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Understanding the functional characteristics of antibodies produced against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) will assist in the determination of disease outcomes for this virus. In this study, the ability of antibodies to inhibit viral entry into the host cell through the interaction of the receptor binding domain of the viral spike protein and the angiotensin-converting enzyme 2 receptor on the human cell surface was investigated. The SARS-CoV-2 IgG levels in 20 SARS-CoV-2 positive patients were measured using an enzyme-linked immunosorbent assay, and the samples were further analyzed using a functional binding assay. Inhibition of viral infectivity was also measured using a pseudovirus neutralization assay against a D614G SARS-CoV-2 virus strain. A significant correlation between IgG levels and neutralizing antibody 50% inhibitory concentration (IC50) titers was observed (p < 0.05). Similarly, the IC50 titers obtained in the neutralization and binding assays were significantly correlated (p < 0.001). Varying levels of IgG and IC50 titers were observed for the SARS-CoV-2 antibody-positive samples, with one sample not showing any neutralizing capability despite detectable IgG levels. Gender comparisons showed no statistical differences in any of the assays. These results suggest that increased SARS-CoV-2 IgG levels correlate with greater protection against the entry of the virus into cells; however, further investigations in larger studies are needed to confirm the correlates of protection. [ABSTRACT FROM AUTHOR]

Adult T-cell leukemia/lymphoma (ATL) is an intractable T-cell malignancy accompanied by massive invasion of lymphoma cells into various tissues. We demonstrate here that ATL cells cultured on a layer of epithelial-like feeder cells form anchorage-dependent multicellular aggregates (Ad-MCAs) and that a fraction of MCA-forming ATL cells acquire CD44 high cancer stem cell-like phenotypes. ATL cells forming Ad-MCAs displayed extracellular microvesicles with enhanced expression of CD44v9 at cell synapses, augmented expression of multidrug resistance protein 1, and increased NF-κB activity. Blockade of the NF-κB pathway dramatically reduced Ad-MCA formation by ATL cells and the emergence of CD44 high ATL cells, but left a considerable number of ATL cells adhering to the feeder layer. Disruption of vimentin cytoskeleton by treatment with withaferin A, a natural steroidal lactone, suppressed not only the adhesion of ATL cells to the feeder layer but also subsequent Ad-MCA formation by ATL cells, suggesting the involvement of vimentin in anchoring ATL cells to the feeder layer. Ad-MCA formation by ATL cells on a layer of epithelial-like feeder cells may mimic critical events that occur in metastatic colonization. [ABSTRACT FROM AUTHOR]

HTLV-1 is the etiologic agent of the adult T cell leukemia-lymphoma (ATLL). The viral regulatory protein Tax plays a central role in leukemogenesis as a transcriptional transactivator of both viral and cellular gene expression, and this requires Tax activity in both the cytoplasm and the nucleus. In the present study, we have investigated the mechanisms involved in the nuclear localization of Tax. Employing a GFP fusion expression system and a range of Tax mutants, we could confirm that the N-terminal 60 amino acids, and specifically residues within the zinc finger motif in this region, are important for nuclear localization. Using an in vitro nuclear import assay, it could be demonstrated that the transportation of Tax to the nucleus required neither energy nor carrier proteins. Specific and direct binding between Tax and p62, a nucleoporin with which the importin beta family of proteins have been known to interact was also observed. The nuclear import activity of wild type Tax and its mutants and their binding affinity for p62 were also clearly correlated, suggesting that the entry of Tax into the nucleus involves a direct interaction with nucleoporins within the nuclear pore complex (NPC). The nuclear export of Tax was also shown to be carrier independent. It could be also demonstrated that Tax it self may have a carrier function and that the NF-κB subunit p65 could be imported into the nucleus by Tax. These studies suggest that Tax could alter the nucleocytoplasmic distribution of cellular proteins, and this could contribute to the deregulation of cellular processes observed in HTLV-1 infection. [ABSTRACT FROM AUTHOR]

