RESUMEN
Cell-cell fusion proteins are essential in development. Here we show that the C. elegans cell-cell fusion protein EFF-1 is structurally homologous to viral class II fusion proteins. The 2.6 Å crystal structure of the EFF-1 trimer displays the same 3D fold and quaternary conformation of postfusion class II viral fusion proteins, although it lacks a nonpolar "fusion loop," indicating that it does not insert into the target membrane. EFF-1 was previously shown to be required in both cells for fusion, and we show that blocking EFF-1 trimerization blocks the fusion reaction. Together, these data suggest that whereas membrane fusion driven by viral proteins entails leveraging of a nonpolar loop, EFF-1-driven fusion of cells entails trans-trimerization such that transmembrane segments anchored in the two opposing membranes are brought into contact at the tip of the EFF-1 trimer to then, analogous to SNARE-mediated vesicle fusion, zip the two membranes into one.
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Proteínas de Caenorhabditis elegans/química , Glicoproteínas de Membrana/química , Secuencia de Aminoácidos , Animales , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Fusión Celular , Cristalografía por Rayos X , Evolución Molecular , Células Gigantes/metabolismo , Fusión de Membrana , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Modelos Moleculares , Datos de Secuencia Molecular , Mutagénesis , Polimerizacion , Estructura Terciaria de Proteína , Alineación de Secuencia , Proteínas Virales de Fusión/química , Proteínas Virales de Fusión/genética , Proteínas Virales de Fusión/metabolismoRESUMEN
The liver is an organ with strong regenerative capacity, yet primary hepatocytes have a low amplification potential in vitro, a major limitation for the cell-based therapy of liver disorders and for ex vivo biological screens. Induced pluripotent stem cells (iPSCs) may help to circumvent this obstacle but often harbor genetic and epigenetic abnormalities, limiting their potential. Here, we describe the pharmacological induction of proliferative human hepatic progenitor cells (HPCs) through a cocktail of growth factors and small molecules mimicking the signaling events involved in liver regeneration. Human HPCs from healthy donors and pediatric patients proliferated vigorously while maintaining their genomic stability and could be redifferentiated in vitro into metabolically competent cells that supported the replication of hepatitis B and delta viruses. Redifferentiation efficiency was boosted by three-dimensional culture. Finally, transcriptome analysis showed that HPCs were more closely related to mature hepatocytes than iPSC-derived hepatocyte-like cells were. Conclusion: HPC induction holds promise for a variety of applications such as ex vivo disease modeling, personalized drug testing or metabolic studies, and development of a bioartificial liver.
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Técnicas de Cultivo de Célula , Medios de Cultivo/química , Hepatocitos/fisiología , Hígado/citología , Células Madre , Animales , Estudios de Casos y Controles , Masculino , Ratones Endogámicos NOD , Cultivo Primario de CélulasRESUMEN
Hantaviruses are zoonotic viruses transmitted to humans by persistently infected rodents, giving rise to serious outbreaks of hemorrhagic fever with renal syndrome (HFRS) or of hantavirus pulmonary syndrome (HPS), depending on the virus, which are associated with high case fatality rates. There is only limited knowledge about the organization of the viral particles and in particular, about the hantavirus membrane fusion glycoprotein Gc, the function of which is essential for virus entry. We describe here the X-ray structures of Gc from Hantaan virus, the type species hantavirus and responsible for HFRS, both in its neutral pH, monomeric pre-fusion conformation, and in its acidic pH, trimeric post-fusion form. The structures confirm the prediction that Gc is a class II fusion protein, containing the characteristic ß-sheet rich domains termed I, II and III as initially identified in the fusion proteins of arboviruses such as alpha- and flaviviruses. The structures also show a number of features of Gc that are distinct from arbovirus class II proteins. In particular, hantavirus Gc inserts residues from three different loops into the target membrane to drive fusion, as confirmed functionally by structure-guided mutagenesis on the HPS-inducing Andes virus, instead of having a single "fusion loop". We further show that the membrane interacting region of Gc becomes structured only at acidic pH via a set of polar and electrostatic interactions. Furthermore, the structure reveals that hantavirus Gc has an additional N-terminal "tail" that is crucial in stabilizing the post-fusion trimer, accompanying the swapping of domain III in the quaternary arrangement of the trimer as compared to the standard class II fusion proteins. The mechanistic understandings derived from these data are likely to provide a unique handle for devising treatments against these human pathogens.
