Biology subjects

Morikawa, T.

Publications and source records attributed to Morikawa, T..

3 recordsLinked to original sources

bioRxiv2021

Influenza virus ribonucleoprotein complex formation occurs in the nucleolus

Influenza A virus double-helical ribonucleoprotein complex (vRNP) performs transcription and replication of viral genomic RNA (vRNA). Unlike most RNA viruses, vRNP formation accompanied by vRNA replication is carried out in the nucleus of virus-infected cell. However, the precise subnuclear site remains unknown. Here, we report the subnuclear site of vRNP formation in influenza virus. We found that all vRNP components were colocalized in the nucleolus of virus-infected cells at early stage of infection. Mutational analysis showed that nucleolar localization of viral nucleoprotein, a major vRNP component, is critical for functional double-helical vRNP formation. Furthermore, nucleolar disruption of virus-infected cells inhibited vRNP component assembly into double-helical vRNPs, resulting in decreased vRNA transcription and replication. Collectively, our findings demonstrate that the vRNA replication-coupled vRNP formation occurs in the nucleolus, demonstrating the importance of the nucleolus for influenza virus life cycle.

microbiology↗

bioRxiv2020

vRNA-vRNA interactions in influenza A virus HA vRNA packaging

The genome of the influenza A virus is composed of eight single-stranded negative-sense RNA segments (vRNAs). The eight different vRNAs are selectively packaged into progeny virions. This process likely involves specific interactions among vRNAs via segment-specific packaging signals located in the 3 and 5 terminal coding regions of vRNAs. To identify vRNA(s) that interact with hemagglutinin (HA) vRNA during genome packaging, we generated a mutant virus, HA 5m2, which possessed five silent mutations in the 5 packaging signal region of HA vRNA. The HA 5m2 virus had a specific defect in HA vRNA incorporation, which reduced the viral replication efficiency. After serial passaging in cells, the virus acquired additional mutations in the 5 terminal packaging signal regions of both HA and PB2 vRNAs. These mutations contributed to recovery of viral growth and packaging efficiency of HA vRNA. A direct RNA-RNA interaction between the 5 ends of HA and PB2 vRNAs was confirmed in vitro. Our results indicate that direct interactions of HA vRNA with PB2 vRNA via their packaging signal regions are important for selective genome packaging and enhance our knowledge on the emergence of pandemic influenza viruses through genetic reassortment.

microbiology↗

bioRxiv2018

Environmental Optimization Enables Maintenance of Quiescent Hematopoietic Stem Cells Ex Vivo

Hematopoietic stem cells (HSCs) maintain lifelong hematopoiesis by remaining quiescent in the bone marrow niche. Recapitulation of a quiescent state in culture has not been achieved, as cells rapidly proliferate and differentiate in vitro. After exhaustive analysis of different environmental factor combinations and concentrations as a way to mimic physiological conditions, we were able to maintain engraftable quiescent HSCs for 1 month in culture under very low cytokine concentrations, hypoxia, and very high fatty acid levels. Exogenous fatty acids were required likely due to suppression of intrinsic fatty acid synthesis by hypoxia and low cytokine conditions. By contrast, high cytokine concentrations or normoxia induced HSC proliferation and differentiation. Our novel culture system provides a means to evaluate properties of steady state HSCs and test effects of defined factors in vitro under near-physiological conditions.

cell biology↗