International Journal of Molecular Sciences

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18 pages, 3165 KiB

Open AccessCommunication

High Monocyte Count Associated with Human Cytomegalovirus Replication In Vivo and Glucocorticoid Therapy May Be a Hallmark of Disease

by Przemyslaw Zdziarski and Andrzej Gamian

Int. J. Mol. Sci. 2022, 23(17), 9595; https://doi.org/10.3390/ijms23179595 - 24 Aug 2022

Cited by 2 | Viewed by 1776

Abstract

Cytomegalovirus (CMV) syndrome and infectious disease are defined as pathogen detection with appropriate clinical symptoms, but there are not pathognomonic signs of CMV disease. Although the prodrome of acute minor viral infections leukopenia (lymphopenia and neutropenia) is noted with onset of fever, followed [...] Read more.

Cytomegalovirus (CMV) syndrome and infectious disease are defined as pathogen detection with appropriate clinical symptoms, but there are not pathognomonic signs of CMV disease. Although the prodrome of acute minor viral infections leukopenia (lymphopenia and neutropenia) is noted with onset of fever, followed by monocytosis, the role of monocytosis in CMV disease has not been described. Furthermore, under influence of corticosteroid therapy, CMV reactivation and monocytosis are described, but without a strict relationship with steroids dose. In the study, the monocyte level was investigated during the CMV infectious process. Regrettably, a non-selected group of 160 patients with high CMV viremia showed high dispersion of monocyte level and comparable with the median value for healthy subjects. Therefore, we investigated monocyte level in CMV-infected patients in relation to the logarithmic phase of the infectious process. Samples from patients with active CMV replication (exponential growth of CMV viremia) were tested. Significant monocytosis (above 1200/µL) during the logarithmic phase of CMV infection (with exponent between 3.23 and 5.77) was observed. Increased count and percentage of monocytes correlated with viral replication in several clinical situations except when there was a rapid recovery without relapse. Furthermore, glucocorticoids equivalent to 10 and 20 mg of dexamethasone during a 2–3-week period caused monocytosis—significant increase (to 1604 and 2214/µL, respectively). Conclusion: In light of the logarithmic increase of viral load, high monocytosis is a hallmark of CMV replication. In the COVID-19 era, presence of high virus level, especially part of virome (CMV) in the molecular technique, is not sufficient for the definition of either proven or probable CMV replication at any site. These preliminary observations merit additional studies to establish whether this clinical response is mediated by monocyte production or by decrease of differentiation to macrophages. Full article

(This article belongs to the Special Issue Virus Replication)

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16 pages, 3122 KiB

Open AccessArticle

Differential miRNA Expression Profiling Reveals Correlation of miR125b-5p with Persistent Infection of Japanese Encephalitis Virus

by Chih-Wei Huang, Kuen-Nan Tsai, Yi-Shiuan Chen and Ruey-Yi Chang

Int. J. Mol. Sci. 2021, 22(8), 4218; https://doi.org/10.3390/ijms22084218 - 19 Apr 2021

Cited by 16 | Viewed by 2773

Abstract

MicroRNAs (miRNAs) play versatile roles in multiple biological processes. However, little is known about miRNA’s involvement in flavivirus persistent infection. Here, we used an miRNA array analysis of Japanese encephalitis virus (JEV)-infected cells to search for persistent infection-associated miRNAs in comparison to acute [...] Read more.

MicroRNAs (miRNAs) play versatile roles in multiple biological processes. However, little is known about miRNA’s involvement in flavivirus persistent infection. Here, we used an miRNA array analysis of Japanese encephalitis virus (JEV)-infected cells to search for persistent infection-associated miRNAs in comparison to acute infection. Among all differentially expressed miRNAs, the miR-125b-5p is the most significantly increased one. The high level of miR-125b-5p in persistently JEV-infected cells was confirmed by Northern analysis and real-time quantitative polymerase chain reaction. As soon as the cells established a persistent infection, a significantly high expression of miR-125b-5p was readily observed. Transfecting excess quantities of a miR-125b-5p mimic into acutely infected cells reduced genome replication and virus titers. Host targets of miR125b-5p were analyzed by target prediction algorithms, and six candidates were confirmed by a dual-luciferase reporter assay. These genes were upregulated in the acutely infected cells and sharply declined in the persistently infected cells. The transfection of the miR125b-5p mimic reduced the expression levels of Stat3, Map2k7, and Triap1. Our studies indicated that miR-125b-5p targets both viral and host sequences, suggesting its role in coordinating viral replication and host antiviral responses. This is the first report to characterize the potential roles of miR-125b-5p in persistent JEV infections. Full article

