Publications by authors named "Erica Diani"

29 Publications

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Assessment of SARS-CoV-2 IgG and IgM antibody detection with a lateral flow immunoassay test.

Heliyon 2021 Oct 19;7(10):e08192. Epub 2021 Oct 19.

Department of Diagnostics and Public Health, Microbiology Section, University of Verona, Italy.

The dramatic impact of SARS-CoV-2 infection on the worldwide public health has elicited the rapid assessment of molecular and serological diagnostic methods. Notwithstanding the diagnosis of SARS-CoV-2 infection is based on molecular biology approaches including multiplex or singleplex real time RT-PCR, there is a real need for affordable and rapid serological methods to support diagnostics, and surveillance of infection spreading. In this study, we performed a diagnostic accuracy analysis of COVID-19 IgG/IgM rapid test cassette lateral flow immunoassay test (LFIA) assay. To do so, we analyzed different cohorts of blood samples obtained from 151 SARS-CoV-2 RT-PCR assay positive patients (group 1) and 51 SARS-CoV-2 RT-PCR assay negative patients (group 2) in terms of sensitivity, specificity, PPV, NPV and likelihood ratios. In addition, we challenged LFIA with plasma from 99 patients stored during 2015-2017 period. Our results showed that this LFIA detected SARS-CoV-2 IgM and/or IgG in 103 out of 151 (68.21%) samples of group 1, whereas no IgM and/or IgG detection was displayed both in the group 2 and in pre-pandemic samples. Interestingly, IgM and/or IgG positivity was detected in 86 out of 94 (91.49%) group 1 samples collected after 10 days from symptoms onset whereas only 17 out of 57 of group 1 samples obtained before day 10 were positive to SARS-CoV-2 specific antibodies. We also compared the performance of this LFIA test with respect to other four different LFIA assays in 40 serum samples from multiplex RT-PCR positive individuals. Within the limits of the study size, the results demonstrated that COVID-19 IgG/IgM rapid test cassette LFIA assay displayed valid performance in IgM and IgG detection when compared with the other four LFIA assays. Hence, this approach might be considered as an alternative point-of-care procedure for SARS-CoV-2 serological investigation.
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http://dx.doi.org/10.1016/j.heliyon.2021.e08192DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8525011PMC
October 2021

Cross-Immunization Against Respiratory Coronaviruses May Protect Children From SARS-CoV2: More Than a Simple Hypothesis?

Front Pediatr 2020 18;8:595539. Epub 2021 Jan 18.

Department of Diagnostic and Public Health, Unit of Microbiology, Verona University, Verona, Italy.

In January 2020, a new coronavirus was identified as responsible for a pandemic acute respiratory syndrome. The virus demonstrated a high infectious capability and not-neglectable mortality in humans. However, similarly to previous SARS and MERS, the new disease COVID-19 caused by SARS-CoV-2 seemed to relatively spare children and younger adults. Some hypotheses have been proposed to explain the phenomenon, including lower ACE2 expression in children, cross-immunization from measles/rubella/mumps and BCG-vaccination, as well as the integrity of respiratory mucosa. Herein, we hypothesize that an additional mechanism might contribute to children's relative protection from SARS-CoV-2, the cross-immunization conferred by previous exposures to other common respiratory coronaviruses. To support our hypothesis, we show a statistically significant similarity in genomic and protein sequences, including epitopes for B- and T-cell immunity, of SARS-CoV-2 and the other beta coronaviruses. Since these coronaviruses are highly diffused across pediatric populations, cross-reactive immunity might reasonably induce an at least partial protection from SARS-CoV-2 in children.
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http://dx.doi.org/10.3389/fped.2020.595539DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7849449PMC
January 2021

Prevalence of SARS-CoV-2 infection in the obstetric population before the hospital admission.

Minerva Ginecol 2020 12 24;72(6):429-430. Epub 2020 Sep 24.

Department of Obstetrics and Gynecology, University Hospital of Verona, University of Verona, Verona, Italy.

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http://dx.doi.org/10.23736/S0026-4784.20.04654-7DOI Listing
December 2020

Interferon gamma inducible protein 16 (IFI16) expression is reduced in mantle cell lymphoma.

Heliyon 2019 Nov 14;5(11):e02643. Epub 2019 Nov 14.

Department of Diagnostic and Public Health, Unit of Microbiology, University of Verona, Verona, Italy.

IFI16, member of the IFN-inducible PYHIN-200 gene family, modulates proliferation, survival and differentiation of different cell lineages. In particular, IFI16 expression, which is regulated during the differentiation of B cells, was recently studied in B-CLL as well. Here, we compared IFI16 expression in several lymphomas including Burkitt lymphoma, diffuse large B-cell lymphoma, follicular lymphoma, marginal zone lymphoma and mantle cell lymphoma with respect to normal cell counterparts. We observed that IFI16 expression was significantly deregulated only in mantle cell lymphoma (p < 0.05). Notably, IFI16 was associated with the expression of genes involved in interferon response, cell cycle, cell death and proliferation and, interestingly, lipid and glucose metabolism, suggesting that IFI16 deregulation might be associated with relevant changes in cell biology. In our group of mantle cell lymphoma samples a correlation between patient survival and IFI16 expression was not detected even though mantle cell lymphoma prognosis is known to be associated with cell proliferation. Altogether, these results suggest a complex relationship between IFI16 expression and MCL which needs to be analyzed in further studies.
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http://dx.doi.org/10.1016/j.heliyon.2019.e02643DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6893061PMC
November 2019

'High' antiretroviral deintensification strategy and cellular HIV DNA levels.

