ICDP workshop on the Lake Tanganyika Scientific Drilling Project: a late Miocene–present record of climate, rifting, and ecosystem evolution from the world's oldest tropical lake

The Neogene and Quaternary are characterized by enormous changes in global climate and environments, including global cooling and the establishment of northern high-latitude glaciers. These changes reshaped global ecosystems, including the emergence of tropical dry forests and savannahs that are found in Africa today, which in turn may have influenced the evolution of humans and their ancestors. However, despite decades of research we lack long, continuous, well-resolved records of tropical climate, ecosystem changes, and surface processes necessary to understand their interactions and influences on evolutionary processes. Lake Tanganyika, Africa, contains the most continuous, long continental climate record from the mid-Miocene (∼10 Ma) to the present anywhere in the tropics and has long been recognized as a top-priority site for scientific drilling. The lake is surrounded by the Miombo woodlands, part of the largest dry tropical biome on Earth. Lake Tanganyika also harbors incredibly diverse endemic biota and an entirely unexplored deep microbial biosphere, and it provides textbook examples of rift segmentation, fault behavior, and associated surface processes. To evaluate the interdisciplinary scientific opportunities that an ICDP drilling program at Lake Tanganyika could offer, more than 70 scientists representing 12 countries and a variety of scientific disciplines met in Dar es Salaam, Tanzania, in June 2019. The team developed key research objectives in basin evolution, source-to-sink sedimentology, organismal evolution, geomicrobiology, paleoclimatology, paleolimnology, terrestrial paleoecology, paleoanthropology, and geochronology to be addressed through scientific drilling on Lake Tanganyika. They also identified drilling targets and strategies, logistical challenges, and education and capacity building programs to be carried out through the project. Participants concluded that a drilling program at Lake Tanganyika would produce the first continuous Miocene–present record from the tropics, transforming our understanding of global environmental change, the environmental context of human origins in Africa, and providing a detailed window into the dynamics, tempo and mode of biological diversification and adaptive radiations.© Author(s) 2020. This open access article is distributed under the Creative Commons Attribution 4.0 License

Characterization of Nonstructural Protein Membrane Anchor Deletion Mutants Expressed in the Context of the Hepatitis C Virus Polyprotein

Protein-protein interactions involved in formation of the membrane-associated hepatitis C virus (HCV) replication complex are poorly understood. Here, we investigated nonstructural proteins with deletions in their membrane anchor domains when expressed in the context of the entire HCV polyprotein. Interactions among cytosolic domains of HCV nonstructural proteins were found not to be sufficiently strong to rescue such mutants to the membrane. Thus, the membrane anchor domains of nonstructural proteins are essential for incorporation of these proteins into the HCV replication complex while interactions among the cytosolic domains appear to be relatively weak. This feature may provide the nonstructural proteins with a certain flexibility to perform their multiple functions during HCV replication

Upregulation of Protein Phosphatase 2Ac by Hepatitis C Virus Modulates NS3 Helicase Activity through Inhibition of Protein Arginine Methyltransferase 1

Hepatitis C virus (HCV) is a major cause of chronic liver disease, cirrhosis, and hepatocellular carcinoma worldwide. HCV has a positive-strand RNA genome of about 9.4 kb in size, which serves as a template for replication and for translation of a polyprotein of about 3,000 amino acids. The polyprotein is cleaved co- and posttranslationally by cellular and viral proteases into at least 10 different mature proteins. One of these proteins, nonstructural protein 3 (NS3), has serine protease and NTPase/RNA helicase activity. Arginine 467 in the helicase domain of NS3 (arginine 1493 in the polyprotein) can be methylated by protein arginine methyltransferase 1 (PRMT1). Here we report that the methylation of NS3 inhibits the enzymatic activity of the helicase. Furthermore, we found that PRMT1 activity itself is regulated by protein phosphatase 2A (PP2A). PP2A inhibits PRMT1 enzymatic activity and therefore increases the helicase activity of NS3. This is important, because we found an increased expression of PP2A in cell lines with inducible HCV protein expression, in transgenic mice expressing HCV proteins in hepatocytes, and in liver biopsy samples from patients with chronic hepatitis C. Interestingly, up-regulation of PP2A not only modulates the enzymatic activity of an important viral protein, NS3 helicase, but also interferes with the cellular defense against viruses by inhibiting interferon-induced signaling through signal transducer and activator of transcription 1 (STAT1). We conclude that up-regulation of PP2A might be crucial for the efficient replication of HCV and propose PP2A as a potential target for anti-HCV treatment strategies

Novel Potent Capsid Assembly Modulators Regulate Multiple Steps of the Hepatitis B Virus Life Cycle

The assembly of hepatitis B virus (HBV) core protein (HBc) into capsids represents a critical step of viral replication. HBc has multiple functions during the HBV life cycle, which makes it an attractive target for antiviral therapies

