Interactions between p27 and p88 replicase proteins of Red clover necrotic mosaic virus play an essential role in viral RNA replication and suppression of RNA silencing via the 480-kDa viral replicase complex assembly

AbstractRed clover necrotic mosaic virus (RCNMV), a positive-sense RNA virus with a bipartite genome, encodes p27 and p88 replicase proteins that are required for viral RNA replication and suppression of RNA silencing. In this study, we indentified domains in p27 and p88 responsible for their protein–protein interactions using in vitro pull-down assays with the purified recombinant proteins. Coimmunoprecipitation analysis in combination with blue-native polyacrylamide gel electrophoresis using mutated p27 proteins showed that both p27–p27 and p27–p88 interactions are essential for the formation of the 480-kDa complex, which has RCNMV-specific RNA-dependent RNA polymerase activity. Furthermore, we found a good correlation between the accumulated levels of the 480-kDa complex and replication levels and the suppression of RNA silencing activity. Our results indicate that interactions between RCNMV replicase proteins play an essential role in viral RNA replication and in suppressing RNA silencing via the 480-kDa replicase complex assembly

Roles of Phosphatidic Acid in Virus RNA Replication

Eukaryotic positive-strand RNA [(+)RNA] viruses are intracellular obligate parasites replicate using the membrane-bound replicase complexes that contain multiple viral and host components. To replicate, (+)RNA viruses exploit host resources and modify host metabolism and membrane organization. Phospholipase D (PLD) is a phosphatidylcholine- and phosphatidylethanolamine-hydrolyzing enzyme that catalyzes the production of phosphatidic acid (PA), a lipid second messenger that modulates diverse intracellular signaling in various organisms. PA is normally present in small amounts (less than 1% of total phospholipids), but rapidly and transiently accumulates in lipid bilayers in response to different environmental cues such as biotic and abiotic stresses in plants. However, the precise functions of PLD and PA remain unknown. Here, we report the roles of PLD and PA in genomic RNA replication of a plant (+)RNA virus, Red clover necrotic mosaic virus (RCNMV). We found that RCNMV RNA replication complexes formed in Nicotiana benthamiana contained PLDα and PLDβ. Gene-silencing and pharmacological inhibition approaches showed that PLDs and PLDs-derived PA are required for viral RNA replication. Consistent with this, exogenous application of PA enhanced viral RNA replication in plant cells and plant-derived cell-free extracts. We also found that a viral auxiliary replication protein bound to PA in vitro, and that the amount of PA increased in RCNMV-infected plant leaves. Together, our findings suggest that RCNMV hijacks host PA-producing enzymes to replicate

Femtoscopic study of coupled-channel baryon-baryon interactions with S=−2S=-2

Particles and Nuclei International Conference - PANIC2021, 5 - 10 September, 2021, OnlineThe correlation functions of pΞ− and ΛΛ pairs from pp and pA collisions are studied in the coupled-channel framework using the NΞ-ΛΛ coupled-channel baryon-baryon potentials obtained in the lattice QCD calculation at almost physical quark masses. The pΞ− correlation function is calculated to be significantly enhanced from the pure Coulomb case, while the ΛΛ correlation function is slightly enhanced from that of the pure fermion quantum statistics. These features reflect the large and small scattering lengths in the pΞ− and ΛΛ channels in magnitude, and agree with the observed data by the ALICE collaboration. The agreement confirms the S = −2 baryon-baryon potentials from lattice QCD