J Chemother 2020 Feb 6;32(1):49-51. Epub 2019 Dec 6.

Microbiology and Virology Section, Department of Diagnostic and Public Health, University of Verona, Verona, Italy.

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http://dx.doi.org/10.1080/1120009X.2019.1692996DOI Listing
February 2020

Immunovirological outcome and HIV-1 DNA decay in a small cohort of HIV-1-infected patients deintensificated from Abacavir/Lamivudine/Dolutegravir to Lamivudine plus Dolutegravir.

New Microbiol 2018 10 12;41(4):262-267. Epub 2018 Oct 12.

Microbiology and Virology Section, Department of Diagnostic and Public Health, University of Verona, Verona, Italy.

Combination abacavir/lamivudine/dolutegravir (ABC/3TC/DTG) is approved as a first-line treatment for antiretroviral naïve patients. This report investigated the immunovirological outcome and total HIV-1 DNA decay in a small cohort of naïve HIV-1-positive patients treated with this regimen. In the presence of viral suppression and increased lymphocyte T CD4+ cells, the quantitative analysis of total HIV-1 DNA content revealed a significant decay after 12 months of treatment. Subsequently, we deintensificated the treatment of these patients from (ABC/3TC/DTG) to lamivudine plus dolutegravir (3TC/DTG) after 12 months of virological suppression, as a strategy of "induction-maintenance" therapy. The analysis of HIV-1 RNA viral load, total HIV-1 DNA, CD4+ T lymphocyte count and CD8+ HLA-DR+ T lymphocyte percentage after a mean 3.5 months of therapy deintensification showed no significant difference with respect to data detected after 12 months of ABC/3TC/DTG treatment in the presence of continuous viral suppression. These results indicate that the deintensification of highly active antiretroviral therapy (HAART) from ABC/ 3TC/DTG to 3TC/DTG effectively controls HIV-1 replication and in the early period does not induce any significant variations of total HIV-1 DNA. This suggests that HAART deintensification might be proposed as a therapeutic evolution in the treatment of HIV-1 infection.
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October 2018

HIV-1-Associated Neurocognitive Disorders: Is HLA-C Binding Stability to β-Microglobulin a Missing Piece of the Pathogenetic Puzzle?

Front Neurol 2018 21;9:791. Epub 2018 Sep 21.

Department of Diagnostics and Public Health, University of Verona, Verona, Italy.

AIDS dementia complex (ADC) and HIV-associated neurocognitive disorders (HAND) are complications of HIV-1 infection. Viral infections are risk factors for the development of neurodegenerative disorders. Aging is associated with low-grade inflammation in the brain, i.e., the inflammaging. The molecular mechanisms linking immunosenescence, inflammaging and the pathogenesis of neurodegenerative disorders, such as Alzheimer's disease (AD) and Parkinson's disease, are largely unknown. ADC and HAND share some pathological features with AD and may offer some hints on the relationship between viral infections, neuroinflammation, and neurodegeneration. β-microglobulin (βm) is an important pro-aging factor that interferes with neurogenesis and worsens cognitive functions. Several studies published in the 80-90s reported high levels of βm in the cerebrospinal fluid of patients with ADC. High levels of βm have also been detected in AD. Inflammatory diseases in elderly people are associated with polymorphisms of the MHC-I locus encoding HLA molecules that, by associating with βm, contribute to cellular immunity. We recently reported that HLA-C, no longer associated with βm, is incorporated into HIV-1 virions, determining an increase in viral infectivity. We also documented the presence of HLA-C variants more or less stably linked to βm. These observations led us to hypothesize that some variants of HLA-C, in the presence of viral infections, could determine a greater release and accumulation of βm, which in turn, may be involved in triggering and/or sustaining neuroinflammation. ADC is the most severe form of HAND. To explore the role of HLA-C in ADC pathogenesis, we analyzed the frequency of HLA-C variants with unstable binding to βm in a group of patients with ADC. We found a higher frequency of unstable HLA-C alleles in ADC patients, and none of them was harboring stable HLA-C alleles in homozygosis. Our data suggest that the role of HLA-C variants in ADC/HAND pathogenesis deserves further studies. If confirmed in a larger number of samples, this finding may have practical implication for a personalized medicine approach and for developing new therapies to prevent HAND. The exploration of HLA-C variants as risk factors for AD and other neurodegenerative disorders may be a promising field of study.
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http://dx.doi.org/10.3389/fneur.2018.00791DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6160745PMC
September 2018

Duplex real-time polymerase chain reaction assay for the detection of human KIPyV and WUPyV in nasopharyngeal aspirate pediatric samples.

Mol Cell Probes 2018 08 5;40:13-18. Epub 2018 Jun 5.

Microbiology and Virology Unit, Department of Diagnostics and Public Health, University of Verona, Strada delle Grazie 8, 37134 Verona, Italy.