The human polyomavirus JC virus (JCV) is ubiquitous and can be shed in the urine of more than 40% of the healthy population. Amplification and sequencing of JCV from urine has allowed a distinctive map of the distribution of JCV genotypes worldwide. To define the frequency of JCV urinary excretion and genotype distribution in Ireland, urines from 121 healthy individuals and from 94 immunocompromised individuals (human immunodeficiency virus [HIV]-positive patients and rheumatoid arthritis patients) were collected. JCV DNA was detected by polymerase-chain reaction (PCR) with subsequent nucleotide sequencing of a fragment of the major capsid protein (VP1). JCV was detected in 20.7% of healthy individuals and was found significantly more often in the urine of HIV-positive patients (54.2%; P [ABSTRACT FROM AUTHOR]

Background: The Tax proteins encoded by human T lymphotropic virus type 1 (HTLV-1) and type 2 (HTLV-2) are transcriptional activators of both the viral long terminal repeat (LTR) and cellular promoters via the CREB and NFkB pathways. In contrast to HTLV-1, HTLV-2 has been classified into four distinct genetic subtypes A, B, C and D defined by phylogenetic analysis of their nucleotide sequences and the size and amino acid sequence of their Tax proteins. In the present study we have analysed and compared the transactivating activities of three Tax 2A and one Tax 2B proteins using LTR and NFkB reporter assays. Results: We found that with the exception of the prototype Tax 2A Mo protein, the other two Tax 2A proteins failed to transactivate either the viral LTR or NFkB promoter in Jurkat and 293T cells. Loss of activity was not associated with either expression levels or an alteration in subcellular distribution as all Tax 2 proteins were predominantly located in the cytoplasm of transfected cells. Analysis of the sequence of the two inactive Tax 2A proteins relative to Mo indicated that one had six amino acid changes and the other had one change in the central region of the protein. Mutations present at the amino and the extreme carboxy termini of Mo resulted in the loss of LTR but not NFkB activation whereas those occurring in the central region of the protein appeared to abolish transactivation of both promoters. Analysis of the transactivation phenotypes of Tax 1, Tax 2A Mo and Tax 2B containing mutations identified in the present study or previously characterised Tax mutations showed that domains required for LTR and NFkB activation are very similar but not identical in all three Tax proteins. Conclusion: Our results suggest that loss of activity of two Tax 2A proteins derived from different isolates is associated with multiple amino acid changes relative to Mo in domains required for the activation of the CREB or CREB and NFkB pathways and that these domains are very similar but not identical in Tax 2B and Tax 1. The loss of Tax function in 2A viruses may have implications for their biological and pathogenic properties. [ABSTRACT FROM AUTHOR]

The primary function of the HIV-1 regulatory protein Tat, activation of transcription from the viral LTR, is highly regulated by complex interactions between Tat and a number of host cell proteins. Tat nuclear import, a process mediated by importin β, is a prerequisite for its activity. Here, we report and characterize the interaction of the human inhibitor of MyoD family domain-containing protein (l-mfa), HIC, with Tat at a biochemical and a functional level. This interaction was shown to occur in vivo and in vitro and to involve the nuclear localization signal and the transactivation responsive element-binding domains of Tat and the I-mfa domain of HIC. Coexpression of HIC and Tat resulted in the down-regulation of transactivation of the HIV-1 LTR, and colocalization studies revealed the cytoplasmic sequestration of Tat by HIC. Functionally this sequestration appears to be the underlying mechanism of LTR transcriptional repression by HIC and represents a unique mechanism for the control of Tat activity and regulation of HIV-1 replication. [ABSTRACT FROM AUTHOR]

The function of the transactivator protein Tax from HTLV-II subtype A, subtype B, Brazilian subtype C, and African subtype D isolates was compared to that of Tax from an HTLV-I isolate. HTLV-II subtypes A, B, and C were less active in the transactivation of a NF-κB reporter compared to HTLV-I Tax in 293T but not Jurkat T cells. In both cell types there were no significant differences between the functions of HTLV-II B, C, and D and HTLV-I Tax proteins on either a full-length HTLV-I LTR or a 21-bp repeat reporter, suggesting that there is equivalent CREB-mediated transactivation between the viruses and these subtypes. In contrast, Tax of some but not all HTLV-II subtype A isolates, including the prototype Mo, had a greatly decreased ability to transactivate, and this could be directly correlated with a decrease in protein expression. Employment of cDNA clones encoding both Rex and Tax demonstrated that Rex was unable to rescue the expression or activity of the IIA Mo isolate. These studies demonstrate that with the exception of some HTLV-IIA subtypes there are no significant differences in Tax transactivation via the CREB and NF-κB pathways between the two viruses and suggest that the HTLV-II Tax may have a pathogenic potential equivalent to that of HTLV-I. [Copyright &y& Elsevier]