(This article belongs to the Special Issue Virus Replication)

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15 pages, 5147 KiB

Open AccessArticle

Heat Shock Protein 90 Chaperones E1A Early Protein of Adenovirus 5 and Is Essential for Replication of the Virus

by Iga Dalidowska, Olga Gazi, Dorota Sulejczak, Maciej Przybylski and Pawel Bieganowski

Int. J. Mol. Sci. 2021, 22(4), 2020; https://doi.org/10.3390/ijms22042020 - 18 Feb 2021

Cited by 8 | Viewed by 2540

Abstract

Adenovirus infections tend to be mild, but they may pose a serious threat for young and immunocompromised individuals. The treatment is complicated because there are no approved safe and specific drugs for adenovirus infections. Here, we present evidence that 17-(Allylamino)-17-demethoxygeldanamycin (17-AAG), an inhibitor [...] Read more.

Adenovirus infections tend to be mild, but they may pose a serious threat for young and immunocompromised individuals. The treatment is complicated because there are no approved safe and specific drugs for adenovirus infections. Here, we present evidence that 17-(Allylamino)-17-demethoxygeldanamycin (17-AAG), an inhibitor of Hsp90 chaperone, decreases the rate of human adenovirus 5 (HAdV-5) replication in cell cultures by 95%. 17-AAG inhibited the transcription of early and late genes of HAdV-5, replication of viral DNA, and expression of viral proteins. 6 h after infection, Hsp90 inhibition results in a 6.3-fold reduction of the newly synthesized E1A protein level without a decrease in the E1A mRNA level. However, the Hsp90 inhibition does not increase the decay rate of the E1A protein that was constitutively expressed in the cell before exposure to the inhibitor. The co-immunoprecipitation proved that E1A protein interacted with Hsp90. Altogether, the presented results show, for the first time. that Hsp90 chaperones newly synthesized, but not mature, E1A protein. Because E1A serves as a transcriptional co-activator of adenovirus early genes, the anti-adenoviral activity of the Hsp90 inhibitor might be explained by the decreased E1A level. Full article

(This article belongs to the Special Issue Virus Replication)

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21 pages, 5364 KiB

Open AccessArticle

Discovery of a Small Molecule Inhibitor of Human Adenovirus Capable of Preventing Escape from the Endosome

by Jimin Xu, Judith Berastegui-Cabrera, Marta Carretero-Ledesma, Haiying Chen, Yu Xue, Eric A. Wold, Jerónimo Pachón, Jia Zhou and Javier Sánchez-Céspedes

Int. J. Mol. Sci. 2021, 22(4), 1617; https://doi.org/10.3390/ijms22041617 - 5 Feb 2021

Cited by 6 | Viewed by 2762

Abstract

Human adenoviruses (HAdVs) display a wide range of tissue tropism and can cause an array of symptoms from mild respiratory illnesses to disseminated and life-threatening infections in immunocompromised individuals. However, no antiviral drug has been approved specifically for the treatment of HAdV infections. [...] Read more.

Human adenoviruses (HAdVs) display a wide range of tissue tropism and can cause an array of symptoms from mild respiratory illnesses to disseminated and life-threatening infections in immunocompromised individuals. However, no antiviral drug has been approved specifically for the treatment of HAdV infections. Herein, we report our continued efforts to optimize salicylamide derivatives and discover compound 16 (JMX0493) as a potent inhibitor of HAdV infection. Compound 16 displays submicromolar IC₅₀ values, a higher selectivity index (SI > 100) and 2.5-fold virus yield reduction compared to our hit compound niclosamide. Moreover, unlike niclosamide, our mechanistic studies suggest that the antiviral activity of compound 16 against HAdV is achieved through the inhibition of viral particle escape from the endosome, which bars subsequent uncoating and the presentation of lytic protein VI. Full article

(This article belongs to the Special Issue Virus Replication)

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15 pages, 3550 KiB

Open AccessArticle

Comparison of Antiviral Activity of Gemcitabine with 2′-Fluoro-2′-Deoxycytidine and Combination Therapy with Remdesivir against SARS-CoV-2

by Yejin Jang, Jin Soo Shin, Myoung Kyu Lee, Eunhye Jung, Timothy An, Uk-Il Kim, Kyungjin Kim and Meehyein Kim

Int. J. Mol. Sci. 2021, 22(4), 1581; https://doi.org/10.3390/ijms22041581 - 4 Feb 2021