GAPDH-A Facilitates Intercellular Movement of RCNMV

The formation of virus movement protein (MP)-containing punctate structures on the cortical endoplasmic reticulum is required for efficient intercellular movement of Red clover necrotic mosaic virus (RCNMV), a bipartite positive-strand RNA plant virus. We found that these cortical punctate structures constitute a viral replication complex (VRC) in addition to the previously reported aggregate structures that formed adjacent to the nucleus. We identified host proteins that interacted with RCNMV MP in virus-infected Nicotiana benthamiana leaves using a tandem affinity purification method followed by mass spectrometry. One of these host proteins was glyceraldehyde 3-phosphate dehydrogenase-A (NbGAPDH-A), which is a component of the Calvin-Benson cycle in chloroplasts. Virus-induced gene silencing of NbGAPDH-A reduced RCNMV multiplication in the inoculated leaves, but not in the single cells, thereby suggesting that GAPDH-A plays a positive role in cell-to-cell movement of RCNMV. The fusion protein of NbGAPDH-A and green fluorescent protein localized exclusively to the chloroplasts. In the presence of RCNMV RNA1, however, the protein localized to the cortical VRC as well as the chloroplasts. Bimolecular fluorescence complementation assay and GST pulldown assay confirmed in vivo and in vitro interactions, respectively, between the MP and NbGAPDH-A. Furthermore, gene silencing of NbGAPDH-A inhibited MP localization to the cortical VRC. We discuss the possible roles of NbGAPDH-A in the RCNMV movement process

Identification of a Novel Quinvirus in the Family Betaflexiviridae That Infects Winter Wheat

Yellow mosaic disease in winter wheat is usually attributed to the infection by bymoviruses or furoviruses; however, there is still limited information on whether other viral agents are also associated with this disease. To investigate the wheat viromes associated with yellow mosaic disease, we carried out de novo RNA sequencing (RNA-seq) analyses of symptomatic and asymptomatic wheat-leaf samples obtained from a field in Hokkaido, Japan, in 2018 and 2019. The analyses revealed the infection by a novel betaflexivirus, which tentatively named wheat virus Q (WVQ), together with wheat yellow mosaic virus (WYMV, a bymovirus) and northern cereal mosaic virus (a cytorhabdovirus). Basic local alignment search tool (BLAST) analyses showed that the WVQ strains (of which there are at least three) were related to the members of the genus Foveavirus in the subfamily Quinvirinae (family Betaflexiviridae). In the phylogenetic tree, they form a clade distant from that of the foveaviruses, suggesting that WVQ is a member of a novel genus in the Quinvirinae. Laboratory tests confirmed that WVQ, like WYMV, is potentially transmitted through the soil to wheat plants. WVQ was also found to infect rye plants grown in the same field. Moreover, WVQ-derived small interfering RNAs accumulated in the infected wheat plants, indicating that WVQ infection induces antiviral RNA silencing responses. Given its common coexistence with WYMV, the impact of WVQ infection on yellow mosaic disease in the field warrants detailed investigation

Development of an X-ray HARP–FEA detector system for high-throughput protein crystallography

A new detector system for protein crystallography based on an X-ray HARP–FEA is presented

Usefulness of the Palliative Prognostic Index in patients with lung cancer.

The usefulness of the Palliative Prognostic Index (PPI) has been successfully validated in a variety of clinical settings. However, while lung cancer is the leading cause of death worldwide, patients with lung cancer accounted for only 6.9-25.8 % of the study populations in these previous studies. We conducted a retrospective study to evaluate the usefulness of the PPI for survival prediction in patients with lung cancer. Patients with lung cancer who were admitted to our hospital between 2009 and 2013 to receive palliative care were enrolled. The association between the Palliative Prognostic Index, determined based on the data recorded in the clinical charts at the last admission to our hospital, and survival was evaluated. The patient group with a PPI of >6 showed a significantly shorter survival time than the patient group with a PPI of ≤ 6 (P < 0.0001, log-rank test). The sensitivity and specificity of the PPI determined using the cutoff value of 6 for predicting less than 3 weeks of survival were 61.3 and 86.8 %, respectively. However, the sensitivity decreased to 50.0 % when the assessment was carried out in only patients with small cell lung carcinoma. Our findings suggest the existence of a close association between the PPI and survival in patients with lung cancer receiving palliative care. However, the sensitivity of the index for predicting less than 3 weeks of survival was relatively low in patients with small cell lung carcinoma

Three dimensional analysis of air hydrate in ice core sample by using phase-contrast X-ray imaging techniques

第2回極域科学シンポジウム　氷床コアセッション 11月16日（水） 国立極地研究所 2階大会議室前フロ