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Orthobunyavirus/química , Orthohantavirus/química , Proteínas del Envoltorio Viral/química , Animales , Cristalografía , Glicoproteínas/química , Humanos , Conformación Proteica , Relación Estructura-Actividad , Resonancia por Plasmón de SuperficieAsunto(s)
Hepatitis C Crónica/epidemiología , Hepatitis D Crónica/epidemiología , Virus de la Hepatitis Delta/aislamiento & purificación , Necrosis Hepática Masiva/epidemiología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Niño , Femenino , Hepacivirus/aislamiento & purificación , Anticuerpos Antihepatitis/aislamiento & purificación , Virus de la Hepatitis B/aislamiento & purificación , Hepatitis B Crónica/diagnóstico , Hepatitis B Crónica/virología , Hepatitis C Crónica/complicaciones , Hepatitis C Crónica/diagnóstico , Hepatitis C Crónica/virología , Hepatitis D Crónica/complicaciones , Hepatitis D Crónica/diagnóstico , Hepatitis D Crónica/virología , Virus de la Hepatitis Delta/genética , Virus de la Hepatitis Delta/inmunología , Humanos , Pueblos Indígenas/estadística & datos numéricos , Masculino , Necrosis Hepática Masiva/diagnóstico , Necrosis Hepática Masiva/virología , Persona de Mediana Edad , ARN Viral/aislamiento & purificación , Venezuela/epidemiología , Adulto JovenRESUMEN
UNLABELLED: The structure of adenovirus outer capsid was revealed recently at 3- to 4-Å resolution (V. Reddy, S. Natchiar, P. Stewart, and G. Nemerow, Science 329:1071-1075, 2010, http://dx.doi.org/10.1126/science.1187292); however, precise details on the function and biochemical and structural features for the inner core still are lacking. Protein V is one the most important components of the adenovirus core, as it links the outer capsid via association with protein VI with the inner DNA core. Protein V is a highly basic protein that strongly binds to DNA in a nonspecific manner. We report the expression of a soluble protein V that exists in monomer-dimer equilibrium. Using reversible cross-linking affinity purification in combination with mass spectrometry, we found that protein V contains multiple DNA binding sites. The binding sites from protein V mediate heat-stable nucleic acid associations, with some of the binding sites possibly masked in the virus by other core proteins. We also demonstrate direct interaction between soluble proteins V and VI, thereby revealing the bridging of the inner DNA core with the outer capsid proteins. These findings are consistent with a model of nucleosome-like structures proposed for the adenovirus core and encapsidated DNA. They also suggest an additional role for protein V in linking the inner nucleic acid core with protein VI on the inner capsid shell. IMPORTANCE: Scant knowledge exists of how the inner core of adenovirus containing its double-stranded DNA (dsDNA) genome and associated proteins is organized. Here, we report a purification scheme for a recombinant form of protein V that allowed analysis of its interactions with the nucleic acid core region. We demonstrate that protein V exhibits stable associations with dsDNA due to the presence of multiple nucleic acid binding sites identified both in the isolated recombinant protein and in virus particles. As protein V also binds to the membrane lytic protein VI molecules, this core protein may serve as a bridge from the inner dsDNA core to the inner capsid shell.