The association of the severity of liver disease and the molecular evolution of hepatitis C virus (HCV) during chronic infection remains unclear and controversial. To address this we have studied the interpatient variability in the nucleotide sequence of two regions of the HCV genome, E1/E2, which contain the hypervariable region 1 and the nonstructural NS5b region, in a cohort of Irish female patients who were all recipients of a single source of HCV genotype 1b-contaminated anti-D immunoglobulin in 1977 and 1978 and who over the subsequent 20 years developed a spectrum of liver disease. In addition, quasispecies analysis was used to evaluate intrapatient variability in the E1/E2 region in four patients with mild and four with severe disease. Phylogenetic and evolutionary rate analyses of the nucleotide sequences demonstrated that there was no significant difference between those who developed mild disease and those who had progressed to severe disease or cirrhosis. These findings suggest that other factors, either additional viral or host, may be important in the pathogenesis and clinical outcome of chronic hepatitis C virus infection. [Copyright &y& Elsevier]

The divergent clinical outcomes of human T cell leukemia virus type 1 (HTLV-1) and HTLV-2 infections have been attributed to functional differences in their antisense proteins. In contrast to HTLV-1 bZIP factor (HBZ), the role of the antisense protein of HTLV-2 (APH-2) in HTLV-2 infection is poorly understood. In previous studies, we identified the endosomal sorting complex required for transport 0 (ESCRT-0) subunit HRS as a novel interaction partner of APH-2 but not HBZ. HRS is a master regulator of endosomal protein sorting for lysosomal degradation and is hijacked by many viruses to promote replication. However, no studies to date have shown a link between HTLVs and HRS. In this study, we sought to characterize the interaction between HRS and APH-2 and to investigate the impact of HRS on the life cycle of HTLV-2. We confirmed a direct specific interaction between APH-2 and HRS and showed that the CC2 domain of HRS and the N-terminal domain of APH-2 mediate their interaction. We demonstrated that HRS recruits APH-2 to early endosomes, possibly furnishing an entry route into the endosomal/lysosomal pathway. We demonstrated that inhibition of this pathway using either bafilomycin or HRS overexpression substantially extends the half-life of APH-2 and stabilizes Tax2B expression levels. We found that HRS enhances Tax2B-mediated long terminal repeat (LTR) activation, while depletion of HRS enhances HTLV-2 production and release, indicating that HRS may have a negative impact on HTLV-2 replication. Overall, our study provides important new insights into the role of the ESCRT-0 HRS protein, and by extension the ESCRT machinery and the endosomal/lysosomal pathway, in HTLV-2 infection. [ABSTRACT FROM AUTHOR]