In this study, we describe a duplex real-time PCR assay for the simultaneous detection of KIPyV and WUPyV polyomaviruses based on TaqMan probes. This assay detected 500 copies/mL both for KIPyV and WUPyV in 100% of tested positive samples. We assessed this technique on 482 nasopharyngeal aspirate specimens from hospitalized pediatric patients with respiratory symptoms, previously analyzed with commercial multiplex assay for 16 major respiratory viruses. Our assay detected KIPyV genome in 15 out of 482 samples (3.1%) and WUPyV genome in 24 out of 482 samples (4.9%), respectively, and in three samples the coinfection of the two viruses was found. Interestingly, 29 out of 36 of samples with KIPyV and/or WUPyV infection exhibited a co-infection with one or more respiratory viruses confirming that KIPyV and WUPyV were often detected in association to other viral infections. Of note, KIPyV and WUPyV were detected singularly in 4 out of 15 cases and 3 out of 24 cases, respectively, suggesting a possible direct role of these viruses in the respiratory diseases. In conclusion, this method could be taken into account as an alternative technical approach to detect KIPyV and/or WUPyV in respiratory samples for epidemiological and diagnostic analyses.
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http://dx.doi.org/10.1016/j.mcp.2018.06.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7172048PMC
August 2018

IFI16 reduced expression is correlated with unfavorable outcome in chronic lymphocytic leukemia.

APMIS 2017 Jun 18;125(6):511-522. Epub 2017 May 18.

Department of Diagnostic and Public Health, Unit of Microbiology, University of Verona, Verona, Italy.

Chronic lymphocytic leukemia (CLL) is the most common leukemia in adults. Its clinical course is typically indolent; however, based on a series of pathobiological, clinical, genetic, and phenotypic parameters, patient survival varies from less than 5 to more than 20 years. In this paper, we show for the first time that the expression of the interferon-inducible DNA sensor IFI16, a member of the PYHIN protein family involved in proliferation inhibition and apoptosis regulation, is associated with the clinical outcome in CLL. We studied 99 CLLs cases by immunohistochemistry and 10 CLLs cases by gene expression profiling. We found quite variable degrees of IFI16 expression among CLLs cases. Noteworthy, we observed that a reduced IFI16 expression was associated with a very poor survival, but only in cases with ZAP70/CD38 expression. Furthermore, we found that IFI16 expression was associated with a specific gene expression signature. As IFI16 can be easily detected by immunohistochemistry or flow cytometry, it may become a part of phenotypic screening in CLL patients if its prognostic role is confirmed in independent series.
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http://dx.doi.org/10.1111/apm.12692DOI Listing
June 2017

Assessment of NS1 gene-specific real time quantitative TaqMan PCR for the detection of Human Bocavirus in respiratory samples.

Mol Cell Probes 2017 08 27;34:53-55. Epub 2017 Apr 27.

Microbiology and Virology Unit, Department of Pathology and Diagnostic, University of Verona, Strada delle Grazie 8, 37134 Verona, Italy.

Human Bocaviruses (HBoV) were associated with respiratory diseases. Here, we assessed a TaqMan-based PCR for the detection of all four HBoV subtype infections with a sensitivity up to 15 copies/reaction. To evaluate this assay on clinical samples, 178 nasopharyngeal aspirate specimens from pediatric cases were analyzed and HBoV genome was detected in 13 out of 178 patients with a viral load range between 1.6 × 10 and 9.4 × 10 copies/ml. These results indicated that this method could be used as an alternative technique for the diagnosis of HBoV infection.
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http://dx.doi.org/10.1016/j.mcp.2017.04.005DOI Listing
August 2017

A Luciferase Functional Quantitative Assay for Measuring NF-ĸB Promoter Transactivation Mediated by HTLV-1 and HTLV-2 Tax Proteins.

Methods Mol Biol 2017 ;1582:79-87

Section of Biology and Genetics, Department of Neurosciences, Biomedicine and Movement Sciences, University of Verona, Strata le Grazie 8, 37134, Verona, Italy.

HTLV-1 and HTLV-2 viruses express Tax transactivator proteins required for viral genome transcription and capable of transforming cells in vivo and in vitro. Although Tax oncogenic potential needs to be further elucidated, it is well established that Tax proteins activate, among others, transcription factors of the NF-ĸB family, which are involved in immune and inflammatory responses, cell growth, apoptosis, stress responses and oncogenesis. Here, we describe a reporter gene assay applied for quantitative analysis of Tax-dependent NF-ĸB activation. The procedure is based on co-transfection of two individual vectors containing the cDNA for firefly and Renilla luciferase enzymes and vectors expressing Tax proteins. The luciferase expression is driven by cis-NF-ĸB promoter regulatory elements responsive to Tax transactivating factor. This assay is particularly useful to investigate Tax influence on NF-ĸB activation mediated by viral or host factors.
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http://dx.doi.org/10.1007/978-1-4939-6872-5_6DOI Listing
February 2018

M48U1 and Tenofovir combination synergistically inhibits HIV infection in activated PBMCs and human cervicovaginal histocultures.

Sci Rep 2017 02 1;7:41018. Epub 2017 Feb 1.

Institut Galien Paris Sud, CNRS UMR 8612, Univ. Paris-Sud, Université Paris Saclay, 5 rue J-B. Clément, 92296, Châtenay-Malabry cedex, France.

