Akio Nomoto

Human Cytomegalovirus Glycoprotein UL141 Targets the TRAIL Death Receptors to Thwart Host Innate Antiviral Defenses

Summary Death receptors (DRs) of the TNFR superfamily contribute to antiviral immunity by promoting apoptosis and regulating immune homeostasis during infection, and viral inhibition of DR signaling can alter immune defenses. Here we identify the human cytomegalovirus (HCMV) UL141 glycoprotein as necessary and sufficient to restrict TRAIL DR function. Despite showing no primary sequence homology to TNF family cytokines, UL141 binds the ectodomains of both human TRAIL DRs with affinities comparable to the natural ligand TRAIL. UL141 binding promotes intracellular retention of the DRs, thus protecting virus infected cells from TRAIL and TRAIL-dependent NK cell-mediated killing. The identification of UL141 as a herpesvirus modulator of the TRAIL DRs strongly implicates this pathway as a regulator of host defense to HCMV and highlights UL141 as a pleiotropic inhibitor of NK cell effector function.

Tripropeptin C Blocks the Lipid Cycle of Cell Wall Biosynthesis by Complex Formation with Undecaprenyl Pyrophosphate

ABSTRACT Tripropeptin C (TPPC) is a naturally occurring cyclic lipodepsipeptide antibiotic produced by a Lysobacter sp. TPPC exhibits potent antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant enterococci (VRE), and penicillin-resistant Streptococcus pneumoniae. This antibiotic also inhibits the incorporation of N-acetylglucosamine into the peptidoglycan of S. aureus at a 50% inhibitory concentration (IC50) of 0.7 μM, which is proportional to its MIC (0.87 μM; equivalent to 1.0 μg/ml). Treatment of exponential-phase S. aureus cells with TPPC resulted in accumulation of UDP-MurNAc-pentapeptide in the cytoplasm. The antimicrobial activity of TPPC was weakened by the addition of prenyl pyrophosphates but not by prenyl phosphates, UDP-linked sugars, or the pentapeptide of peptidoglycan. The direct interaction between TPPC and undecaprenyl pyrophosphate (C55-PP) was observed by mass spectrometry and thin-layer chromatography analysis, indicating that TPPC can potentially inhibit C55-PP phosphatase activity, which plays a crucial role in the lipid cycle of peptidoglycan synthesis. As expected, TPPC inhibits this enzymatic reaction at an IC50 of 0.03 to 0.1 μM in vitro, as does bacitracin. From the analysis of accumulation of lipid carrier-related compounds, TPPC was found to cause the accumulation of C55-PP in situ, leading to the accumulation of a glycine-containing lipid intermediate. This suggested that the TPPC/C55-PP complex also inhibits the transglycosylation step or flippase activity, adding to the inhibition of C55-PP dephosphorylation. This mode of action is different from that of currently available drugs such as vancomycin, daptomycin, and bacitracin.

Waldiomycin, a novel WalK-histidine kinase inhibitor from Streptomyces sp. MK844-mF10.

WalK, a histidine kinase, and WalR, a response regulator, make up a two-component signal transduction system that is indispensable for the cell-wall metabolism of low GC Gram-positive bacteria. WalK inhibitors are likely to show bactericidal effects against methicillin-resistant Staphylococcus aureus . We discovered a new WalK inhibitor, designated waldiomycin, by screening metabolites from actinomycetes. Waldiomycin belongs to the family of angucycline antibiotics and is structurally related to dioxamycin. Waldiomycin inhibits WalK from S. aureus and Bacillus subtilis at IC50s 8.8 and 10.2 μM, respectively, and shows antibacterial activity with MICs ranging from 4 to 8 μg ml−1 against methicillin-resistant S. aureus and B. subtilis.

Synthesis of intervenolin, an antitumor natural quinolone with unusual substituents.

Synthesis of intervenolin, a natural quinolone discovered by screening for selective growth inhibitors of cancer cells cocultured with stromal cells over monocultured cells, was achieved. The synthesis utilized a thiocyanate-isothiocyanate rearrangement and Suzuki-Miyaura coupling to furnish the characteristic substituents, the iminodithiocarbonate moiety, and the geranyl side chain, respectively. In vivo studies showed that intervenolin inhibited tumor tissue growth in model mice.

Poliovirus trafficking toward central nervous system via human poliovirus receptor-dependent and -independent pathway

In humans, paralytic poliomyelitis results from the invasion of the central nervous system (CNS) by circulating poliovirus (PV) via the blood–brain barrier (BBB). After the virus enters the CNS, it replicates in neurons, especially in motor neurons, inducing the cell death that causes paralytic poliomyelitis. Along with this route of dissemination, neural pathway has been reported in humans, monkeys, and PV-sensitive human PV receptor (hPVR/CD155)-transgenic (Tg) mice. We demonstrated that a fast retrograde axonal transport process is required for PV dissemination through the sciatic nerve of hPVR-Tg mice and that intramuscularly inoculated PV causes paralysis in a hPVR-dependent manner. We also showed that hPVR-independent axonal transport of PV exists in hPVR-Tg and non-Tg mice, indicating that several different pathways for PV axonal transport exist in these mice. Circulating PV after intravenous inoculation in mice cross the BBB at a high rate in a hPVR-independent manner. We will implicate an involvement of a new possible receptor for PV to permeate the BBB based on our recent findings.