The human T-cell leukemia virus type 1 (HTLV-1) regulatory proteins Tax and HBZ play indispensable roles in regulating viral and cellular gene expression. BRG1, the ATPase subunit of the SWI/SNF chromatin remodeling complex, has been demonstrated to be essential not only for Tax transactivation but also for viral replication. We sought to investigate the physical interaction between HBZ and BRG1 and to determine the effect of these interactions on Tax-mediated long terminal repeat (LTR) activation. We reveal that HTLV-1 cell lines and adult T-cell leukemia (ATL) cells harbor high levels of BRG1. Using glutathione S-transferase (GST) pulldown and coimmunoprecipitation assays, we have demonstrated physical interactions between BRG1 and HBZ and characterized the protein domains involved. Moreover, we have identified the PBAF signature subunits BAF200 and BAF180 as novel interaction partners of HBZ, suggesting that the PBAF complex may be required for HTLV-1 transcriptional repression by HBZ. Additionally, we found that BRG1 expression translocates HBZ into distinct nuclear foci. We show that HBZ substantially represses HTLV-1 LTR activation by Tax/BRG1. Interestingly, we found that Tax stabilizes the expression of exogenous and endogenous BRG1 and that HBZ reverses this effect. Finally, using a chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR) assay, we illustrate that HBZ facilitates the downregulation of HTLV-1 transcription by deregulating the recruitment of SWI/SNF complexes to the promoter. Overall, we conclude that SWI/SNF complexes, in addition to other cellular transcription factors, are involved in HBZ-mediated suppression of HTLV-1 viral gene expression. IMPORTANCE The pathogenic potential of HTLV-1 is linked to the indispensable multifaceted functions of the viral regulatory proteins Tax and HBZ, encoded by the sense and antisense viral transcripts, respectively. The interaction between Tax and the SWI/SNF family of chromatin remodeling complexes has been associated with HTLV-1 transcriptional activation. To date, the relationship between the SWI/SNF chromatin remodeling family and HBZ, the only viral protein that is consistently expressed in infected cells and ATL cells, has not been elucidated. Here, we have characterized the biological significance of the SWI/SNF family in regard to viral transcriptional repression by HBZ. This is important because it provides a better understanding of the function and role of HBZ in downregulating viral transcription and, hence, its contribution to viral latency and persistence in vivo, a process that may ultimately lead to the development of ATL. [ABSTRACT FROM AUTHOR]

The pathogenesis of human T cell leukemia virus type 1 (HTLV-1) is strongly linked to the viral regulatory proteins Tax1 and HBZ, whose opposing functions contribute to the clinical outcome of infection. Type I interferons alpha and beta (IFN-α and IFN-β) are key cytokines involved in innate immunity, and IFN-α, in combination with other antivirals, is extensively used in the treatment of HTLV-1 infection. The relationship between HTLV-1 and IFN signaling is unclear, and to date the effect of HBZ on this pathway has not been examined. Here we report that HBZ significantly enhances interferon regulatory factor 7 (IRF7)-induced IFN-α- and IFNstimulated response element (ISRE) promoter activities and IFN-α production and can counteract the inhibitory effect of Tax1. In contrast to this, we show that HBZ and Tax1 cooperate to inhibit the induction of IFN-β and ISRE promoters by IRF3 and IFN-β production. In addition, we reveal that HBZ enhances ISRE activation by IFN-α. We further show that HBZ enhances IRF7 and suppresses IRF3 activation by TBK1 and IKK+. We demonstrate that HBZ has no effect on virus-induced nuclear accumulation of IRF3, suggesting that it may inhibit IRF3 activity at a transcriptional level. We show that HBZ physically interacts with IRF7 and IKK but not with IRF3 or TBK1. Overall, our findings suggest that both HBZ and Tax1 are negative regulators of immediate early IFN-β innate immune responses, while HBZ but not Tax1 positively regulates the induction of IFN-α and downstream IFN-α signaling. [ABSTRACT FROM AUTHOR]

The human T-cell leukaemia virus type 1 and type 2 (HTLV-1/HTLV-2) antisense proteins HBZ and APH-2 play key roles in the HTLV lifecycles and persistence in the host. Nuclear Factors Associated with double-stranded RNA (NFAR) proteins NF90/110 function in the lifecycles of several viruses and participate in host innate immunity against infection and oncogenesis. Using GST pulldown and co-immunoprecipitation assays we demonstrate specific novel interactions between HBZ/APH-2 and NF90/110 and characterised the protein domains involved. Moreover we show that NF90/110 significantly enhance Tax mediated LTR activation, an effect that was abolished by HBZ but enhanced by APH-2. Additionally we found that HBZ and APH-2 modulate the promoter activity of survivin and are capable of antagonising NF110-mediated survivin activation. Thus interactions between HTLV antisense proteins and the NFAR protein family have an overall positive impact on HTLV infection. Hence NFARs may represent potential therapeutic targets in HTLV infected cells. [ABSTRACT FROM AUTHOR]