Microbicides are considered a promising strategy for preventing human immunodeficiency virus (HIV-1) transmission and disease. In this report, we first analyzed the antiviral activity of the miniCD4 M48U1 peptide formulated in hydroxyethylcellulose (HEC) hydrogel in activated peripheral blood mononuclear cells (PBMCs) infected with R5- and X4-tropic HIV-1 strains. The results demonstrate that M48U1 prevented infection by several HIV-1 strains including laboratory strains, and HIV-1 subtype B and C strains isolated from the activated PBMCs of patients. M48U1 also inhibited infection by two HIV-1 transmitted/founder infectious molecular clones (pREJO.c/2864 and pTHRO.c/2626). In addition, M48U1 was administered in association with tenofovir, and these two antiretroviral drugs synergistically inhibited HIV-1 infection. In the next series of experiments, we tested M48U1 alone or in combination with tenofovir in HEC hydrogel with an organ-like structure mimicking human cervicovaginal tissue. We demonstrated a strong antiviral effect in absence of significant tissue toxicity. Together, these results indicate that co-treatment with M48U1 plus tenofovir is an effective antiviral strategy that may be used as a new topical microbicide to prevent HIV-1 transmission.
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http://dx.doi.org/10.1038/srep41018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5286506PMC
February 2017

Molecular characterization of HIV-1 Nef and ACOT8 interaction: insights from in silico structural predictions and in vitro functional assays.

Sci Rep 2016 Mar 1;6:22319. Epub 2016 Mar 1.

Department of Neurosciences, Biomedicine and Movement Sciences, University of Verona, Strada le Grazie 8, 37134 Verona, Italy.

HIV-1 Nef interacts with several cellular proteins, among which the human peroxisomal thioesterase 8 (ACOT8). This interaction may be involved in the endocytosis regulation of membrane proteins and might modulate lipid composition in membrane rafts. Nef regions involved in the interaction have been experimentally characterized, whereas structural details of the ACOT8 protein are unknown. The lack of structural information hampers the comprehension of the functional consequences of the complex formation during HIV-1 infection. We modelled, through in silico predictions, the ACOT8 structure and we observed a high charge complementarity between Nef and ACOT8 surfaces, which allowed the identification of the ACOT8 putative contact points involved in the interaction. The predictions were validated by in vitro assays through the development of ACOT8 deletion mutants. Coimmunoprecipitation and immunofluorescence analyses showed that ACOT8 Arg(45)-Phe(55) and Arg(86)-Pro(93) regions are involved in Nef association. In addition, K91S mutation abrogated the interaction with Nef, indicating that Lys(91) plays a key role in the interaction. Finally, when associated with ACOT8, Nef may be preserved from degradation. These findings improve the comprehension of the association between HIV-1 Nef and ACOT8, helping elucidating the biological effect of their interaction.
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http://dx.doi.org/10.1038/srep22319DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4772117PMC
March 2016

HTLV-1 Tax protein recruitment into IKKε and TBK1 kinase complexes enhances IFN-I expression.

Virology 2015 Feb 19;476:92-99. Epub 2014 Dec 19.

Department of Life and Reproduction Sciences, Section of Biology and Genetics, University of Verona, Strada le Grazie 8, 37134 Verona, Italy. Electronic address:

The Tax protein expressed by human T-cell leukemia virus type 1 (HTLV-1) plays a pivotal role in the deregulation of cellular pathways involved in the immune response, inflammation, cell survival, and cancer. Many of these effects derive from Tax multiple interactions with host factors, including the subunits of the IKK-complex that are required for NF-κB activation. IKKɛ and TBK1 are two IKK-related kinases that allow the phosphorylation of interferon regulatory factors that trigger IFN type I gene expression. We observed that IKKɛ and TBK1 recruit Tax into cellular immunocomplexes. We also found that TRAF3, which regulates cell receptor signaling effectors, forms complexes with Tax. Transactivation analyses revealed that expression of Tax, in presence of IKKɛ and TBK1, enhances IFN-β promoter activity, whereas the activation of NF-κB promoter is not modified. We propose that Tax may be recruited into the TBK1/IKKɛ complexes as a scaffolding-adaptor protein that enhances IFN-I gene expression.
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http://dx.doi.org/10.1016/j.virol.2014.12.005DOI Listing
February 2015

New insights into functional roles of the polypyrimidine tract-binding protein.

Int J Mol Sci 2013 Nov 20;14(11):22906-32. Epub 2013 Nov 20.

Department of Life and Reproduction Sciences, Section of Biology and Genetics, University of Verona, Strada Le Grazie 8, 37134 Verona, Italy.

Polypyrimidine Tract Binding Protein (PTB) is an intensely studied RNA binding protein involved in several post-transcriptional regulatory events of gene expression. Initially described as a pre-mRNA splicing regulator, PTB is now widely accepted as a multifunctional protein shuttling between nucleus and cytoplasm. Accordingly, PTB can interact with selected RNA targets, structural elements and proteins. There is increasing evidence that PTB and its paralog PTBP2 play a major role as repressors of alternatively spliced exons, whose transcription is tissue-regulated. In addition to alternative splicing, PTB is involved in almost all steps of mRNA metabolism, including polyadenylation, mRNA stability and initiation of protein translation. Furthermore, it is well established that PTB recruitment in internal ribosome entry site (IRES) activates the translation of picornaviral and cellular proteins. Detailed studies of the structural properties of PTB have contributed to our understanding of the mechanism of RNA binding by RNA Recognition Motif (RRM) domains. In the present review, we will describe the structural properties of PTB, its paralogs and co-factors, the role in post-transcriptional regulation and actions in cell differentiation and pathogenesis. Defining the multifunctional roles of PTB will contribute to the understanding of key regulatory events in gene expression.
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http://dx.doi.org/10.3390/ijms141122906DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3856098PMC
November 2013

Polymorphism -2604G>A variants in TLR4 promoter are associated with different gene expression level in peripheral blood of atherosclerotic patients.