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Adenoviridae/genética , Adenoviridae/metabolismo , ADN Viral/metabolismo , Proteínas del Núcleo Viral/genética , Proteínas del Núcleo Viral/metabolismo , Proteínas Virales/genética , Proteínas Virales/metabolismo , Secuencia de Aminoácidos , Secuencia de Bases , Sitios de Unión/genética , Cápside/metabolismo , Proteínas de la Cápside/genética , Proteínas de la Cápside/metabolismo , ADN Viral/genética , Datos de Secuencia Molecular , Unión Proteica/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Virión/genética , Virión/metabolismoRESUMEN
SARS-CoV-2 Omicron subvariants with increased transmissibility and immune evasion are spreading globally with alarming persistence. Whether the mutations and evolution of spike (S) Omicron subvariants alter the viral hijacking of human TMPRSS2 for viral entry remains to be elucidated. This is particularly important to investigate because of the large number and diversity of mutations of S Omicron subvariants reported since the emergence of BA.1. Here we report that human TMPRSS2 is a molecular determinant of viral entry for all the Omicron clinical isolates tested in human lung cells, including ancestral Omicron subvariants (BA.1, BA.2, BA.5), contemporary Omicron subvariants (BQ.1.1, XBB.1.5, EG.5.1) and currently circulating Omicron BA.2.86. First, we used a co-transfection assay to demonstrate the endoproteolytic cleavage by TMPRSS2 of spike Omicron subvariants. Second, we found that N-0385, a highly potent TMPRSS2 inhibitor, is a robust entry inhibitor of virus-like particles harbouring the S protein of Omicron subvariants. Third, we show that N-0385 exhibits nanomolar broad-spectrum antiviral activity against live Omicron subvariants in human Calu-3 lung cells and primary patient-derived bronchial epithelial cells. Interestingly, we found that N-0385 is 10-20 times more potent than the repositioned TMPRSS2 inhibitor, camostat, against BA.5, EG.5.1, and BA.2.86. We further found that N-0385 shows broad synergistic activity with clinically approved direct-acting antivirals (DAAs), i.e., remdesivir and nirmatrelvir, against Omicron subvariants, demonstrating the potential therapeutic benefits of a multi-targeted treatment based on N-0385 and DAAs.
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Benzotiazoles , COVID-19 , Sulfonamidas , Humanos , Anticuerpos Neutralizantes , Anticuerpos Antivirales , Antivirales , SARS-CoV-2 , Serina EndopeptidasasRESUMEN
SARS-CoV-2, the causative virus of COVID-19, continues to threaten global public health. COVID-19 is a multi-organ disease, causing not only respiratory distress, but also extrapulmonary manifestations, including gastrointestinal symptoms with SARS-CoV-2 RNA shedding in stool long after respiratory clearance. Despite global vaccination and existing antiviral treatments, variants of concern are still emerging and circulating. Of note, new Omicron BA.5 sublineages both increasingly evade neutralizing antibodies and demonstrate an increased preference for entry via the endocytic entry route. Alternative to direct-acting antivirals, host-directed therapies interfere with host mechanisms hijacked by viruses, and enhance cell-mediated resistance with a reduced likelihood of drug resistance development. Here, we demonstrate that the autophagy-blocking therapeutic berbamine dihydrochloride robustly prevents SARS-CoV-2 acquisition by human intestinal epithelial cells via an autophagy-mediated BNIP3 mechanism. Strikingly, berbamine dihydrochloride exhibited pan-antiviral activity against Omicron subvariants BA.2 and BA.5 at nanomolar potency, providing a proof of concept for the potential for targeting autophagy machinery to thwart infection of current circulating SARS-CoV-2 subvariants. Furthermore, we show that autophagy-blocking therapies limited virus-induced damage to intestinal barrier function, affirming the therapeutic relevance of autophagy manipulation to avert the intestinal permeability associated with acute COVID-19 and post-COVID-19 syndrome. Our findings underscore that SARS-CoV-2 exploits host autophagy machinery for intestinal dissemination and indicate that repurposed autophagy-based antivirals represent a pertinent therapeutic option to boost protection and ameliorate disease pathogenesis against current and future SARS-CoV-2 variants of concern.