Human T-cell leukemia virus type 1 (HTLV-1) is the etiologic agent of adult T-cell leukemia (ATL). The viral regulatory protein Taxi plays a pivotal role in T-cell transformation and ATL development. Previous studies in our laboratory, using the yeast 2-hy-brid approach to screen a T-cell library for Taxi-interacting partners, identified the cellular Four and a Half Lim domain protein 3 (FHL3) as a possible Taxi-interacting candidate. FHL3 is a member of the FHL family of proteins, which function as transcrip-tional coactivators and cytoskeleton regulators and have a role in cancer progression and development. The aim of this study was to investigate the physical and functional interaction between Taxi and members of the FHL family of proteins. We show that Taxi and FHL3 interact both in vitro and in vivo. Furthermore, both FHL1 and -2 also interact with Taxi. We have demon-strated that FHL3 enhances Taxi-mediated activation of the viral long terminal repeat (LTR) without affecting basal activity and that FHL1 to -3 regulate NF-KB activation by Taxi in a cell-specific manner. In addition, we have found that the interaction be-tween Taxi and FHL1 to -3 affects the localization of these proteins, leading to their redistribution in cells. Taxi also affected FHL3 cytoskeleton function by increasing FHL3-mediated cell spreading. Overall, our results suggest that the interaction be-tween Taxi and the FHL family alters both the transactivating activity and the subcellular localization of Taxi and provide new insights into molecular mechanisms that underlie the oncogenic nature of this HTLV-1 protein. [ABSTRACT FROM AUTHOR]

An abstract of the research paper titled,"Physical and functional interaction between the Human T-cell leukemia virus type 1 (HTLV-1) Tax oncoprotein and the cellular LIM domain protein Four and a Half Lim domain protein 3 (FHL3)," is presented.

An abstract of the research paper titled,"Regulation of AP-1 activity by the Human T-cell leukemia virus type 2 (HTLV-2) APH-2 protein," is presented.

Background: In contrast with human T-cell leukemia virus type 1 (HTLV-1) that causes ATL (adult T-cell leukemia), HTLV-2 has not been causally linked to malignant disease. The minus strand of the HTLV genomes encode the regulatory proteins HTLV-1 bZIP factor (HBZ) for HTLV-1 and antisense protein of HTLV-2 (APH-2) for HTLV-2. Unlike the viral proteins Tax1 and Tax2, both HBZ and APH-2 are constitutively expressed in infected cells suggesting that they may play important roles in the pathogenesis of these viruses. To date, very little is known about the function of APH-2 except that it inhibits Tax2-mediated transcription of HTLV-2 genes. In the present study, we investigated the role of APH-2 in basal and Tax2B-mediated activation of the AP-1 pathway. Results: We demonstrate that, unlike HBZ, APH-2 stimulates basal AP-1 transcription by interacting with c-Jun and JunB through its non-conventional bZIP domain. In addition, when Tax2 and APH-2 are co-expressed, they physically interact in vivo and in vitro and APH-2 acts as an inhibitor of Tax2-mediated activation of AP-1 transcription. Conclusions: This report is the first to document that HTLV-2 can modulate the AP-1 pathway. Altogether our results reveal that, in contrast with HBZ, APH-2 regulates AP-1 activity in a Tax2-dependant manner. As the AP-1 pathway is involved in numerous cellular functions susceptible to affect the life cycle of the virus, these distinct biological properties between HBZ and APH-2 may contribute to the differential pathogenic potential of HTLV-1 and HTLV-2. [ABSTRACT FROM AUTHOR]