J Hum Genet 2013 Dec 10;58(12):812-4. Epub 2013 Oct 10.

Section of Neurology, Department of Neurological and Movement Sciences, University of Verona, Verona, Italy.

Toll-like receptor-4 (TLR4) is a primary receptor of the innate immune reaction and compelling evidence demonstrates its involvement in the pathogenesis of atherosclerosis and stroke. TLR4 is constitutively expressed on monocytes and endothelial cells; it is highly expressed in atherosclerotic plaques and in peripheral blood of patients after ischemic stroke. Polymorphisms in the promoter region that alter the transcriptional regulation of this gene may represent genetic risk factors involved in the predisposition to atherosclerotic disease. In this study we investigated the effect on TLR4 gene expression of three polymorphisms in the upstream regulatory region at positions -1607T>C/rs10759932, -2026A>G/rs1927914 and -2604G>A/rs10759931 in peripheral blood of atherosclerotic patients. RNA from individuals homozygous for the -2604A allele showed a lower expression of the gene when compared to patients carrying the counterparts GG+GA. Electrophoretic mobility shift assays showed differences in the electrophoretic mobility of the DNA-nuclear protein complexes formed by the G>A variants, suggesting that the two alleles differ in their binding affinity to transcriptional factors.
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http://dx.doi.org/10.1038/jhg.2013.98DOI Listing
December 2013

Highlights on distinctive structural and functional properties of HTLV Tax proteins.

Front Microbiol 2013 Sep 9;4:271. Epub 2013 Sep 9.

Department of Life and Reproduction Sciences, University of Verona Verona, Italy.

Human T cell leukemia viruses (HTLVs) are complex human retroviruses of the Deltaretrovirus genus. Four types have been identified thus far, with HTLV-1 and HTLV-2 much more prevalent than HTLV-3 or HTLV-4. HTLV-1 and HTLV-2 possess strictly related genomic structures, but differ significantly in pathogenicity, as HTLV-1 is the causative agent of adult T cell leukemia and of HTLV-associated myelopathy/tropical spastic paraparesis, whereas HTLV-2 is not associated with neoplasia. HTLVs code for a protein named Tax that is responsible for enhancing viral expression and drives cell transformation. Much effort has been invested to dissect the impact of Tax on signal transduction pathways and to identify functional differences between the HTLV Tax proteins that may explain the distinct oncogenic potential of HTLV-1 and HTLV-2. This review summarizes our current knowledge of Tax-1 and Tax-2 with emphasis on their structure, role in activation of the NF-κB (nuclear factor kappa-B) pathway, and interactions with host factors.
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http://dx.doi.org/10.3389/fmicb.2013.00271DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3766827PMC
September 2013

Ubiquitination and sumoylation of the HTLV-2 Tax-2B protein regulate its NF-κB activity: a comparative study with the HTLV-1 Tax-1 protein.

Retrovirology 2012 Dec 7;9:102. Epub 2012 Dec 7.

Department of Life and Reproduction Sciences, Section of Biology and Genetics, University of Verona, Strada Le Grazie 8, 37134, Verona, Italy.

Background: Retroviruses HTLV-1 and HTLV-2 have homologous genomic structures but differ significantly in pathogenicity. HTLV-1 is associated with Adult T cell Leukemia (ATL), whereas infection by HTLV-2 has no association with neoplasia. Transformation of T lymphocytes by HTLV-1 is linked to the capacity of its oncoprotein Tax-1 to alter cell survival and cell cycle control mechanisms. Among these functions, Tax-1-mediated activation of cellular gene expression via the NF-κB pathway depends on Tax-1 post-translational modifications by ubiquitination and sumoylation. The Tax-2 protein of HTLV-2B (Tax-2B) is also modified by ubiquitination and sumoylation and activates the NF-κB pathway to a level similar to that of Tax-1. The present study aims to understand whether ubiquitination and sumoylation modifications are involved in Tax-2B-mediated activation of the NF-κB pathway.

Results: The comparison of Tax-1 and Tax-2B lysine to arginine substitution mutants revealed conserved patterns and levels of ubiquitination with notable difference in the lysine usage for sumoylation. Neither Tax-1 nor Tax-2B ubiquitination and sumoylation deficient mutants could activate the NF-κB pathway and fusion of ubiquitin or SUMO-1 to the C-terminus of the ubiquitination and sumoylation deficient Tax-2B mutant strikingly restored transcriptional activity. In addition, ubiquitinated forms of Tax-2B colocalized with RelA and IKKγ in prominent cytoplasmic structures associated with the Golgi apparatus, whereas colocalization of Tax-2B with the RelA subunit of NF-κB and the transcriptional coactivator p300 in punctate nuclear structures was dependent on Tax-2B sumoylation, as previously observed for Tax-1.

Conclusions: Both Tax-1 and Tax-2 activate the NF-κB pathway via similar mechanisms involving ubiquitination and sumoylation. Therefore, the different transforming potential of HTLV-1 and HTLV-2 is unlikely to be related to different modes of activation of the canonical NF-κB pathway.
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http://dx.doi.org/10.1186/1742-4690-9-102DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3543174PMC
December 2012

Comparison of the Genetic Organization, Expression Strategies and Oncogenic Potential of HTLV-1 and HTLV-2.

