Respiratory epithelial cell death by influenza virus infection is responsible for the induction of inflammatory responses, but the exact cell death mechanism is not understood. Here we showed that ...influenza virus infection induces apoptosis and pyroptosis in normal or precancerous human bronchial epithelial cells. Apoptosis was induced only in malignant tumor cells infected with influenza virus. In human precancerous respiratory epithelial cells (PL16T), the number of apoptotic cells increased at early phases of infection, but pyroptotic cells were observed at late phases of infection. These findings suggest that apoptosis is induced at early phases of infection but the cell death pathway is shifted to pyroptosis at late phases of infection. We also found that the type I interferon (IFN)-mediated JAK-STAT signaling pathway promotes the switch from apoptosis to pyroptosis by inhibiting apoptosis possibly through the induced expression of the
anti-apoptotic gene. Further, the inhibition of JAK-STAT signaling repressed pyroptosis but enhanced apoptosis in infected PL16T cells. Collectively, we propose that type I IFN signaling pathway triggers pyroptosis but not apoptosis in the respiratory epithelial cells in a mutually exclusive manner to initiate proinflammatory responses against influenza virus infection.
Respiratory epithelium functions as a sensor of infectious agents to initiate inflammatory responses along with cell death. However, the exact cell death mechanism responsible for inflammatory responses by influenza virus infection is still unclear. We showed that influenza virus infection induced apoptosis and pyroptosis in normal or precancerous human bronchial epithelial cells. Apoptosis was induced at early phases of infection, but the cell death pathway was shifted to pyroptosis at late phases of infection under the regulation of type I IFN signaling to promote proinflammatory cytokine production. Taken together, our results indicate that the type I IFN signaling pathway plays an important role to induce pyroptosis but represses apoptosis in the respiratory epithelial cells to initiate proinflammatory responses against influenza virus infection.
Abstract
Telomere, the terminus of linear chromosome in eukaryotes, is composed of specific repeat DNA which is mainly synthesized by a protein complex called telomerase. The maintenance of telomere ...DNA is important for unlimited proliferative capacity of cancer cells. The telomerase activity is controlled by the expression level of
telomerase reverse transcriptase
(
TERT
), a catalytic unit of telomerase, in some species including human. Therefore, to reveal the regulatory mechanisms of the transcription of
TERT
gene is important for understanding the tumor development. We found that template activating factor-I (TAF-I), a multifunctional nuclear protein, is involved in the transcriptional activation of
TERT
for the maintenance of telomere DNA in HeLa cells. TAF-I maintains the histone H3 modifications involved in transcriptional activation and hypomethylated cytosines in CpG dinucleotides around the transcription start site (TSS) in the
TERT
gene locus. Collectively, TAF-I is involved in the maintenance of telomere DNA through the regulation of
TERT
transcription, then consequently the occurrence and/or recurrence of cancer cells.
Although thousands of long noncoding RNAs (lncRNAs) are localized in the nucleus, only a few dozen have been functionally characterized. Here we show that nuclear enriched abundant transcript 1 ...(NEAT1), an essential lncRNA for the formation of nuclear body paraspeckles, is induced by influenza virus and herpes simplex virus infection as well as by Toll-like receptor3-p38 pathway-triggered poly I:C stimulation, resulting in excess formation of paraspeckles. We found that NEAT1 facilitates the expression of antiviral genes including cytokines such as interleukin-8 (IL8). We found that splicing factor proline/glutamine-rich (SFPQ), a NEAT1-binding paraspeckle protein, is a repressor of IL8 transcription, and that NEAT1 induction relocates SFPQ from the IL8 promoter to the paraspeckles, leading to transcriptional activation of IL8. Together, our data show that NEAT1 plays an important role in the innate immune response through the transcriptional regulation of antiviral genes by the stimulus-responsive cooperative action of NEAT1 and SFPQ.
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•Poly I:C stimulation and virus infection induce NEAT1v2 expression•Poly I:C stimulation and virus infection induces excess formation of paraspeckles•NEAT1 facilitates the expression of antiviral genes including cytokines such as IL8•NEAT1 induction relocates SFPQ from the IL8 promoter to the paraspeckles
Imamura et al. show that NEAT1, a nuclear long noncoding RNA, can cooperatively function with a paraspeckle protein, SFPQ (splicing factor proline/glutamine-rich), to regulate antiviral gene expression.
Replication of influenza viral genomic RNA (vRNA) is catalyzed by viral RNA-dependent RNA polymerase (vRdRP). Complementary RNA (cRNA) is first copied from vRNA, and progeny vRNAs are then amplified ...from the cRNA. Although vRdRP and viral RNA are minimal requirements, efficient cell-free replication could not be reproduced using only these viral factors. Using a biochemical complementation assay system, we found a novel activity in the nuclear extracts of uninfected cells, designated IREF-2, that allows robust unprimed vRNA synthesis from a cRNA template. IREF-2 was shown to consist of host-derived proteins, pp32 and APRIL. IREF-2 interacts with a free form of vRdRP and preferentially upregulates vRNA synthesis rather than cRNA synthesis. Knockdown experiments indicated that IREF-2 is involved in in vivo viral replication. On the basis of these results and those of previous studies, a plausible role(s) for IREF-2 during the initiation processes of vRNA replication is discussed.
