Binding of HSV-1 Glycoprotein K Allen, Sariah J; Mott, Kevin R; Matsuura, Yoshiharu ...
PloS one,
01/2014, Letnik:
9, Številka:
1
Journal Article
Recenzirano
Glycoprotein K (gK) is a virion envelope protein of herpes simplex virus types 1 (HSV-1) and 2 (HSV-2), which plays important roles in virion entry, morphogenesis and egress. Two-hybrid and pull-down ...assays were utilized to demonstrate that gK and no other HSV-1 genes specifically binds to signal peptide peptidase (SPP), also known as minor histocompatibility antigen H13. SPP dominant negative mutants, shRNA against SPP significantly reduced HSV-1 replication in vitro. SPP also affected lysosomes and ER responses to HSV-1 infection. Thus, in this study we have shown for the first time that gK, despite its role in fusion and egress, is also involved in binding the cytoplasmic protein SPP. These results also suggest that SPP plays an important role in viral replication and possibly virus pathogenesis. This makes SPP unique in that its function appears to be required by the virus as no other protein can compensate its loss in terms of viral replication.
We previously have described a model of multiple sclerosis (MS) in which constitutive expression of murine interleukin (IL)-2 by herpes simplex virus type 1 (HSV-1) (HSV-IL-2) causes central nervous ...system (CNS) demyelination in different strains of mice. In the current study, we investigated whether this HSV-IL-2-induced demyelination can be blocked using recombinant viruses expressing different cytokines or by injection of plasmid DNA. We have found that coinfection of HSV-IL-2-infected mice with recombinant viruses expressing IL-12p35, IL-12p40 or IL-12p35+IL-12p40 did not block the CNS demyelination, and that coinfection with a recombinant virus expressing interferon (IFN)-γ exacerbated it. In contrast, coinfection with a recombinant virus expressing IL-4 reduced demyelination, whereas coinfection of HSV-IL-2-infected mice with a recombinant HSV-1 expressing the IL-12 heterodimer (HSV-IL-12p70) blocked the CNS demyelination in a dose-dependent manner. Similarly, injection of IL-12p70 DNA blocked HSV-IL-2-induced CNS demyelination in a dose-dependent manner and injection of IL-35 DNA significantly reduced CNS demyelination. Injection of mice with IL-12p35 DNA, IL-12p40 DNA, IL-12p35+IL-12p40 DNA or IL-23 DNA did not have any effect on HSV-IL-2-induced demyelination, whereas injection of IL-27 DNA increased the severity of the CNS demyelination in the HSV-IL-2-infected mice. This study demonstrates for the first time that IL-12p70 can block HSV-IL-2-induced CNS demyelination and that IL-35 can also reduce this demyelination, whereas IFN-γ and IL-27 exacerbated the demyelination in the CNS of the HSV-IL-2-infected mice. Our results suggest a potential role for IL-12p70 and IL-35 signaling in the inhibition of HSV-IL-2-induced immunopathology by preventing development of autoaggressive T cells.
We have shown previously that exacerbation of corneal scarring (CS) in HSV-1 glycoprotein K (gK) immunized mice was associated with CD8 super(+) T cells. In this study, we investigated the type and ...the nature of the immune responses that are involved in the exacerbation of CS in gK-immunized animals. BALB/c mice were vaccinated with baculovirus expressed gK, gD, or mock-immunized. Twenty-one days after the third immunization, mice were ocularly infected with 2 x 10 super(5) PFU/eye of virulent HSV-1 strain McKrae. Infiltration of the cornea by CD4 super(+), CD8 super(+), CD25 super(+), CD4 super(+)CD25 super(+), CD8 super(+)CD25 super(+), CD19 super(+), CD40 super(+), CD40L super(+), CD62L super(+), CD95 super(+), B7-1 super(+), B7-2 super(+), MHC-I super(+), and MHC-II super(+) cells was monitored by immunohistochemistry, qRT-PCR and FACS at various times post-infection (PI). This study demonstrated for the first time that the presence of CD8 super(+)CD25 super(+) T cells in the cornea is correlated with exacerbation of CS in the gK-immunized group.
Glycoprotein K (gK) is a virion envelope protein of herpes simplex virus types 1 (HSV-1) and 2 (HSV-2), which plays important roles in virion entry, morphogenesis and egress. Two-hybrid and pull-down ...assays were utilized to demonstrate that gK and no other HSV-1 genes specifically binds to signal peptide peptidase (SPP), also known as minor histocompatibility antigen H13. SPP dominant negative mutants, shRNA against SPP significantly reduced HSV-1 replication in vitro. SPP also affected lysosomes and ER responses to HSV-1 infection. Thus, in this study we have shown for the first time that gK, despite its role in fusion and egress, is also involved in binding the cytoplasmic protein SPP. These results also suggest that SPP plays an important role in viral replication and possibly virus pathogenesis. This makes SPP unique in that its function appears to be required by the virus as no other protein can compensate its loss in terms of viral replication.
