Resistance to maribavir is associated with the exclusion of pUL27 from nucleoli during human cytomegalovirus infection

Select mutations in the human cytomegalovirus (HCMV) gene UL27 confer low-grade resistance to the HCMV UL97 kinase inhibitor maribavir (MBV). It has been reported that the 608-amino acid UL27 gene product (pUL27) normally localizes to cell nuclei and nucleoli, whereas its truncation at codon 415, as found in a MBV-resistant mutant, results in cytoplasmic localization. We now show that in the context of full-length pUL27, diverse single amino acid substitutions associated with MBV resistance result in loss of its nucleolar localization when visualized after transient transfection, whereas substitutions representing normal interstrain polymorphism had no such effect. The same differences in localization were observed during a complete infection cycle with recombinant HCMV strains over-expressing full-length fluorescent pUL27 variants. Nested UL27 C-terminal truncation expression plasmids showed that amino acids 596-599 were required for the nucleolar localization of pUL27. These results indicate that the loss of a nucleolar function of pUL27 may contribute to MBV resistance, and that the nucleolar localization of pUL27 during HCMV infection depends not only on a carboxy-terminal domain but also on a property of pUL27 that is affected by MBV-resistant mutations, such as an interaction with component(s) of the nucleolus.     

精神发育迟滞患儿母亲生活质量的调查分析

目的探讨精神发育迟滞患儿母亲的生活质量状况。方法采用生活质量综合评定问卷-74(GQOLI-74)对26例精神发育迟滞患儿母亲及与之相匹配的26例正常儿童的母亲进行评定。结果①精神发育迟滞患儿母亲GQOLI-74维度评分和总体生活状态评分均明显低于正常对照组(t=2.11,2.17,2.88,2.94,3.05;P均<0.05或<0.01),具有统计学意义,其中以躯体和心理功能尤为显著;②文化程度低的患儿母亲GQOLI-74维度评分和总体生活状态评分均低于文化程度高者,大部分因子和总体生活状态差异显著(t=2.08,2.10,2.12,2.20;P均<0.05),具有统计学意义。结论精神发育迟滞患儿母亲的生活质量较正常对照者差。     

RNase P体外靶定切割人巨细胞病毒UL54 mRNA片段研究

目的：观察大肠埃希菌RNase P催化亚基M1 RNA在细胞外水平上对人巨细胞病毒（human cytomegalovirus,HCMV）DNA聚合酶UL54 mRNA的切割效果,探讨核酶作为新型抗病毒制剂的应用前景。方法：针对HCMVUL54 mRNA设计特异性的外部引导序列（external guide sequences,EGSs）EGS-T6,观察其引导M1 RNA特异的细胞外切割活性。结果：在EGS-T6引导下,大肠埃希菌RNase P催化亚基M1 RNA在细胞外可特异地对靶mRNA进行有效切割。结论：EGS-T6具备引导M1 RNA对UL54 mRNA特异切割的能力,可发展成一种新型抗病毒制剂。     

Induction of angiogenic chemokine CCL2 by human herpesvirus 8 chemokine receptor

Human herpesvirus 8 (HHV-8) is associated with Kaposi's sarcoma (KS), an endothelial cell lesion believed to be initiated and driven primarily by cytokine dysregulation. Among the viral proteins suspected as contributing to viral pathogenesis is the lytically expressed viral G protein-coupled receptor (vGPCR), which can induce various cellular cytokines. CC ligand-2 (CCL2/MCP-1) is a vGPCR-regulated angiogenic chemokine found at elevated levels in KS lesions and induced by HHV-8 infection of endothelial cells. Here we show that vGPCR induces CCL2 in endothelial cells via activation of C/EBPβ and that vGPCR and C/EBPβ are critical components of CCL2 induction by HHV-8 infection of endothelial cultures. To our knowledge, this is the first report of vGPCR-mediated cytokine induction, and its characterization, in the context of virus infection. Our results identify a mechanism by which vGPCR can contribute, in a host cell shutoff-independent manner, to viral pathogenesis.     

Human cytomegalovirus (HCMV) UL139 open reading frame: Sequence variants are clustered into three major genotypes

Human cytomegalovirus (HCMV) infects a number of organs and cell types, leading to the hypothesis that HCMV disease and tissue tropism may be related to specific sequence variability. This study examined the genomic variability of a new polymorphic locus in HCMV, UL139 open reading frame (ORF). Detailed analysis showed that a large number of nucleotide insertions and non-synonymous substitutions occurred in the UL139 ORF, particularly in the 5' half, using the Toledo strain as the reference sequence. The UL139 variants were not distributed randomly, but were clustered clearly into three major groups: G1 (G1a, G1b, and G1c), G2 (G2a, G2b), and G3. In this study, it was found that the predicted UL139 product shared sequence homology with human CD24, a signal transducer modulating B-cell activation responses, and the sequences in G1c contained a specific attachment site of prokaryotic membrane lipoprotein lipid. The precise definition of UL139 genotypes and its putative function would be helpful in understanding better HCMV.     