Leuk Res Treatment 2012 29;2012:876153. Epub 2011 Dec 29.

Department of Oncology and Surgical Sciences, The University of Padova, 35128 Padova, Italy.

Human T cell leukemia virus types 1 and 2 (HTLV-1 and HTLV-2) are genetically related complex retroviruses that are capable of immortalizing human T-cells in vitro and establish life-long persistent infections in vivo. In spite of these apparent similarities, HTLV-1 and HTLV-2 exhibit a significantly different pathogenic potential. HTLV-1 is recognized as the causative agent of adult T-cell leukemia/lymphoma (ATLL) and tropical spastic paraparesis/HTLV-1-associated myelopathy (TSP/HAM). In contrast, HTLV-2 has not been causally linked to human malignancy, although it may increase the risk of developing inflammatory neuropathies and infectious diseases. The present paper is focused on the studies aimed at defining the viral genetic determinants of the pathobiology of HTLV-1 and HTLV-2 through a comparison of the expression strategies and functional properties of the different gene products of the two viruses.
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http://dx.doi.org/10.1155/2012/876153DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3504254PMC
December 2012

Transcriptional regulation of the human Raver2 ribonucleoprotein gene.

Gene 2012 Feb 28;493(2):243-52. Epub 2011 Nov 28.

Department of Life and Reproduction Sciences, Section of Biology and Genetics, University of Verona, Strada le Grazie 8, 37134 Verona, Italy.

Raver2 is a putative modulator of the activity of the polypyrimidine-tract binding protein (PTB), one of the most intensively studied splicing repressors. Little is known about Raver2 expression, and all current data is from mice where it shows tissue specificity. In the present study, by comparing Raver2 transcript expression in human and mouse tissues, we found that human Raver2 is ubiquitously expressed in adult tissues. In order to investigate human Raver2 transcription regulation, we identified and characterized a putative promoter region in a 1000bp region upstream of the transcription starting site of the gene. Dual luciferase reporter assays demonstrated that this region had promoter activity conferred by the first 160bp. By mutagenic analyses of putative cis-acting regulatory sequences, we identified an individual site that decreased the promoter activity by up to 40% when mutated. Together, our results suggest that regulation of human Raver2 expression involves TATA-less transcriptional activity.
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http://dx.doi.org/10.1016/j.gene.2011.11.036DOI Listing
February 2012

Low penetrance and effect on protein secretion of LGI1 mutations causing autosomal dominant lateral temporal epilepsy.

Epilepsia 2011 Jul 19;52(7):1258-64. Epub 2011 Apr 19.

Department of Neurological Sciences, University of Rome Sapienza, Rome, Italy.

Purpose: To describe the clinical and genetic findings of four families with autosomal dominant lateral temporal epilepsy.

Methods: A personal and family history was obtained from each affected and unaffected subject along with a physical and neurologic examination. Routine electroencephalography and magnetic resonance imaging (MRI) studies were performed in almost all patients. DNAs from family members were screened for LGI1 mutations. The effects of mutations on Lgi1 protein secretion were determined in transfected culture cells.

Key Findings: The four families included a total of 11 patients (two deceased), six of whom had lateral temporal epilepsy with auditory aura. Age at onset was in the second decade of life; seizures were well controlled by antiepileptic treatment and MRI studies were normal. We found two pathogenic LGI1 mutations with uncommonly low penetrance: the R136W mutation, previously detected in a sporadic case with telephone-induced partial seizures, gave rise to the epileptic phenotype in three of nine mutation carriers in one family; the novel C179R mutation caused epilepsy in an isolated patient from a family where the mutation segregated. Another novel pathogenic mutation, I122T, and a nonsynonymous variant, I359V, were found in the two other families. Protein secretion tests showed that the R136W and I122T mutations inhibited secretion of the mutant proteins, whereas I359V had no effect on protein secretion; C179R was not tested, because of its predictable effect on protein folding.

Significance: These findings suggest that some LGI1 mutations may have a weak penetrance in families with complex inheritance pattern, or isolated patients, and that the protein secretion test, together with other predictive criteria, may help recognize pathogenic LGI1 mutations.
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http://dx.doi.org/10.1111/j.1528-1167.2011.03071.xDOI Listing
July 2011

Drug resistant ADLTE and recurrent partial status epilepticus with dysphasic features in a family with a novel LGI1mutation: electroclinical, genetic, and EEG/fMRI findings.

Epilepsia 2009 Nov 22;50(11):2481-6. Epub 2009 Jun 22.

Department of Neurological Sciences, University of Rome, Rome, Italy.

Purpose: We characterized a family with autosomal dominant lateral temporal epilepsy (ADLTE) whose proband presented uncommon electroclinical findings such as drug-resistant seizures and recurrent episodes of status epilepticus with dysphasic features.

Methods: The electroclinical characteristics and LGI1 genotype were defined in the family. In the proband, the ictal pattern was documented during video-EEG monitoring and epileptic activity was mapped by EEG/fMRI.

Results: The affected members who were studied had drug-resistant seizures. In the proband, seizures with predominant dysphasic features often occurred as partial status epilepticus. The video-EEG-documented ictal activity and fMRI activation clearly indicated the elective involvement of the left posterior lateral temporal cortex. Sequencing of LGI1 exons revealed a heterozygous c.367G>A mutation in exon 4, resulting in a Glu123Lys substitution in the protein sequence.

