Comparison of OspA Serotypes for Borrelia burgdorferi Sensu Lato from Japan,Europe and North America

Sixty-one Borrelia burgdorferi sensu lato strains from various sources (ticks, human, and wild animals) in Japan and two strains from ticks in Far Eastern Russia were classified on the basis of reactivity with 16 monoclonal antibodies (mAb) to outer surface protein A (OspA) and by DNA-DNA hybridization assay. Eleven OspA serotypes (J1 to J11) were recognized among the Japanese and the Far East Russian isolates (serotypes J1 to J9 were identified as B. garinii, serotype J10 was identified as B. afzelii, and serotype J11 corresponded to B. japonica), whereas 7 OspA serotypes for North American and European isolates previously reported (Bettina Wilske et al, J. Clin. Microbiol. 31:340-350, 1993) were not observed except for OspA serotype 2 which showed identical reactivity with OspA serotype J10. This finding provides helpful information for understanding the geographical distribution of Lyme disease borrelia and the development of vaccine and diagnostic tests. In conclusion: 1. B. burgdorferi sensu stricto has not been observed in Japan, 2. Japanese B. afzelii isolates are closely related to those from Europe, 3. B. garinii isolates from Japan are highly heterogeneous and apparently different from European B. garinii isolates.     

Oviposition-stimulatory activity of phenanthroindolizidine alkaloids of host-plant origin to a danaid butterfly, Ideopsis similis

Oviposition response of Ideopsis similis (L.) (Lepidoptera: Danaidae) was examined for 12 phenanthroindolizidine alkaloids present in its host plant, Tylophora tanakae (Maxim.) (Asclepiadaceae). At least five alkaloids, i.e. (+)‐isotylocrebrine (3,4,6,7‐tetramethoxyphenanthroindolizidine; l ), (+)‐3‐demethyliso‐ tylocrebrine ( 3 ), (+)‐isotylocrebrine N‐oxide ( 5 ), (+)‐6‐demethyltylocrebrine ( 8 ) and (–)‐7‐demethyltylophorine ( 10 ), were found to individually stimulate oviposition by females. Of these, compounds 1, 3 and 10 were regarded as key components most responsible for host recognition or preference. However, female egg‐laying was much higher in response to a mixture of the five alkaloids. In two‐choice bioassays, more eggs were deposited on samples comprising the five alkaloids than on samples consisting of a single alkaloid. This suggests strongly that host selection by the butterfly is mediated by the synergistic action of several phenanthroindolizidine alkaloids present in the host plant.     

Structure of the Dengue virus helicase/nucleoside triphosphatase catalytic domain at a resolution of 2.4 A

Dengue fever is an important emerging public health concern, with several million viral infections occurring annually, for which no effective therapy currently exists. The NS3 protein from Dengue virus is a multifunctional protein of 69 kDa, endowed with protease, helicase, and nucleoside 5'-triphosphatase (NTPase) activities. Thus, NS3 plays an important role in viral replication and represents a very interesting target for the development of specific antiviral inhibitors. We present the structure of an enzymatically active fragment of the Dengue virus NTPase/helicase catalytic domain to 2.4 A resolution. The structure is composed of three domains, displays an asymmetric distribution of charges on its surface, and contains a tunnel large enough to accommodate single-stranded RNA. Its C-terminal domain adopts a new fold compared to the NS3 helicase of hepatitis C virus, which has interesting implications for the evolution of the Flaviviridae replication complex. A bound sulfate ion reveals residues involved in the metal-dependent NTPase catalytic mechanism. Comparison with the NS3 hepatitis C virus helicase complexed to single-stranded DNA would place the 3' single-stranded tail of a nucleic acid duplex in the tunnel that runs across the basic face of the protein. A possible model for the unwinding mechanism is proposed.     

