Biology subjects

Maruzuru, Y.

Publications and source records attributed to Maruzuru, Y..

6 recordsLinked to original sources

bioRxiv2026

Calmodulin controls the tempo of HSV-1 gene-expression cascade to reshape infection heterogeneity

Viral replication exhibits substantial heterogeneity among individual cells. Although temporally ordered gene-expression cascades are thought to contribute to such heterogeneity in DNA viruses, whether and how their tempo is regulated, and how cell-to-cell variability shapes population-level infection outcomes, remain unknown. Here, we developed a high-throughput live-cell imaging platform to track HSV-1 gene-expression dynamics in individual cells and, in combination with single-cell RNA sequencing, identified calmodulin (CaM) as a host regulator of HSV-1 gene-expression tempo. CaM accelerates progression from immediate-early to early and late phases at the single-cell level by advancing their onset. By increasing the fraction of virus-producing cells without altering burst size, CaM reshapes the distribution of infection outcomes, thereby promoting population-level viral replication. Pharmacological inhibition of CaM protects mice from lethal infection, exceeding acyclovir. These findings establish CaM-mediated temporal control as a key determinant of population-level viral output and a regulator of infection heterogeneity.

microbiology↗

bioRxiv2025

Identification of Viral Activators of the HSV-2 UL13 Protein Kinase

Although previous studies reported that the herpes simplex virus 2 (HSV-2) UL13 protein kinase mediates the phosphorylation of elongation factor 1{delta} (EF-1{delta}) in infected cells, we show here that individual expression of UL13 was insufficient to induce phosphorylation of EF-1{delta} in mammalian cells. This led us to hypothesize that HSV-2 UL13 requires viral cofactors for full kinase activity and prompted us to identify such cofactors. Our results were as follows. (i) Co-expression of UL13 with UL55 or Us10 significantly enhanced phosphorylation of EF-1{delta} compared to UL13 alone. (ii) UL13 was co-precipitated with UL55 or Us10 upon co-expression, and its kinase activity was significantly increased in their presence, as demonstrated by in vitro kinase assays. (iii) In HSV-2-infected cells, UL13 was specifically co-precipitated with Us10 and UL55. (iv) The UL55-null mutation significantly reduced phosphorylation of EF-1{delta} in HSV-2-infected cells, whereas the Us10-null mutation had little effect; however, the double-null mutation further decreased the phosphorylation compared to the UL55-null mutation alone. (v) The UL55-null mutation, but not the Us10-null mutation, significantly reduced HSV-2 replication and cell-cell spread in U2OS cells to levels comparable to those observed with the UL13 kinase-dead mutation. These results suggest that UL55 acts as a principal activator of UL13 in HSV-2-infected cells, whereas Us10 serves as an auxiliary activator. Moreover, the role of UL13 kinase activity in HSV-2 replication and cell-cell spread in U2OS cells appears to be largely dependent on UL55. ImportanceHerpesviruses encode conserved protein kinases (CHPKs) that often target cellular cyclin-dependent kinase (CDK) phosphorylation sites. CHPKs from beta-and gammaherpesviruses can exhibit these CDK-like functions even when individually expressed in mammalian cells. In contrast, CHPKs from alphaherpesviruses display these CDK-like functions in infected cells, but not upon individual expression, suggesting that they require additional viral factors to exhibit full kinase activity. In this study, we focused on HSV-2 UL13, an alphaherpesvirus CHPK, and identified HSV-2 UL55 and Us10 as viral activators of UL13. In HSV-2-infected cells, UL55 functions as a principal activator of UL13, while Us10 serves as an auxiliary activator. Importantly, the contribution of UL13 kinase activity to HSV-2 replication and cell-cell spread appears to be largely dependent on the presence of UL55. Our findings uncover a previously unrecognized mechanism of CHPK regulation in alphaherpesviruses and provide new insights into the evolutionary diversification of viral kinase control.

microbiology↗

bioRxiv2023

Regulatory Mimicry of Cyclin-Dependent Kinases by Conserved Herpesvirus Protein Kinases

Herpesviruses encode conserved protein kinases (CHPKs) that target cellular cyclin-dependent kinase (CDK) phosphorylation sites; thus, they are termed viral CDK-like kinases. Tyrosine 15 in the GxGxxG motifs of CDK1 and CDK2, whose phosphorylation down-regulates their catalytic activities, is conserved in the corresponding motifs of CHPKs. We found that herpes simplex virus 2 (HSV-2) CHPK UL13 mimicked the regulatory mechanism of CDKs. This regulatory mimicry was conserved in CHPKs encoded by herpesviruses subclassified into subfamilies other than HSV-2, suggesting CHPKs have regulatory and functional mimicry with CDKs. Phosphorylation of the corresponding Tyr in HSV-2 UL13 was required for the down-regulation of viral replication and pathogenicity, specifically in the central nervous system of mice, and for efficient viral recurrence in guinea pigs. These data highlight the dual impact of the regulatory mimicry of CDKs by CHPK on the fine-tuned regulation of lytic and latent HSV-2 infections in vivo.