Conclusions: The uncommon clinical pattern (high seizure frequency, drug-resistance) highlights the variability of the ADLTE phenotype and extends our knowledge of the clinical spectrum associated with LGI1 mutations.
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http://dx.doi.org/10.1111/j.1528-1167.2009.02181.xDOI Listing
November 2009

Characterization and functional analysis of cis-acting elements of the human farnesyl diphosphate synthetase (FDPS) gene 5' flanking region.

Genomics 2009 Mar 12;93(3):227-34. Epub 2008 Dec 12.

Department of Mother and Child, Biology and Genetics, University of Verona, Verona, Italy.

Farnesyl diphosphate synthetase (FDPS) is a key enzyme in the isoprenoid pathway responsible for cholesterol biosynthesis, post-translational protein modifications and synthesis of steroid hormones, whose expression is regulated by phorbol esters and polyunsaturated fatty acids. Genomic comparison of the 5' upstream sequence of the FDPS genes identifies conserved binding sites for NF-Y, SP1, SRE3, and YY1 regulatory elements in rat, mouse, dog and chimpanzee. Two additional specific consensus sequences, upstream of the core promoter that had not been analysed previously, are shared only by human and chimpanzee genomes. The work presented here aimed at characterizing these genomic sequence elements in the human FDPS promoter region and their contribution to gene expression. We have characterized functionally the minimal basal promoter of the human FDPS gene by means of deletion mutants and we have identified two cis-acting elements which modulate the FDPS gene expression and are recognized by Pax5 and OCT-1 transcription factors.
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http://dx.doi.org/10.1016/j.ygeno.2008.11.002DOI Listing
March 2009

A novel loss-of-function LGI1 mutation linked to autosomal dominant lateral temporal epilepsy.

Arch Neurol 2008 Jul;65(7):939-42

Muscular and Neurodegenerative Diseases Unit, Institute G. Gaslini, University of Genoa, Genoa, Italy.

Background: Mutations responsible for autosomal dominant lateral temporal epilepsy have been found in the leucine-rich, glioma-inactivated 1 (LGI1) gene.

Objectives: To describe the clinical and genetic findings in a family with autosomal dominant lateral temporal epilepsy and to determine the functional effects of a novel LGI1 mutation in culture cells.

Design: Clinical, genetic, and functional investigations.

Setting: University hospital and laboratory.

Patients: An Italian family with autosomal dominant lateral temporal epilepsy.

Main Outcome Measure: Mutation analysis.

Results: A novel LGI1 mutation, c.365T>A (Ile122Lys), segregating with the disease was identified. The mutant Lgi1 protein was not secreted by culture cells.

Conclusion: Our data provide further evidence that mutations in LGI1 hamper secretion of the Lgi1 protein, thereby precluding its normal function.
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http://dx.doi.org/10.1001/archneur.65.7.939DOI Listing
July 2008

Familial epilepsy and developmental dysphasia: description of an Italian pedigree with autosomal dominant inheritance and screening of candidate loci.

Epilepsy Res 2008 Jul 27;80(1):9-17. Epub 2008 May 27.

Department of Neurosciences, Division of Neurology, Via Altura 3, Bellaria Hospital, 40139 Bologna, Italy.

Purpose: To describe a familial epileptic condition combining a peculiar electro-clinical pattern with developmental language dysfunction in a large Italian kindred.

Methods: We studied the clinical and neurophysiological features of a 4-generation family with 10 affected members (3 deceased). We also analysed in 7 affected and 7 healthy members microsatellite markers for 51 candidate loci for epilepsy, including 42 loci containing ion channel genes expressed in the brain, as well as the SPCH1 and SRPX2 loci.

Results: Five of the seven living affected members (aged 20-58 years) had the full phenotype (seizures, EEG epileptiform abnormalities and dysphasia). The language dysfunction was the first symptom, becoming evident since the period of language development and mainly consisting of phonemic and syntactic paraphasias, difficulty of expression and reduced verbal fluency. The seizures had their onset between 2 and 23 years and were reported as epileptic falls (4) associated or not with myoclonic features, absences (3), tonic-clonic (1) and complex partial seizures (1). The seizures were easily controlled by antiepileptic treatment in all patients except one. In the five patients with a good response of seizures to treatment, the EEG tracings showed the coexistence of focal and generalized epileptiform abnormalities; in the refractory patient the interictal EEG demonstrated bilateral asynchronous fronto-temporal paroxysms with left predominance and ictal SEEG recording suggested a multifocal origin of the discharges. MRI of the brain was normal in all patients. Linkage analysis provided negative LOD scores for all the investigated loci.

Conclusion: We have described a novel familial pattern of epilepsy and developmental dysphasia which is not genetically linked to epilepsy or speech disorder loci, as documented by a candidate-gene linkage approach.
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http://dx.doi.org/10.1016/j.eplepsyres.2008.03.014DOI Listing
July 2008

Autosomal dominant lateral temporal epilepsy: absence of mutations in ADAM22 and Kv1 channel genes encoding LGI1-associated proteins.

Epilepsy Res 2008 Jul 28;80(1):1-8. Epub 2008 Apr 28.

CNR-Institute of Neurosciences, Section of Padua, Padova, Italy.