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COVID-19 , Hepatitis C Crónica , Humanos , SARS-CoV-2 , Antivirales/farmacología , Síndrome Post Agudo de COVID-19 , ARN Viral , Anticuerpos Neutralizantes , Autofagia , Anticuerpos Antivirales , Glicoproteína de la Espiga del Coronavirus , Proteínas de la MembranaRESUMEN
The protein kinase C-activating sponge natural product alotaketal C (1) potently inhibits the infection of human Calu-3 lung cells by SARS-CoV-2 Omicron BA.1 and BA.5 variants. Simplified analogs of 1 have been synthesized and tested for anti-SARS-CoV-2 activity providing SAR data for the antiviral pharmacophore of 1. Analogs 19 and 23, which are missing the C-11 substituents in 1 and have modified C-13 appendages, are â¼2- to 7-fold more potent than 1 and have equal or larger selectivity indices.
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Productos Biológicos , COVID-19 , Humanos , SARS-CoV-2 , Antivirales/farmacología , FarmacóforoRESUMEN
Antivirals with broad coronavirus activity are important for treating high-risk individuals exposed to the constantly evolving SARS-CoV-2 variants of concern (VOCs) as well as emerging drug-resistant variants. We developed and characterized a novel class of active-site-directed 3-chymotrypsin-like protease (3CLpro) inhibitors (C2-C5a). Our lead direct-acting antiviral (DAA), C5a, is a non-covalent, non-peptide with a dissociation constant of 170 nM against recombinant SARS-CoV-2 3CLpro. The compounds C2-C5a exhibit broad-spectrum activity against Omicron subvariants (BA.5, BQ.1.1, and XBB.1.5) and seasonal human coronavirus-229E infection in human cells. Notably, C5a has median effective concentrations of 30-50â nM against BQ.1.1 and XBB.1.5 in two different human cell lines. X-ray crystallography has confirmed the unique binding modes of C2-C5a to the 3CLpro, which can limit virus cross-resistance to emerging Paxlovid-resistant variants. We tested the effect of C5a with two of our newly discovered host-directed antivirals (HDAs): N-0385, a TMPRSS2 inhibitor, and bafilomycin D (BafD), a human vacuolar H+-ATPase [V-ATPase] inhibitor. We demonstrated a synergistic action of C5a in combination with N-0385 and BafD against Omicron BA.5 infection in human Calu-3 lung cells. Our findings underscore that a SARS-CoV-2 multi-targeted treatment for circulating Omicron subvariants based on DAAs (C5a) and HDAs (N-0385 or BafD) can lead to therapeutic benefits by enhancing treatment efficacy. Furthermore, the high-resolution structures of SARS-CoV-2 3CLpro in complex with C2-C5a will facilitate future rational optimization of our novel broad-spectrum active-site-directed 3C-like protease inhibitors.
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COVID-19 , Hepatitis C Crónica , Humanos , Inhibidores de Proteasas/farmacología , Antivirales/farmacología , SARS-CoV-2RESUMEN
The COVID-19 pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), remains a global public health crisis. The reduced efficacy of therapeutic monoclonal antibodies against emerging SARS-CoV-2 variants of concern (VOCs), such as omicron BA.5 subvariants, has underlined the need to explore a novel spectrum of antivirals that are effective against existing and evolving SARS-CoV-2 VOCs. To address the need for novel therapeutic options, we applied cell-based high-content screening to a library of natural products (NPs) obtained from plants, fungi, bacteria, and marine sponges, which represent a considerable diversity of chemical scaffolds. The antiviral effect of 373 NPs was evaluated using the mNeonGreen (mNG) reporter SARS-CoV-2 virus in a lung epithelial cell line (Calu-3). The screening identified 26 NPs with half-maximal effective concentrations (EC50) below 50 µM against mNG-SARS-CoV-2; 16 of these had EC50 values below 10 µM and three NPs (holyrine A, alotaketal C, and bafilomycin D) had EC50 values in the nanomolar range. We demonstrated the pan-SARS-CoV-2 activity of these three lead antivirals against SARS-CoV-2 highly transmissible Omicron subvariants (BA.5, BA.2 and BA.1) and highly pathogenic Delta VOCs in human Calu-3 lung cells. Notably, holyrine A, alotaketal C, and bafilomycin D, are potent nanomolar inhibitors of SARS-CoV-2 Omicron subvariants BA.5 and BA.2. The pan-SARS-CoV-2 activity of alotaketal C [protein kinase C (PKC) activator] and bafilomycin D (V-ATPase inhibitor) suggest that these two NPs are acting as host-directed antivirals (HDAs). Future research should explore whether PKC regulation impacts human susceptibility to and the severity of SARS-CoV-2 infection, and it should confirm the important role of human V-ATPase in the VOC lifecycle. Interestingly, we observed a synergistic action of bafilomycin D and N-0385 (a highly potent inhibitor of human TMPRSS2 protease) against Omicron subvariant BA.2 in human Calu-3 lung cells, which suggests that these two highly potent HDAs are targeting two different mechanisms of SARS-CoV-2 entry. Overall, our study provides insight into the potential of NPs with highly diverse chemical structures as valuable inspirational starting points for developing pan-SARS-CoV-2 therapeutics and for unravelling potential host factors and pathways regulating SARS-CoV-2 VOC infection including emerging omicron BA.5 subvariants.
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Productos Biológicos , COVID-19 , Humanos , SARS-CoV-2 , Pandemias , Adenosina Trifosfatasas , Antivirales/farmacología , Antivirales/uso terapéutico , Productos Biológicos/farmacología , Glicoproteína de la Espiga del CoronavirusRESUMEN
Rotaviruses, the single most important agents of acute severe gastroenteritis in children, are nonenveloped viruses formed by a three-layered capsid that encloses a genome formed by 11 segments of double-stranded RNA. The mechanism of entry of these viruses into the host cell is not well understood. The best-studied strain, RRV, which is sensitive to neuraminidase (NA) treatment of the cells, uses integrins alpha2 beta1 and alphav beta3 and the heat shock protein hsc70 as receptors and enters MA104 cells through a non-clathrin-, non-caveolin-mediated pathway that depends on a functional dynamin and on the presence of cholesterol on the cell surface. In this work, using a combination of pharmacological, biochemical, and genetic approaches, we compared the entry characteristics of four rotavirus strains known to have different receptor requirements. We chose four rotavirus strains that represent all phenotypic combinations of NA resistance or sensitivity and integrin dependence or independence. We found that even though all the strains share their requirements for hsc70, dynamin, and cholesterol, three of them differ from the simian strain RRV in the endocytic pathway used. The human strain Wa, porcine strain TFR-41, and bovine strain UK seem to enter the cell through clathrin-mediated endocytosis, since treatments that inhibit this pathway block their infectivity; consistent with this entry route, these strains were sensitive to changes in the endosomal pH. The inhibition of other endocytic mechanisms, such as macropinocytosis or caveola-mediated uptake, had no effect on the internalization of the rotavirus strains tested here.
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Endocitosis , Células Epiteliales/virología , Rotavirus/fisiología , Internalización del Virus , Animales , Bovinos , Línea Celular , Colesterol/metabolismo , Vesículas Cubiertas por Clatrina/virología , Dinaminas/metabolismo , Proteínas del Choque Térmico HSC70/metabolismo , Haplorrinos , Humanos , PorcinosRESUMEN
Hepatitis delta virus (HDV) is a defective human virus that lacks the ability to produce its own envelope proteins and is thus dependent on the presence of a helper virus, which provides its surface proteins to produce infectious particles. Hepatitis B virus (HBV) was so far thought to be the only helper virus described to be associated with HDV. However, recent studies showed that divergent HDV-like viruses could be detected in fishes, birds, amphibians, and invertebrates, without evidence of any HBV-like agent supporting infection. Another recent study demonstrated that HDV can be transmitted and propagated in experimental infections ex vivo and in vivo by different enveloped viruses unrelated to HBV, including hepatitis C virus (HCV) and flaviviruses such as Dengue and West Nile virus. All this new evidence, in addition to the identification of novel virus species within a large range of hosts in absence of HBV, suggests that deltaviruses may take advantage of a large spectrum of helper viruses and raises questions about HDV origins and evolution.
