Foreword

Art and Design Research for the Future: Innovation and Art & Design ; September 26, 2017Conference: Tsukuba Global Science Week 2017Date: September 25-27, 2017Venue: Tsukuba International Congress CenterSponsored: University of Tsukub

Identification and characterization of host factors involved in transcription and replication of the influenza virus genome

科学研究費助成事業(科学研究費補助金)研究成果報告書：基盤研究(A)2008-2011課題番号：2024902

Silencing of IFN-stimulated gene transcription is regulated by histone H1 and its chaperone TAF-I

Chromatin structure and its alteration play critical roles in the regulation of transcription. However, the transcriptional silencing mechanism with regard to the chromatin structure at an unstimulated state of the interferon (IFN)-stimulated gene (ISG) remains unclear. Here we investigated the role of template activating factor-I (TAF-I, also known as SET) in ISG transcription. Knockdown (KD) of TAF-I increased ISG transcript and simultaneously reduced the histone H1 level on the ISG promoters during the early stages of transcription after IFN stimulation from the unstimulated state. The transcription factor levels on the ISG promoters were increased in TAF-I KD cells only during the early stages of transcription. Furthermore, histone H1 KD also increased ISG transcript. TAF-I and histone H1 double KD did not show the additive effect in ISG transcription, suggesting that TAF-I and histone H1 may act on the same regulatory pathway to control ISG transcription. In addition, TAF-I KD and histone H1 KD affected the chromatin structure near the ISG promoters. On the basis of these findings, we propose that TAF-I and its target histone H1 are key regulators of the chromatin structure at the ISG promoter to maintain the silent state of ISG transcription

Replication-Uncoupled Histone Deposition during Adenovirus DNA Replication

In infected cells, the chromatin structure of the adenovirus genome DNA plays critical roles in its genome functions. Previously, we reported that in early phases of infection, incoming viral DNA is associated with both viral core protein VII and cellular histones. Here we show that in late phases of infection, newly synthesized viral DNA is also associated with histones. We also found that the knockdown of CAF-1, a histone chaperone that functions in the replication-coupled deposition of histones, does not affect the level of histone H3 bound on viral chromatin, although CAF-1 is accumulated at viral DNA replication foci together with PCNA. Chromatin immunoprecipitation assays using epitope-tagged histone H3 demonstrated that histone variant H3.3, which is deposited onto the cellular genome in a replication-independent manner, is selectively associated with both incoming and newly synthesized viral DNAs. Microscopic analyses indicated that histones but not USF1, a transcription factor that regulates viral late gene expression, are excluded from viral DNA replication foci and that this is achieved by the oligomerization of the DNA binding protein (DBP). Taken together, these results suggest that histone deposition onto newly synthesized viral DNA is most likely uncoupled with viral DNA replication, and a possible role of DBP oligomerization in this replication-uncoupled histone deposition is discussed

Foreword

Title: Capacity for Depiction: The Role of Art and Design in Science　「描出力ー科学研究におけるアートおよびデザインの役割」 Conference: Tsukuba Global Science Week 2020 Date: November 21 2020 (Held Online) Sponsored: University of Tsukuba Tsukuba Global Science Week 2020 Art & Design Session Proceedingsarticl

Intrinsically disordered regions of nucleophosmin/B23 regulate its RNA binding activity through their inter- and intra-molecular association

Nucleophosmin (NPM1/B23) is a nucleolar protein implicated in growth-associated functions, in which the RNA binding activity of B23 plays essential roles in ribosome biogenesis. The C-terminal globular domain (CTD) of B23 has been believed to be the RNA binding domain because the splicing variant B23.2 lacking the CTD binds considerably less efficiently to RNA. However, the recognition of target RNAs by B23 remains poorly understood. Herein, we report a novel mechanism by which B23 recognizes specific RNA targets. We observed that the nucleolar retention of B23.3 lacking the basic region of B23.1 was lower than that of B23.1 because of its low RNA binding activity. Circular dichroism measurements indicated that the basic region and adjacent acidic regions of B23 are intrinsically disordered regions (IDRs). Biochemical analyses revealed that the basic IDR alone strongly binds to RNA with low specificity. The excessive RNA binding activity of the basic IDR was restrained by intra-molecular interaction with the acidic IDR of B23. Chemical cross-linking experiments and fluorescent labeling of bipartite tetracysteine-tagged proteins suggested that the inter- and intra-molecular interactions between the two IDRs contribute to the regulation of the RNA binding activity of CTD to control the cellular localization and functions of B23

DNA replication-dependent binding of CTCF plays a critical role in adenovirus genome functions

The expression of adenovirus late genes is shown to require viral DNA replication, but its mechanism remains elusive. Here we found that knockdown of CTCF suppresses viral DNA replication as well as late, but not early, gene expression. Chromatin immunoprecipitation assays indicated that CTCF binds to viral chromatin depending on viral DNA replication. These findings depict CTCF as a critical regulator for adenovirus genome functions in late phases of infection

An influenza virus replicon system in yeast identified Tat-SF1 as a stimulatory host factor for viral RNA synthesis

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YB-1 Functions as a Porter To Lead Influenza Virus Ribonucleoprotein Complexes to Microtubules

De novo-synthesized RNAs are under the regulation of multiple posttranscriptional processes by a variety of RNA-binding proteins. The influenza virus genome consists of single-stranded RNAs and exists as viral ribonucleoprotein (vRNP) complexes. After the replication of vRNP in the nucleus, it is exported to the cytoplasm and then reaches the budding site beneath the cell surface in a process mediated by Rab11a-positive recycling endosomes along microtubules. However, the regulatory mechanisms of the postreplicational processes of vRNP are largely unknown. Here we identified, as a novel vRNP-interacting protein, Y-box-binding protein 1 (YB-1), a cellular protein that is involved in regulation of cellular transcription and translation. YB-1 translocated to the nucleus from the cytoplasm and accumulated in PML nuclear bodies in response to influenza virus infection. vRNP assembled into the exporting complexes with YB-1 at PML nuclear bodies. After nuclear export, using YB-1 knockdown cells and in vitro reconstituted systems, YB-1 was shown to be required for the interaction of vRNP exported from the nucleus with microtubules around the microtubule-organizing center (MTOC), where Rab11a-positive recycling endosomes were located. Further, we also found that YB-1 overexpression stimulates the production of progeny virions in an Rab11a-dependent manner. Taking these findings together, we propose that YB-1 is a porter that leads vRNP to microtubules from the nucleus and puts it into the vesicular trafficking system