Mutations in the LGI1 gene are linked to autosomal dominant lateral temporal epilepsy (ADTLE) in about half of the families tested, suggesting that ADLTE is genetically heterogeneous. Recently, the Lgi1 protein has been found associated with different protein complexes and two distinct molecular mechanisms possibly underlying ADLTE have been hypothesized: the one recognizes Lgi1 as a novel subunit of the presynaptic Kv1 potassium channel implicated in the regulation of channel inactivation, the other suggests that Lgi1 acts as a ligand that selectively binds to the postsynaptic receptor ADAM22, thereby regulating the glutamate-AMPA neurotransmission. Both mechanisms imply that LGI1 mutations result in alteration of synaptic currents, though of different types. Since their protein products have been found associated with Lgi1, the Kv1 channel subunit genes KCNA1, KCNA4, and KCNAB1 and ADAM22 can be considered strong candidates for ADLTE. We sequenced their coding exons and flanking splice sites in the probands of 9 carefully ascertained ADLTE families negative for LGI1 mutations. We failed to detect any mutation segregating with the disease, but identified several previously unreported polymorphisms. An association study of four non-synonymous variants (three found in ADAM22, one in KCNA4) in a population of 104 non-familial lateral temporal epilepsy cases did not show any modification of susceptibility to this disorder. Altogether, our results suggest that neither ADAM22 nor any of the three Kv1 channel genes are major causative genes for ADLTE.
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http://dx.doi.org/10.1016/j.eplepsyres.2008.03.001DOI Listing
July 2008

Analysis of LGI1 promoter sequence, PDYN and GABBR1 polymorphisms in sporadic and familial lateral temporal lobe epilepsy.

Neurosci Lett 2008 May 4;436(1):23-6. Epub 2008 Mar 4.

CNR-Institute of Neurosciences, Section of Padua, Padova, Italy.

Autosomal dominant lateral temporal epilepsy (ADTLE) is a genetically transmitted epileptic syndrome characterized by focal seizures with predominant auditory symptoms likely originating from the lateral region of the temporal lobe. Mutations in coding region or exon splice sites of the leucine-rich, glioma-inactivated 1 (LGI1) gene account for about 50% of ADLTE families. De novo LGI1 mutations of the same kind have also been found in about 2.5% of non-familial cases with idiopathic partial epilepsy with auditory features (IPEAF). In both conditions, mutations in the LGI1 promoter region have not been reported. We sequenced the minimal promoter region of LGI1 in the probands of 16 ADLTE families and in 104 sporadic IPEAF patients and no mutations clearly linked to the disease were found. However, two polymorphisms, -500G>A and -507G>A, with potential functional implications were identified and analysed in the cohort of sporadic IPEAF patients but their frequencies did not differ from those found in a control population of similar age, gender and geographic origin. We also analysed in our study population the GABA(B) receptor 1 c.1465G>A and the prodynorphin promoter 68-bp repeat polymorphisms, previously associated with temporal lobe epilepsy. None of these polymorphisms showed a significant association with IPEAF, whereas a tendency towards association with the prodynorphin low expression (L) alleles was found in the small group of ADLTE index cases, in agreement with previous studies suggesting that this polymorphism is a susceptibility factor in familial forms of temporal lobe epilepsy.
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http://dx.doi.org/10.1016/j.neulet.2008.02.045DOI Listing
May 2008

Familial mesial temporal lobe epilepsy (FMTLE) : a clinical and genetic study of 15 Italian families.

J Neurol 2008 Jan 21;255(1):16-23. Epub 2007 Nov 21.

Epilepsy Center, Department of Neurological Sciences Federico II University, Via Pansini 5, 80131, Napoli, Italy.

Introduction: Familial mesial temporal lobe epilepsy (FMTLE) is characterized by prominent psychic and autonomic seizures, often without hippocampal sclerosis (HS) or a previous history of febrile seizures (FS), and good prognosis. The genetics of this condition is largely unknown.We present the electroclinical and genetic findings of 15 MTLE Italian families.

Patients And Methods: FMTLE was defined when two or more first-degree relatives had epilepsy suggesting a mesial temporal lobe origin. The occurrence of seizures with auditory auras was considered an exclusion criterion. Patients underwent video-EEG recordings, 1.5-Tesla MRI particularly focused on hippocampal analysis, and neuropsychological evaluation. Genetic study included genotyping and linkage analysis of candidate loci at 4q, 18q, 1q, and 12q as well as screening for LGI1/Epitempin mutations.

Results: Most of the families showed an autosomal dominant inheritance pattern with incomplete penetrance. Fifty-four (32 F) affected individuals were investigated. Twenty-one (38.8 %) individuals experienced early FS. Forty-eight individuals fulfilled the criteria for MTLE. Epigastric/visceral sensation (72.9 %) was the most common type of aura, followed by psychic symptoms (35.4 %), and déjà vu (31.2 %). HS occurred in 13.8% of individuals, three of whom belonged to the same family. Prognosis of epilepsy was generally good. Genetic study failed to show LGI1/Epitempin mutations or significative linkage to the investigated loci.

Discussion: FMTLE may be a more common than expected condition, clinically and genetically heterogeneous. Some of the reported families, grouped on the basis of a specific aura, may represent an interesting subgroup on whom to focus future linkage studies.
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http://dx.doi.org/10.1007/s00415-007-0653-1DOI Listing
January 2008
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