Hepatic overexpression of a dominant negative form of raptor enhances Akt phosphorylation and restores insulin sensitivity in K/KAy mice

Several serine/threonine kinases reportedly phosphorylate serine residues of IRS-1 and thereby induce insulin resistance. In this study, to investigate the effect of mTOR/raptor on insulin signaling and metabolism in K/KAy mice with genetic obesity-associated insulin resistance, a dominant negative raptor, COOH-terminally deleted raptor (raptor-DeltaC(T)), was overexpressed in the liver via injection of its adenovirus into the circulation. Hepatic raptor-DeltaC(T) expression levels were 1.5- to 4-fold that of endogenously expressed raptor. Glucose tolerance in raptor-DeltaC(T)-overexpressing mice improved significantly compared with that of LacZ-overexpressing mice. Insulin-induced activation of p70S6 kinase (p70(S6k)) was significantly suppressed in the livers of raptor-DeltaC(T) overexpressing mice. In addition, insulin-induced IRS-1, Ser(307), and Ser(636/639) phosphorylations were significantly suppressed in the raptor-DeltaC(T)-overexpressing liver, whereas tyrosine phosphorylation of IRS-1 was increased. PI 3-kinase activation in response to insulin stimulation was increased approximately twofold, and Akt phosphorylation was clearly enhanced under both basal and insulin-stimulated conditions in the livers of raptor-DeltaC(T) mice. Thus, our data indicate that suppression of the mTOR/p70(S6k) pathway leads to improved glucose tolerance in K/KAy mice. These observations may contribute to the development of novel antidiabetic agents.     

Peptidyl-prolyl cis/trans isomerase NIMA-interacting 1 associates with insulin receptor substrate-1 and enhances insulin actions and adipogenesis

Peptidyl-prolyl cis/trans isomerase NIMA-interacting 1 (Pin1) is a unique enzyme that associates with the pSer/Thr-Pro motif and catalyzes cis-trans isomerization. We identified Pin1 in the immunoprecipitates of overexpressed IRS-1 with myc and FLAG tags in mouse livers and confirmed the association between IRS-1 and Pin1 by not only overexpression experiments but also endogenously in the mouse liver. The analysis using deletion- and point-mutated Pin1 and IRS-1 constructs revealed the WW domain located in the N terminus of Pin1 and Ser-434 in the SAIN (Shc and IRS-1 NPXY binding) domain of IRS-1 to be involved in their association. Subsequently, we investigated the role of Pin1 in IRS-1 mediation of insulin signaling. The overexpression of Pin1 in HepG2 cells markedly enhanced insulin-induced IRS-1 phosphorylation and its downstream events: phosphatidylinositol 3-kinase binding with IRS-1 and Akt phosphorylation. In contrast, the treatment of HepG2 cells with Pin1 siRNA or the Pin1 inhibitor Juglone suppressed these events. In good agreement with these in vitro data, Pin1 knock-out mice exhibited impaired insulin signaling with glucose intolerance, whereas adenoviral gene transfer of Pin1 into the ob/ob mouse liver mostly normalized insulin signaling and restored glucose tolerance. In addition, it was also demonstrated that Pin1 plays a critical role in adipose differentiation, making Pin1 knock-out mice resistant to diet-induced obesity. Importantly, Pin1 expression was shown to be up-regulated in accordance with nutrient conditions such as food intake or a high-fat diet. Taken together, these observations indicate that Pin1 binds to IRS-1 and thereby markedly enhances insulin action, essential for adipogenesis.     

Salt-inducible kinase-mediated regulation of steroidogenesis at the early stage of ACTH-stimulation

Salt-inducible kinase (SIK), expressed in Y1 mouse adrenocortical tumor cells at an early stage of adrenocorticotropic hormone (ACTH)-stimulation, represses the cAMP-responsive element (CRE)-dependent gene expression of CYP11A and StAR by acting on bZIP domain of CRE-binding protein. ACTH induced the SIK’s nuclear to cytosolic translocation in a PKA-dependent manner. A mutant SIK in which the PKA-dependently phosphorylatable Ser577 had been replaced with Ala could not move out of the nucleus. The degree of CRE-reporter repression by SIK was strong as long as SIK was present in the nucleus. These indicated that intracellular translocation of SIK might be an important factor to determine the time-dependent change in the level of steroidogenic gene expression in ACTH-stimulated cells. Promoter analyses suggested that SIK repressed gene expressions not only of CYP11A and StAR but also of CYP11B1, CYP11B2 and SIK itself. We propose here that SIK is one of important molecule regulating expression of steroidogenic genes in the early phase of ACTH treatment.