Niacin: a re-emerging pharmaceutical for the treatment of dyslipidaemia

Dyslipidaemias, particularly those characterized by the ‘atherogenic profile’ of high low-density lipoprotein-cholesterol and triglycerides and low high-density lipoprotein-cholesterol, are the major modifiable risk factor for atherosclerosis. The search for drugs to favourably alter such lipid profiles, reducing the associated morbidity and mortality, remains a major research focus. Niacin (nicotinic acid) is the most effective agent available for increasing high-density lipoprotein-cholesterol, but its use is associated with side effects that negatively affect patient compliance: these appear to arise largely as a result of production of prostaglandin D₂ and its subsequent activation of the DP₁ receptor. Desire to reduce the side effects (and improve pharmacokinetic parameters) has led to the development of a number of agonists that have differing effects, both in terms of clinical potency and the severity of adverse effects. The recent discovery of the niacin G-protein-coupled receptor HM74A (GPR109A) has clarified the distinction between the mechanism whereby niacin exerts its therapeutic effects and the mechanisms responsible for the generation of side effects. This has allowed the development of new drugs that show great potential for the treatment of dyslipidaemia. However, recent advances in understanding of the contribution of prostaglandin metabolism to vascular wall health suggest that some of the beneficial effects of niacin may well result from activation of the same pathways responsible for the adverse reactions. The purpose of this review is to emphasize that the search for agonists that show higher tolerability must take into account all aspects of signalling through this receptor.     

Characterization of molecular determinants for nucleocytoplasmic shuttling of PRV UL54

The pseudorabies virus (PRV) early protein UL54 is a homologue of the herpes simplex virus 1 (HSV-1) immediate-early protein ICP27, which is a multifunctional protein and essential for HSV-1 infection. To determine if UL54 might shuttle between the nucleus and cytoplasm, as has been shown for its homologues in human herpesviruses, the molecular determinants for its nucleocytoplasmic shuttling were investigated. Heterokaryon assays demonstrated that UL54 was a nucleocytoplasmic shuttling protein and this property could not be blocked by leptomycin B, an inhibitor of chromosome region maintenance 1 (CRM1). However, TAP/NXF1 promoted the nuclear export of UL54 and interacted with UL54, suggesting that UL54 shuttles between the nucleus and the cytoplasm via a TAP/NXF1, but not CRM1, dependent nuclear export pathway. Furthermore, UL54 was demonstrated to target to the nucleus through a classic Ran-, importin β1- and α5-dependent nuclear import mechanism.     

Sites and roles of phosphorylation of the human cytomegalovirus DNA polymerase subunit UL44

The human cytomegalovirus DNA polymerase subunit UL44 is a phosphoprotein, but its sites and roles of phosphorylation have not been investigated. We compared sites of phosphorylation of UL44 in vitro by the viral protein kinase UL97 and cyclin-dependent kinase 1 with those in infected cells. Transient treatment of infected cells with a UL97 inhibitor greatly reduced labeling of two minor UL44 phosphopeptides. Viruses containing alanine substitutions of most UL44 residues that are phosphorylated in infected cells exhibited at most modest effects on viral DNA synthesis and yield. However, substitution of highly phosphorylated sites adjacent to the nuclear localization signal abolished viral replication. The results taken together are consistent with UL44 being phosphorylated directly by UL97 during infection, and a crucial role for phosphorylation-mediated nuclear localization of UL44 for viral replication, but lend little support to the widely held hypothesis that UL97-mediated phosphorylation of UL44 is crucial for viral DNA synthesis.     

The early UL3 gene of equine herpesvirus-1 encodes a tegument protein not essential for replication or virulence in the mouse

The UL3 gene of equine herpesvirus-1 (EHV-1) is retained in the genome of defective interfering particles and encodes a ~ 33 kDa myristylated protein. Further characterization showed that the UL3 gene is trans-activated only by the sole immediate early (IE) protein and encodes an early protein that is dispensable for EHV-1 replication and localizes in the tegument of purified virions. UL3-deleted EHV-1 (vL11ΔUL3) exhibits properties of host cell tropism, plaque size, and growth kinetics similar to those of the parental virus. Expression levels of EHV-1 proteins representative of all three gene classes in vL11ΔUL3-infected cells were identical to those in cells infected with parental virus. Mice intranasally infected with vL11ΔUL3 and parental virus showed no significant difference in mortality or virus lung titers. These findings suggest that the UL3 protein does not play a major role in the biology of EHV-1 in cell culture or virulence in the mouse.