Asteltoxins from the Entomopathogenic Fungus Pochonia bulbillosa 8-H-28

New asteltoxins C (3) and D (4) were found in the extract of the entomopathogenic fungus Pochonia bulbillosa 8-H-28. Compound 2, which was spectroscopically identical with the known asteltoxin B, was isolated, and structural analysis led to a revision of the structure of asteltoxin B. Compounds 2 and 4 have a novel tricyclic ring system connected to a dienyl α-pyrone structure. Compound 3 has a 2,8-dioxabicyclo[3.3.0]octane ring similar to that of asteltoxin (1). Compound 3 showed potent antiproliferative activity against NIAS-SL64 cells derived from the fat body of Spodoptera litura larvae, while 2 and 4 were inactive.

Intervenolin, a new antitumor compound with anti-Helicobacter pylori activity, from Nocardia sp. ML96-86F2.

Because stromal cells can regulate the growth and metastasis of tumor cells, a compound that modulates the interaction between the stromal cells and the tumor cells can control the tumor progression. In the course of our screening for such a compound, we have isolated a new compound, intervenolin, from the culture broth of Nocardia sp. ML96-86F2. Intervenolin inhibits the growth of human gastric and colorectal cancer cell lines in the coculture with the respective organ-derived stromal cells more strongly than that of the cancer cells cultured alone. Intervenolin shows antitumor effect against a xenograft model of human colorectal cancer cells in vivo. Furthermore, intervenolin exerts selective anti-Helicobacter pylori effect.

In vivo imaging of proteasome inhibition using a proteasome-sensitive fluorescent reporter

A proteasome degrades numerous regulatory proteins that are critical for tumor growth and is therefore recognized as a promising anticancer target. Determining proteasome activity in the tumors of mice bearing xenografts is essential for the development of novel proteasome inhibitors. We developed a system for in vivo imaging of proteasome inhibition in the tumors of living mice, using a proteasome‐sensitive fluorescent reporter, ZsProSensor‐1. This reporter consists of a green fluorescent protein, ZsGreen, fused to mouse ornithine decarboxylase, which is degraded by the proteasome without being ubiquitinated. In stably transfected cells expressing ZsProSensor‐1, the fluorescent reporter was rapidly degraded under steady‐state conditions, whereas it was stabilized in the presence of proteasome inhibitors. Subcutaneous inoculation of the transfected cells into nude mice resulted in tumor formation. When the proteasome inhibitor bortezomib was intravenously administered to mice bearing these tumors, the ZsProSensor‐1 protein accumulated in the tumors and emitted a fluorescent signal in a dose‐dependent manner. Robust fluorescence was sustained for 3 days and then gradually decreased to baseline levels within 15 days. Intravenous administration of bortezomib also showed potent antitumor activity. In contrast, oral administration of bortezomib did not result in fluorescent protein accumulation in tumors or exhibit any antitumor activity. These results indicate that in vivo imaging using the ZsProSensor‐1 fluorescent protein can be used as an indicator of antitumor activity and will be a powerful tool for the development of novel proteasome inhibitors.

HSC90 is required for nascent hepatitis C virus core protein stability in yeast cells

Hepatitis C virus core protein (Core) contributes to HCV pathogenicity. Here, we demonstrate that Core impairs growth in budding yeast. We identify HSP90 inhibitors as compounds that reduce intracellular Core protein level and restore yeast growth. Our results suggest that HSC90 (Hsc82) may function in the protection of the nascent Core polypeptide against degradation in yeast and the C‐terminal region of Core corresponding to the organelle‐interaction domain was responsible for Hsc82‐dependent stability. The yeast system may be utilized to select compounds that can direct the C‐terminal region to reduce the stability of Core protein.

Sacchathridine A, a prostaglandin release inhibitor from Saccharothrix sp.

Sacchathridine A (1) was isolated from the fermentation broth of strain Saccharothrix sp. MI559-46F5. The structure was determined as a new naphthoquinone derivative with an acetylhydrazino moiety by a combination of NMR, MS spectral analyses, and chemical degradation. Compound 1 showed inhibitory activity of prostaglandin E2 release in a concentration-dependent manner from human synovial sarcoma cells, SW982, with an IC50 value of 1.0 μM, but had no effect on cell growth up to 30 μM.