MAPPIT as a high-throughput screening assay for modulators of protein-protein interactions in HIV and HCV

The discovery of novel antivirals for HIV and HCV has been a focus of intensive research for many years. Where the inhibition of critical viral enzymes by small molecules has proven effective for many viruses, there is considerable merit in pursuing protein-protein interactions (PPIs) as targets for therapeutic intervention. The mammalian protein-protein interaction trap (MAPPIT) is a two-hybrid system used for the study of PPIs. The bait and prey proteins are linked to deficient cytokine receptor chimeras, where the bait and prey interaction and subsequent ligand stimulation restores JAK-STAT signaling, resulting in reporter gene expression controlled by a STAT3-responsive promoter. We report the use of MAPPIT as a high-throughput screening assay for the discovery of inhibitors or stimulators of the Vif-APOBEC3G interaction and the reverse transcriptase heterodimerization (RTp66-RTp51) for HIV and the NS4A-NS3 interaction for HCV

Virologic response and characterisation of HCV genotype 2-6 in patients receiving TMC435 monotherapy (study TMC435-C202).

TMC435 is a potent, once-daily, investigational hepatitis C virus (HCV) NS3/4A protease inhibitor in phase III clinical development. In the phase II trial TMC435-C202 (NCT00812331), TMC435 displayed potent activity in genotype 4, 5 and 6 patients and in 3/6 genotype 2 patients, whereas no activity was observed with genotype 3.Journal ArticleResearch Support, Non-U.S. Gov'tSCOPUS: ar.jinfo:eu-repo/semantics/publishe

Molecular elucidation of drug-induced abnormal assemblies of the hepatitis B virus capsid protein by solid-state NMR

Hepatitis B virus (HBV) capsid assembly modulators (CAMs) represent a recent class of anti-HBV antivirals. CAMs disturb proper nucleocapsid assembly, by inducing formation of either aberrant assemblies (CAM-A) or of apparently normal but genome-less empty capsids (CAM-E). Classical structural approaches have revealed the CAM binding sites on the capsid protein (Cp), but conformational information on the CAM-induced off-path aberrant assemblies is lacking. Here we show that solid-state NMR can provide such information, including for wild-type full-length Cp183, and we find that in these assemblies, the asymmetric unit comprises a single Cp molecule rather than the four quasi-equivalent conformers typical for the icosahedral T = 4 symmetry of the normal HBV capsids. Furthermore, while in contrast to truncated Cp149, full-length Cp183 assemblies appear, on the mesoscopic level, unaffected by CAM-A, NMR reveals that on the molecular level, Cp183 assemblies are equally aberrant. Finally, we use a eukaryotic cell-free system to reveal how CAMs modulate capsid-RNA interactions and capsid phosphorylation. Our results establish a structural view on assembly modulation of the HBV capsid, and they provide a rationale for recently observed differences between in-cell versus in vitro capsid assembly modulation.ISSN:2041-172

Discovery of 1‑((2<i>R</i>,4a<i>R</i>,6<i>R</i>,7<i>R</i>,7a<i>R</i>)‑2-Isopropoxy-2-oxidodihydro‑4<i>H</i>,6<i>H</i>‑spiro[furo[3,2‑<i>d</i>][1,3,2]dioxaphosphinine-7,2′-oxetan]-6-yl)pyrimidine-2,4(1<i>H</i>,3<i>H</i>)‑dione (JNJ-54257099), a 3′-5′-Cyclic Phosphate Ester Prodrug of 2′-Deoxy-2′-Spirooxetane Uridine Triphosphate Useful for HCV Inhibition

JNJ-54257099 (<b>9</b>) is a novel cyclic phosphate ester derivative that belongs to the class of 2′-deoxy-2′-spirooxetane uridine nucleotide prodrugs which are known as inhibitors of the HCV NS5B RNA-dependent RNA polymerase (RdRp). In the Huh-7 HCV genotype (GT) 1b replicon-containing cell line <b>9</b> is devoid of any anti-HCV activity, an observation attributable to inefficient prodrug metabolism which was found to be CYP3A4-dependent. In contrast, in vitro incubation of <b>9</b> in primary human hepatocytes as well as pharmacokinetic evaluation thereof in different preclinical species reveals the formation of substantial levels of 2′-deoxy-2′-spirooxetane uridine triphosphate (<b>8</b>), a potent inhibitor of the HCV NS5B polymerase. Overall, it was found that <b>9</b> displays a superior profile compared to its phosphoramidate prodrug analogues (e.g., <b>4</b>) described previously. Of particular interest is the in vivo dose dependent reduction of HCV RNA observed in HCV infected (GT1a and GT3a) human hepatocyte chimeric mice after 7 days of oral administration of <b>9</b>