It is generally accepted that CD8 T cells play the key role to maintain HSV-1 latency in trigeminal ganglia of ocularly infected mice. Yet, comparably little is known about the role of innate ...immunity in establishment of viral latency. In the current study, we investigated whether CD8 alpha DCs impact HSV-1 latency by examining latency in the trigeminal ganglia (TG) of wild-type (WT) C57BL/6 versus CD8 alpha -/- (lack functional CD8 T cells and CD8 alpha + DCs), CD8 beta -/- (have functional CD8 alpha + T cells and CD8 alpha + DCs), and beta 2m-/- (lack functional CD8 T cells but have CD8 alpha + DCs) mice as well as BXH2 (have functional CD8 T cells but lack CD8 alpha + DCs) versus WT C3H (have functional CD8 alpha T cells and CD8 alpha + DCs) mice. We also determined whether the phenotype of CD8 alpha -/- and BXH2 mice could be restored to that of WT mice by adoptive transfer of WT CD8+ T cells or bone marrow (BM) derived CD8 alpha + DCs. Our results clearly demonstrate that CD8 alpha DCs, rather than CD8 T cells, are responsible for enhanced viral latency and recurrences.
CD80 plays a critical role in stimulation of T cells and subsequent control of infection. To investigate the effect of CD80 on HSV-1 infection, we constructed a recombinant HSV-1 virus that expresses ...two copies of the CD80 gene in place of the latency associated transcript (LAT). This mutant virus (HSV-CD80) expressed high levels of CD80 and had similar virus replication kinetics as control viruses in rabbit skin cells. In contrast to parental virus, this CD80 expressing recombinant virus replicated efficiently in immature dendritic cells (DCs). Additionally, the susceptibility of immature DCs to HSV-CD80 infection was mediated by CD80 binding to PD-L1 on DCs. This interaction also contributed to a significant increase in T cell activation. Taken together, these results suggest that inclusion of CD80 as a vaccine adjuvant may promote increased vaccine efficacy by enhancing the immune response directly and also indirectly by targeting to DC.
Activation induced cytidine deaminase (AID) is required for somatic hypermutation (SHM) and class-switch recombination (CSR). AID initiates the processes that carry out immunoglobulin diversity by ...deaminating cytosine residues within variable (V) and switch (S) regions on the Ig locus during active transcription. The resulting G:U mispairs can then be replicated or repaired by cellular repair mechanisms to give rise to isotype-switched and antigen-specific mature antibodies. In this study I have identified two novel phosphorylation sites, serine 41 and serine 43, and demonstrated their importance in AID activity as well as confirmed the importance of serine 38 phosphorylation. Phosphorylation null mutants generated by replacing serine with alanine are much less active than wild-type AID, as is non-phosphorylated AID purified from E. coli. In contrast, phosphorylation charge mimic mutants generated by replacing serine with aspartic acid, are (3-4) fold more active than wild-type AID. Phosphorylation does not affect the ability of AID to deaminate actively transcribed double-stranded DNA or its ability to preferentially deaminate WRC hot spots. No observed affect on DNA binding strength was detected suggesting that mutant enzymes are folded like wild-type AID. The highly conserved nature of these phosphorylation sites and the identification of Hyper-IgM Type 2 (HIGM2) patients with mutations at serine 43, taken together with our biochemical data suggest that these phosphorylation sites are required for proper AID function in B cells. I propose a model whereby AID phosphorylation is involved in the regulation of AID activity and efficient targeting during SHM and CSR. Additionally, I propose that phosphorylation serves as a gatekeeper for genomic integrity, protecting the B cell genome against inappropriate deamination by providing a rapid activation and targeting mechanism.
Abstract
Radiation-induced high-grade gliomas (RIGs) are an incurable late complication of cranial radiation therapy. We performed DNA methylation profiling, RNA-seq, and DNA sequencing on 32 RIG ...tumors and an in vitro drug screen in two RIG cell lines. We report that based on DNA methylation, RIGs cluster primarily with the pediatric receptor tyrosine kinase I high-grade glioma subtype. Common copy-number alterations include Chromosome (Ch.) 1p loss/1q gain, and Ch. 13q and Ch. 14q loss; focal alterations include
PDGFRA
and
CDK4
gain and
CDKN2A
and
BCOR
loss. Transcriptomically, RIGs comprise a stem-like subgroup with lesser mutation burden and Ch. 1p loss and a pro-inflammatory subgroup with greater mutation burden and depleted DNA repair gene expression. Chromothripsis in several RIG samples is associated with extrachromosomal circular DNA-mediated amplification of
PDGFRA
and
CDK4
. Drug screening suggests microtubule inhibitors/stabilizers, DNA-damaging agents, MEK inhibition, and, in the inflammatory subgroup, proteasome inhibitors, as potentially effective therapies.
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