with OVA(323-339) increased the proportion and number of DP thymocytes, but decreased the proportion and number of SP thymocytes compared with OVA(323-339)-stimulated FTOC without anti-CTLA-4 Ab treatment. In addition, anti-CTLA-4 Ab treatment inhibited OVA(323-339)-induced expression of the early activation marker, CD69, in DP thymocytes, but increased CD69 in SP thymocytes. Similarly, CTLA-4 blockage decreased phosphorylation of ERK in DP thymocytes by Ag-specific TCR engagement, but increased phosphorylation of ERK in SP thymocytes. CTLA-4 blockage inhibited deletion of DP thymocytes treated with a high dose of OVA(323-339), whereas CTLA-4 blockage did not inhibit deletion of DP thymocytes treated with a low dose of OVA(323-339). We conclude that CTLA-4 positively regulates the activation of DP thymocytes, resulting in their deletion, whereas blocking CTLA-4 suppresses the activation of DP thymocytes, leading to inhibition of DP thymocyte deletion. In contrast, CTLA-4 negatively regulates the activation of SP thymocytes.     

Synthesis, characterization and aqueous dispersion of dextran-g-poly(1,4-dioxan-2-one) copolymers

A novel biodegradable graft copolymer, dextran-g-poly(1,4-dioxan-2-one) (PODEX), was synthesized through the ring-opening polymerization (ROP) of 1,4-dioxan-2-one (PDO) onto a dextran backbone. Initially, dextran was silylated with 1,1,1,3,3,3-hexamethyldisilazane. The grafting from pathway was conducted with various (30–70) PDO/OH feed ratios to obtain PODEX copolymers with a various PPDO graft structures. Graft copolymers were characterized by FT-IR, ¹H and ¹³C NMR, DSC, TGA, and WAXD. Molecular weights of the PODEX copolymers (M_W: 94,700–117, 300 Da), glass transition temperature (−29 to −17 °C), melting temperature (82–100 °C), and crystallinity (32–40%) were increased with the content of PPDO. AFM observations revealed that polymeric micelles were spherical and uniform in morphology with around 30–58 nm diameter. Critical micelle concentration (CMC) of self-assembled system was significantly decreased from 3.2 to 1.09 mg/L with the increment of PPDO.