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Virus Helper , Hepatitis D/virología , Virus de la Hepatitis Delta , Animales , Evolución Molecular , Genoma Viral , Virus Helper/fisiología , Virus de la Hepatitis Delta/clasificación , Virus de la Hepatitis Delta/genética , Virus de la Hepatitis Delta/fisiología , Especificidad del Huésped , Humanos , Filogenia , Proteínas Virales/genética , Proteínas Virales/metabolismo , Replicación ViralRESUMEN
Cell entry of enveloped viruses relies on the fusion between the viral and plasma or endosomal membranes, through a mechanism that is triggered by a cellular signal. Here we used a combination of computational and experimental approaches to unravel the main determinants of hepatitis B virus (HBV) membrane fusion process. We discovered that ERp57 is a host factor critically involved in triggering HBV fusion and infection. Then, through modeling approaches, we uncovered a putative allosteric cross-strand disulfide (CSD) bond in the HBV S glycoprotein and we demonstrate that its stabilization could prevent membrane fusion. Finally, we identified and characterized a potential fusion peptide in the preS1 domain of the HBV L glycoprotein. These results underscore a membrane fusion mechanism that could be triggered by ERp57, allowing a thiol/disulfide exchange reaction to occur and regulate isomerization of a critical CSD, which ultimately leads to the exposition of the fusion peptide.
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Antígenos de Superficie de la Hepatitis B/metabolismo , Proteína Disulfuro Isomerasas/metabolismo , Precursores de Proteínas/metabolismo , Proteínas del Envoltorio Viral/metabolismo , Acoplamiento Viral , Animales , Células CHO , Línea Celular Tumoral , Cricetinae , Cricetulus , Femenino , Regulación Viral de la Expresión Génica , Virus de la Hepatitis B , Hepatocitos , Humanos , Masculino , Fusión de Membrana , Ratones , Proteína Disulfuro Isomerasas/genética , Proteínas del Envoltorio Viral/genéticaRESUMEN
Hepatitis D virus (HDV) doesn't encode envelope proteins for packaging of its ribonucleoprotein (RNP) and typically relies on the surface glycoproteins (GPs) from hepatitis B virus (HBV) for virion assembly, envelopment and cellular transmission. HDV RNA genome can efficiently replicate in different tissues and species, raising the possibility that it evolved, and/or is still able to transmit, independently of HBV. Here we show that alternative, HBV-unrelated viruses can act as helper viruses for HDV. In vitro, envelope GPs from several virus genera, including vesiculovirus, flavivirus and hepacivirus, can package HDV RNPs, allowing efficient egress of HDV particles in the extracellular milieu of co-infected cells and subsequent entry into cells expressing the relevant receptors. Furthermore, HCV can propagate HDV infection in the liver of co-infected humanized mice for several months. Further work is necessary to evaluate whether HDV is currently transmitted by HBV-unrelated viruses in humans.
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Coinfección/transmisión , Hepatitis D/transmisión , Virus de la Hepatitis Delta/fisiología , Ensamble de Virus , Animales , Línea Celular Tumoral , Coinfección/virología , Flavivirus/metabolismo , Hepacivirus/metabolismo , Hepacivirus/patogenicidad , Hepatitis D/virología , Virus de la Hepatitis Delta/aislamiento & purificación , Virus de la Hepatitis Delta/patogenicidad , Hepatocitos/trasplante , Hepatocitos/virología , Humanos , Ratones , Ratones Endogámicos NOD , Ratones Transgénicos , Cultivo Primario de Células , ARN Viral/aislamiento & purificación , Ribonucleoproteínas/metabolismo , Vesiculovirus/metabolismo , Proteínas del Envoltorio Viral/metabolismo , Virión/metabolismoRESUMEN
Inactivated influenza vaccines (IIVs) lack broad efficacy. Cellular immunity to a conserved internal antigen, the nucleoprotein (NP), has been correlated to protection against pandemic and seasonal influenza and thus could have the potential to broaden vaccine efficacy. We developed OVX836, a recombinant protein vaccine based on an oligomerized NP, which shows increased uptake by dendritic cells and immunogenicity compared with NP. Intramuscular immunization in mice with OVX836 induced strong NP-specific CD4+ and CD8+ T-cell systemic responses and established CD8+ tissue memory T cells in the lung parenchyma. Strikingly, OVX836 protected mice against viral challenge with three different influenza A subtypes, isolated several decades apart and induced a reduction in viral load. When co-administered with IIV, OVX836 was even more effective in reducing lung viral load.