Cited by 24 | Viewed by 4078

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the coronavirus disease 2019 (COVID-19) pandemic. The virus still spreads globally through human-to-human transmission. Nevertheless, there are no specific treatments clinically approved. This study aimed to compare antiviral activity of gemcitabine [...] Read more.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the coronavirus disease 2019 (COVID-19) pandemic. The virus still spreads globally through human-to-human transmission. Nevertheless, there are no specific treatments clinically approved. This study aimed to compare antiviral activity of gemcitabine and its analogue 2′-fluoro-2′-deoxycytidine (2FdC) against SARS-CoV-2 as well as cytotoxicity in vitro. Fluorescent image-based antiviral assays revealed that gemcitabine was highly potent, with a 50% effective concentration (EC₅₀) of 1.2 μM, more active than the well-known nucleoside monophosphate remdesivir (EC₅₀ = 35.4 μM). In contrast, 2FdC was marginally active (EC₅₀ = 175.2 μM). For all three compounds, the 50% cytotoxic concentration (CC₅₀) values were over 300 μM toward Vero CCL-81 cells. Western blot and quantitative reverse-transcription polymerase chain reaction analyses verified that gemcitabine blocked viral protein expression in virus-infected cells, not only Vero CCL-81 cells but also Calu-3 human lung epithelial cells in a dose-dependent manner. It was found that gemcitabine has a synergistic effect when combined with remdesivir. This report suggests that the difluoro group of gemcitabine is critical for the antiviral activity and that its combination with other evaluated antiviral drugs, such as remdesivir, could be a desirable option to treat SARS-CoV-2 infection. Full article

(This article belongs to the Special Issue Virus Replication)

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22 pages, 2444 KiB

Open AccessArticle

Identification of Important N-Linked Glycosylation Sites in the Hemagglutinin Protein and Their Functional Impact on DC-SIGN Mediated Avian Influenza H5N1 Infection

by Zih-Syuan Yang, Szu-Wei Huang, Wen-Hung Wang, Chih-Yen Lin, Chu-Feng Wang, Aspiro Nayim Urbina, Arunee Thitithanyanont, Sung-Pin Tseng, Po-Liang Lu, Yen-Hsu Chen and Sheng-Fan Wang

Int. J. Mol. Sci. 2021, 22(2), 743; https://doi.org/10.3390/ijms22020743 - 13 Jan 2021

Cited by 11 | Viewed by 3099

Abstract

DC-SIGN, a C-type lectin mainly expressed in dendritic cells (DCs), has been reported to mediate several viral infections. We previously reported that DC-SIGN mediated H5N1 influenza A virus (AIVs) infection, however, the important DC-SIGN interaction with N-glycosylation sites remain unknown. This study aims [...] Read more.

DC-SIGN, a C-type lectin mainly expressed in dendritic cells (DCs), has been reported to mediate several viral infections. We previously reported that DC-SIGN mediated H5N1 influenza A virus (AIVs) infection, however, the important DC-SIGN interaction with N-glycosylation sites remain unknown. This study aims to identify the optimal DC-SIGN interacting N-glycosylation sites in HA proteins of H5N1-AIVs. Results from NetNGlyc program analyzed the H5 hemagglutinin sequences of isolates during 2004–2020, revealing that seven and two conserved N-glycosylation sites were detected in HA1 and HA2 domain, respectively. A lentivirus pseudotyped A/Vietnam/1203/04 H5N1 envelope (H5N1-PVs) was generated which displayed an abundance of HA5 proteins on the virions via immuno-electron microscope observation. Further, H5N1-PVs or reverse-genetics (H5N1-RG) strains carrying a serial N-glycosylated mutation was generated by site-directed mutagenesis assay. Human recombinant DC-SIGN (rDC-SIGN) coated ELISA showed that H5N1-PVs bound to DC-SIGN, however, mutation on the N27Q, N39Q, and N181Q significantly reduced this binding (p < 0.05). Infectivity and capture assay demonstrated that N27Q and N39Q mutations significantly ameliorated DC-SIGN mediated H5N1 infection. Furthermore, combined mutations (N27Q&N39Q) significantly waned the interaction on either H5N1-PVs or -RG infection in cis and in trans (p < 0.01). This study concludes that N27 and N39 are two essential N-glycosylation contributing to DC-SIGN mediating H5N1 infection. Full article

(This article belongs to the Special Issue Virus Replication)

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21 pages, 3121 KiB

Open AccessArticle

Extended Interactions between HIV-1 Viral RNA and tRNA^Lys3 Are Important to Maintain Viral RNA Integrity

by Thomas Gremminger, Zhenwei Song, Juan Ji, Avery Foster, Kexin Weng and Xiao Heng

Int. J. Mol. Sci. 2021, 22(1), 58; https://doi.org/10.3390/ijms22010058 - 23 Dec 2020

Cited by 4 | Viewed by 2787

Abstract

The reverse transcription of the human immunodeficiency virus 1 (HIV-1) initiates upon annealing of the 3′-18-nt of tRNA^Lys3 onto the primer binding site (PBS) in viral RNA (vRNA). Additional intermolecular interactions between tRNA^Lys3 and vRNA have been reported, but their functions [...] Read more.