Retinoic acid inducible gene I (RIG-I)-like receptors (RLRs) function as cytoplasmic sensors for viral RNA to initiate antiviral responses including type I interferon (IFN) production. It has been ...unclear how RIG-I encounters and senses viral RNA. To address this issue, we examined intracellular localization of RIG-I in response to viral infection using newly generated anti-RIG-I antibody. Immunohistochemical analysis revealed that RLRs localized in virus-induced granules containing stress granule (SG) markers together with viral RNA and antiviral proteins. Because of similarity in morphology and components, we termed these aggregates antiviral stress granules (avSGs). Influenza A virus (IAV) deficient in non-structural protein 1 (NS1) efficiently generated avSGs as well as IFN, however IAV encoding NS1 produced little. Inhibition of avSGs formation by removal of either the SG component or double-stranded RNA (dsRNA)-dependent protein kinase (PKR) resulted in diminished IFN production and concomitant enhancement of viral replication. Furthermore, we observed that transfection of dsRNA resulted in IFN production in an avSGs-dependent manner. These results strongly suggest that the avSG is the locus for non-self RNA sensing and the orchestration of multiple proteins is critical in the triggering of antiviral responses.
SET‐Nup214 is a recurrent fusion gene that is mainly observed in T‐cell acute lymphoblastic leukemia (T‐ALL). Dysregulation of homeobox (Hox) genes is frequently observed in patients with leukemia. ...Consistent with this, HoxA genes are upregulated in the SET‐Nup214 + T‐ALL cell line and patients. Although SET‐Nup214 has been reported to be recruited to the promoter regions of HoxA genes, the detailed mechanisms of how SET‐Nup214 specifically binds to HoxA gene promoters and regulates HoxA gene expression are not known. In this study, we demonstrated that SET‐Nup214 interacts with MLL via the SET acidic region of SET‐Nup214. SET‐Nup214 and MLL cooperatively enhance the promoter activity of the HoxA10 gene. Neither the SET region alone nor the Nup214 region alone sufficiently enhanced the HoxA10 gene promoter. Our results indicated that the SET portion of the SET‐Nup214‐fusion protein is important for interactions with MLL and transcription enhancement of the HoxA10 gene. Thus, our study will contribute to the understanding of how SET‐Nup214 and MLL disturb the expression of HoxA10 gene in leukemia.
SET‐Nup214 fusion protein, mainly observed in T‐cell acute lymphoblastic leukemia, interacts with MLL. SET‐Nup214 and MLL cooperatively enhance the promoter activity of the HoxA10 gene.
Upstream binding factor (UBF) is a member of the high-mobility group (HMG) box protein family, characterized by multiple HMG boxes and a C-terminal acidic region (AR). UBF is an essential ...transcription factor for rRNA genes and mediates the formation of transcriptionally active chromatin in the nucleolus. However, it remains unknown how UBF is specifically localized to the nucleolus. Here, we examined the molecular mechanisms that localize UBF to the nucleolus. We found that the first HMG box (HMG box 1), the linker region (LR), and the AR cooperatively regulate the nucleolar localization of UBF1. We demonstrated that the AR intramolecularly associates with and attenuates the DNA binding activity of HMG boxes and confers the structured DNA preference to HMG box 1. In contrast, the LR was found to serve as a nuclear localization signal and compete with HMG boxes to bind the AR, permitting nucleolar localization of UBF1. The LR sequence binds DNA and assists the stable chromatin binding of UBF. We also showed that the phosphorylation status of the AR does not clearly affect the localization of UBF1. Our results strongly suggest that associations of the AR with HMG boxes and the LR regulate UBF nucleolar localization.
The adenovirus (Ad) genome is believed to be packaged into the virion by forming a chromatin‐like structure. The replicated viral genome is likely to be condensed through binding with viral core ...proteins before encapsidation. Replicated viral genomes accumulate in the central region of the nucleus, which we termed virus‐induced postreplication (ViPR) body. However, the molecular mechanism by which the nuclear structure is reorganized and its functional significance in virus production are currently not understood. In this study, we found that viral packaging protein IVa2, but not capsid proteins, accumulated in the ViPR body. In addition, nucleolar chromatin regulatory proteins, nucleophosmin 1 (NPM1), upstream binding factor, and nucleolin accumulated in the ViPR body in late‐stage Ad infection. NPM1 depletion increased the nuclease‐resistant viral genome and delayed the ViPR body formation. These results suggested that structural changes in the infected cell nucleus depend on the formation of viral chromatin by host chromatin regulatory proteins. Because NPM1 depletion decreases production of the infectious virion, we propose that host factor‐mediated viral chromatin remodeling and concomitant ViPR body formation are prerequisites for efficient encapsidation of Ad chromatin.
During late stages of Adenovirus infection, replicated viral DNA accumulates in virus‐induced postreplication (ViPR) bodies in which various host and viral proteins also accumulate. In ViPR bodies, replicated viral DNAs may be remodeled by host chromatin regulatory proteins including nucleophosmin 1. This viral genome remodeling step is likely to be a prerequisite for efficient viral genome encapsidation.
By dissecting and reconstituting a cell‐free influenza virus genome replication system, we have purified and identified the minichromosome maintenance (MCM) complex, which is thought to be a DNA ...replicative helicase, as one of the host factors that regulate the virus genome replication. MCM interacted with the PA subunit of the viral RNA‐dependent RNA polymerase that is found to be involved in the replication genetically. The virus genome replication was decreased in MCM2 knockdown cells. The viral polymerase appeared to be a nonproductive complex, that is, it was capable of initiating replication but produced only abortive short RNA chains. MCM stimulated de novo‐initiated replication reaction by stabilizing a replication complex during its transition from initiation to elongation. Based on the findings, including the result that the MCM‐mediated RNA replication reaction was competed with exogenously added RNA, we propose that MCM functions as a scaffold between the nascent RNA chains and the viral polymerase.
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