Background: The HIV-1 regulatory protein Rev, which is essential for viral replication, mediates the nuclear export of unspliced viral transcripts. Rev nuclear function requires active nucleocytoplasmic shuttling, and Rev nuclear import is mediated by the recognition of its Nuclear Localisation Signal (NLS) by multiple import factors, which include transportin and importin β. However, it remains unclear which nuclear import pathway(s) predominate in vivo, and the cellular environment that modulates Rev nucleocytoplasmic shuttling remains to be characterised. Results: In our study, we have identified the cellular protein HIC (Human I-mfa domain-Containing protein) as a novel interactor of HIV-1 Rev. We demonstrate that HIC selectively interferes with Rev NLS interaction with importin β and impedes its nuclear import and function, but does not affect Rev nuclear import mediated by transportin. Hence, the molecular determinants mediating Rev-NLS recognition by importin β and transportin appear to be distinct. Furthermore, we have employed HIC and M9 M, a peptide specifically designed to inhibit the transportinmediated nuclear import pathway, to characterise Rev nuclear import pathways within different cellular environments. Remarkably, we could show that in 293T, HeLa, COS7, Jurkat, U937, THP-1 and CEM cells, Rev nuclear import is cell type specific and alternatively mediated by transportin or importin β, in a mutually exclusive fashion. Conclusions: Rev cytoplasmic sequestration by HIC may represent a novel mechanism for the control of Rev function. These studies highlight that the multivalent nature of the Rev NLS for different import receptors enables Rev to adapt its nuclear trafficking strategy. [ABSTRACT FROM AUTHOR]

Abstract: The Human I-mfa domain-Containing protein, HIC, is a 246 amino acid protein that functions as a transcriptional regulator. Although the precise function of HIC remains to be clarified, the association of the HIC gene locus with myeloid neoplasms, its interactions with lymphotropic viruses such as EBV, HIV-1 and HTLV-1 and its expression in immune tissues suggest that HIC might have a modulatory role in immune cells. To further characterise the HIC functional relationship with the immune system, we sought to analyse the HIC gene expression profile in immune cells and to determine if immunomodulatory cytokines, such as interleukin (IL)-2, could regulate the expression of HIC mRNA. Relative quantitative real-time RT-PCR revealed that HIC mRNA is highly expressed in PBMCs and in various hematopoietic cell lines. The immunomodulatory cytokine IL-2 up-regulated HIC gene expression in PBMCs, CEM, MT-2 and U937 but markedly reduced HIC gene expression in Raji. Addition of cycloheximide indicated that the IL-2 effects were independent of de novo protein synthesis and that the HIC gene is a direct target of IL-2. Two cell lines (Jurkat and BJAB) displayed a distinct loss in HIC gene expression. However, when these cell lines were subjected to a combination of DNA methyltransferase and histone-deacetylase inhibitors, (5-aza-2-deoxycytidine and trichostatin A, respectively), HIC expression was de-repressed, indicating possible epigenetic control of HIC expression. Overall, our study describes that the immune expression of HIC is cell-specific, dynamic, and identifies the HIC gene as an IL-2 responsive gene. Furthermore, our de-repression studies support the hypothesis that HIC might represent a candidate tumor suppressor gene. Overall, this report provides new insights for a putative role of HIC in the modulation of immune and inflammatory responses and/or hematological malignancies. [Copyright &y& Elsevier]

Adult T-cell leukemia-lymphoma (ATLL) is a group of T-cell malignancies caused by infection with human T-lymphotropic virus type I (HTLV-I). Although the pathogenesis of ATLL remains incompletely understood, the viral regulatory protein Tax is centrally involved in cellular transformation. Here we describe the generation of HTLV-I Tax transgenic mice using the Lck proximal promoter to restrict transgene expression to developing thymocytes. After prolonged latency periods, transgenic mice developed diffuse large-cell lymphomas and leukemia with clinical, pathological and immunological features characteristic of acute ATLL. Transgenic mice were functionally immunocompromised and they developed opportunistic infections. Fulminant disease also developed rapidly in SCID mice after engraftment of lymphomatous cells from transgenic mice. Flow cytometry showed that the cells were CD4− and CD8−, but CD44+, CD25+ and cytoplasmic CD3+. This phenotype is indicative of a thymus-derived pre–T-cell phenotype, and disease development was associated with the constitutive activation of NF-κB. Our model accurately reproduces human disease and will provide a tool for analysis of the molecular events in transformation and for the development of new therapeutics. [ABSTRACT FROM AUTHOR]

An abstract of the research paper titled,"Stromal cells help ATL cells to escape from apoptosis induced by histone deacetylase inhibitors," is presented.