RESUMEN
Rotavirus infection is the cause of severe gastroenteritis of young children worldwide, leading to an estimate of 600,000 deaths a year. Efforts to develop an effective and safe vaccine resulted in licensing in 1998 of a live oral vaccine (RotaShield) that was withdrawn less than 1 year later when reports of cases of intussusception were linked to its application. This led to development of new rotavirus vaccine candidates that are currently in late phase III clinical trials. One of these vaccines, GlaxoSmithKline's Rotarix, was licensed in July 2004 to be used in Mexico. This review describes the general background for rotavirus vaccine development, the different vaccine candidates that have been tested or are currently being evaluated, the association of rotavirus vaccination with the bowel blockage known as intussuception, and discusses the benefits and risks of the fast-track introduction of Rotarix in Latin America, and particularly in Mexico.
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Gastroenteritis/prevención & control , Infecciones por Rotavirus/prevención & control , Vacunas contra Rotavirus/administración & dosificación , Vacunas Atenuadas/administración & dosificación , Niño , Preescolar , Ensayos Clínicos Fase III como Asunto , Femenino , Gastroenteritis/inmunología , Gastroenteritis/virología , Humanos , América Latina , Masculino , México , Infecciones por Rotavirus/inmunología , Vacunas contra Rotavirus/inmunología , Vacunas Atenuadas/inmunologíaAsunto(s)
Virus de la Hepatitis B/fisiología , Hepatitis D/virología , Virus de la Hepatitis Delta/fisiología , Liberación del Virus/fisiología , Replicación Viral/fisiología , Animales , Coinfección , Hepatitis B/complicaciones , Hepatitis B/virología , Hepatitis D/complicaciones , Virus de la Hepatitis Delta/patogenicidad , HumanosRESUMEN
The heat shock cognate protein hsc70 has been implicated as a postattachment cell receptor for rotaviruses. Here we show that hsc70 interacts specifically with rotaviruses through its peptide-binding domain, since a recombinant full-length hsc70 protein and its peptide-binding domain, but not its ATPase domain, bound triple-layered particles in a solid-phase assay, and known ligands of hsc70 competed this binding. The peptide ligands of hsc70 were also shown to block rotavirus infectivity when added to cells before virus infection, suggesting that hsc70 on the surface of MA104 cells also interacts with the virus through its peptide-binding domain and that this interaction is important for virus entry. When purified infectious virus was incubated with soluble hsc70 in the presence of the cochaperone hsp40 and ATP and then pelleted through a sucrose cushion, the recovered virus had lost 60% of its infectivity, even though hsc70 was not detected in the pellet fraction. The hsc70-treated virus showed slightly different reactivities with monoclonal antibodies and was more susceptible to heat and basic pHs than the untreated virus, suggesting that hsc70 induces a subtle conformational change in the virus that results in a reduction of its infectivity. The relevance of the ATPase activity of hsc70 for reducing virus infectivity was demonstrated by the finding that in the presence of a nonhydrolyzable analogue of ATP, virus infectivity was not affected, and a mutant protein lacking ATPase activity failed to reduce virus infection. Altogether, these results suggest that during cell infection, the interaction of the virus with hsc70 on the surface of MA104 cells results in a conformational change of virus particles that facilitates their entry into the cell cytoplasm.