The reverse transcription of the human immunodeficiency virus 1 (HIV-1) initiates upon annealing of the 3′-18-nt of tRNA^Lys3 onto the primer binding site (PBS) in viral RNA (vRNA). Additional intermolecular interactions between tRNA^Lys3 and vRNA have been reported, but their functions remain unclear. Here, we show that abolishing one potential interaction, the A-rich loop: tRNA^Lys3 anticodon interaction in the HIV-1 MAL strain, led to a decrease in viral infectivity and reduced the synthesis of reverse transcription products in newly infected cells. In vitro biophysical and functional experiments revealed that disruption of the extended interaction resulted in an increased affinity for reverse transcriptase (RT) and enhanced primer extension efficiency. In the absence of deoxyribose nucleoside triphosphates (dNTPs), vRNA was degraded by the RNaseH activity of RT, and the degradation rate was slower in the complex with the extended interaction. Consistently, the loss of vRNA integrity was detected in virions containing A-rich loop mutations. Similar results were observed in the HIV-1 NL4.3 strain, and we show that the nucleocapsid (NC) protein is necessary to promote the extended vRNA: tRNA^Lys3 interactions in vitro. In summary, our data revealed that the additional intermolecular interaction between tRNA^Lys3 and vRNA is likely a conserved mechanism among various HIV-1 strains and protects the vRNA from RNaseH degradation in mature virions. Full article

(This article belongs to the Special Issue Virus Replication)

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17 pages, 3450 KiB

Open AccessArticle

Amino Acid Deletions in p6^Gag Domain of HIV-1 CRF07_BC Ameliorate Galectin-3 Mediated Enhancement in Viral Budding

by Wen-Hung Wang, Chun-Sheng Yeh, Chih-Yen Lin, Ruei-Yu Yuan, Aspiro Nayim Urbina, Po-Liang Lu, Yen-Hsu Chen, Yi-Ming Arthur Chen, Fu-Tong Liu and Sheng-Fan Wang

Int. J. Mol. Sci. 2020, 21(8), 2910; https://doi.org/10.3390/ijms21082910 - 21 Apr 2020

Cited by 7 | Viewed by 2733

Abstract

HIV-1 CRF07_BC is a recombinant virus with amino acid (a.a.) deletions in p6^Gag, which are overlapped with the Alix-binding domain. Galectin-3 (Gal3), a β-galactose binding lectin, has been reported to interact with Alix and regulate HIV-1 subtype B budding. This study [...] Read more.

HIV-1 CRF07_BC is a recombinant virus with amino acid (a.a.) deletions in p6^Gag, which are overlapped with the Alix-binding domain. Galectin-3 (Gal3), a β-galactose binding lectin, has been reported to interact with Alix and regulate HIV-1 subtype B budding. This study aims to evaluate the role of Gal3 in HIV-1 CRF07_BC infection and the potential effect of a.a. deletions on Gal3-mediated regulation. A total of 38 HIV-1+ injecting drug users (IDUs) were enrolled in the study. Viral characterization and correlation of Gal3 were validated. CRF07_BC containing 7 a.a. deletions and wild-type in the p6^Gag (CRF07_BC-7d and -wt) were isolated and infectious clones were generated. Viral growth kinetic and budding assays using Jurkat-CCR5/Jurkat-CCR5-Gal3 cells infected with CRF07_BC were performed. Results indicate that 69.4% (25/38) of the recruited patients were identified as CRF07_BC, and CRF07_BC-7d was predominant. Slow disease progression and significantly higher plasma Gal3 were noted in CRF07_BC patients (p < 0.01). Results revealed that CRF07_BC infection resulted in Gal3 expression, which was induced by Tat. Growth dynamic and budding assays indicated that Gal3 expression in Jurkat-CCR5 cells significantly enhanced CRF07_BC-wt replication and budding (p < 0.05), while the promoting effect was ameliorated in CRF07_BC-7d. Co-immunoprecipitation found that deletions in the p6^Gag reduced Gal-3-mediated enhancement of the Alix–Gag interaction. Full article

(This article belongs to the Special Issue Virus Replication)

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