microbiology↗

bioRxiv2023

Evasion of APOBEC1-mediated Intrinsic Immunity by a Herpesvirus Uracil DNA Glycosylase Is a Determinant of Viral Encephalitis

Herpes simplex virus 1 (HSV-1) is the most common cause of viral encephalitis, which can be lethal or result in severe neurological defects, even when treated with antiviral therapy. We demonstrated that activation of HSV-1 uracil-DNA glycosylase (vUNG) by phosphorylation, essential for its enzymatic activity, counteracted APOBEC1 to promote viral replication and encephalitis in the central nervous system (CNS) of mice. The activation of vUNG protected HSV-1 genomes from APOBEC1-mediated DNA editing, allowing efficient viral replication to occur. The presence of APOBEC1 markedly improved lethal encephalitis in mice infected with an HSV-1 mutant carrying a mutation in the phosphorylation site and an UNG inhibitor protected wild-type HSV-1-infected mice from lethal encephalitis. These findings re-define vUNG as an important factor that allows evasion from intrinsic anti-viral immunity mediated by APOBEC1 in the CNS, and suggest a new therapeutic approach for the treatment of fetal and critical HSV-1 encephalitis.

microbiology↗

bioRxiv2023

Direct Relationship between Protein Expression and Progeny Yield of Herpes Simplex Virus 1 Unveils a Rate-limiting Step for Virus Production

Although viral protein expression and progeny virus production were independently shown to be highly heterogenous in individual cells, their direct relationship, analyzed by considering their heterogeneities, has not been investigated to date. This study established a system to fractionate cells infected with a herpesvirus based on the levels of the global expression of viral late proteins, which are largely virion structural proteins, and to titrate virus yields in these fractions. This system demonstrated a direct relationship and indicated there was a threshold for the levels of viral late protein expression for progeny virus production and suggested that viral DNA cleavage/packaging was a rate-limiting step for progeny virus production. These findings, which were masked in previous studies performed at the entire population level, have uncovered a sophisticated viral strategy for efficient progeny virus production and shed new light on an effective target for the development of anti-viral drugs.

microbiology↗

bioRxiv2022

Dual Impacts of a Glycan Shield on the Envelope Glycoprotein B of HSV-1: Evasion from Human Antibodies In Vivo and Neurovirulence

Identification of the mechanisms of viral evasion from human antibodies is crucial both for understanding viral pathogenesis and for designing effective vaccines. However, the in vivo efficacy of the mechanisms of viral evasion from human antibodies has not been well documented. Here we show in cell cultures that an N-glycan shield on the HSV-1 envelope glycoprotein B (gB) mediated evasion from neutralization and antibody-dependent cellular cytotoxicity due to pooled {gamma}-globulins derived from human blood. We also demonstrated that the presence of human {gamma}-globulins in mice and HSV-1 immunity induced by viral infection in mice significantly reduced the replication of a mutant virus lacking the glycosylation site in a peripheral organ but had little effect on the replication of its repaired virus. These results suggest that the glycan shield on the HSV-1 envelope gB mediated evasion from human antibodies in vivo and from HSV-1 immunity induced by viral infection in vivo. Notably, we also found that the glycan shield on HSV-1 gB was significant for HSV-1 neurovirulence and replication in the central nervous system (CNS) of naive mice. Thus, we have identified a critical glycan shield on HSV-1 gB that has dual impacts, namely evasion from human antibodies in vivo and viral neurovirulence. IMPORTANCEHSV-1 establishes lifelong latent and recurrent infections in humans. To produce recurrent infections that contribute to transmission of the virus to new human host(s), the virus must be able to evade the antibodies persisting in latently infected individuals. Here we show that an N-glycan shield on the envelope glycoprotein B of HSV-1 mediates evasion from pooled {gamma}-globulins derived from human blood both in cell cultures and mice. Notably, the N-glycan shield was also significant for HSV-1 neurovirulence in naive mice. Considering the clinical features of HSV-1 infection, these results suggest that the glycan shield not only facilitates recurrent HSV-1 infections in latently infected humans by evading antibodies, but is also important for HSV-1 pathogenesis during the initial infection.

microbiology↗