Dengue virus M and E proteins belonging to genotype II (Cosmopolitan) of serotype 2 are influenced by the nature of M residue 36.

Mosquito-borne dengue disease is caused by the dengue virus serotype-1 to serotype-4. The contemporary dengue outbreaks in the southwestern Indian ocean coincided with the widespread of dengue virus serotype 2 genotype II (Cosmopolitan), including epidemic viral strains DES-14 and RUN-18 isolated in Dar es Salaam (Tanzania) in 2014 and La Reunion Island (France) in 2018, respectively. Heterodimeric interaction between prM (intracellular precursor of surface structural M protein) and envelope E proteins is required during the initial stage of dengue virus assembly. Amino acid 127 of DES-14 prM protein (equivalent to M36) has been identified as an infrequent valine whereas RUN-18 has a common isoleucine. In the present study, we examined the effect of M-I36V mutation on the expression of a recombinant RUN-18 E protein co-expressed with prM in human epithelial A549 cells. The M ectodomain of dengue virus serotype 2 embeds a pro-apoptotic peptide referred as D2AMP. The impact of M-I36V mutation on the death-promoting capability of D2AMP was assessed in A549 cells. We showed that valine at position M36 affects expression of recombinant RUN-18 E protein and potentiates apoptosis-inducing activity of D2AMP. We propose that the nature of M residue 36 influences the virological characteristics of dengue 2 M and E proteins belonging to genotype II that contributes to global dengue burden.

Role of Disulphide Bonds in Membrane Partitioning of a Viral Peptide.

The importance of disulphide bond in mediating viral peptide entry into host cells is well known. In the present work, we elucidate the role of disulphide (SS) bond in partitioning mechanism of membrane-active Hepatitis A Virus-2B (HAV-2B) peptide, which harbours three cysteine residues promoting formation of multiple SS-bonded states. The inclusion of SS-bond not only results in a compact conformation but also induces distorted α-helical hairpin geometry in comparison to SS-free state. Owing to these, the hydrophobic residues get buried, restricting the insertion of SS-bonded HAV-2B peptide into lipid packing defects and thus the partitioning of the peptide is completely or partly abolished. In this way, the disulphide bond can potentially regulate the partitioning of HAV-2B peptide such that the membrane remodelling effects of this viral peptide are significantly reduced. The current findings may have potential implications in drug designing, targeting the HAV-2B protein by promoting disulphide bond formation within its membrane-active region.

Possible role of HLA class-I genotype in SARS-CoV-2 infection and progression: A pilot study in a cohort of Covid-19 Spanish patients.

Human Leukocyte Antigen (HLA) includes a large set of genes with important actions in immune response against viral infection. Numerous studies have revealed the existence of significant associations between certain HLA alleles and the susceptibility and prognosis of different infectious diseases. In this pilot study we analyse the binding affinity between 66 class I HLA alleles and SARS-CoV-2 viral peptides, and its association with the severity of the disease. A total of 45 Spanish patients with mild, moderate and severe SARS-CoV-2 infection were typed for HLA class I; after that, we analysed if an in silico model of HLA I-viral peptide binding affinity and classical HLA supertypes could be correlated to the severity of the disease. Our results suggest that patients with mild disease present Class I HLA molecules with a higher theoretical capacity for binding SARS-Cov-2 peptides and showed greater heterozygosity when comparing them with moderate and severe groups. In this regard, identifying HLA-SARS-CoV-2 peptides binding differences between individuals would help to clarify the heterogeneity of clinical responses to the disease and will also be useful to guide a personalized treatment according to its particular risk.

Molecular challenges imposed by MHC-I restricted long epitopes on T cell immunity.

It has widely been accepted that major histocompatibility complex class I molecules (MHC-I) are limited to binding small peptides of 8-10 residues in length. However, this consensus has recently been challenged with the identification of longer peptides (≥11 residues) that can also elicit cytotoxic CD8+ T cell responses. Indeed, a growing number of studies demonstrate that these non-canonical epitopes are important targets for the immune system. As long epitopes represent up to 10% of the peptide repertoire bound to MHC-I molecules, here we review their impact on antigen presentation by MHC-I, TCR recognition, and T cell immunity.

Peculiarities of the Presentation of the Encephalitogenic MBP Peptide by HLA-DR Complexes Providing Protection and Predisposition to Multiple Sclerosis.

Predisposition to multiple sclerosis (MS), a chronic autoimmune disease of the central nervous system, is due to various factors. The genetic component is considered one of the most important factors. HLA class II genes contribute the most to the development of MS. The HLA-DRB1*15 allele group is considered one of the main genetic risk factors predisposing to MS. The group of HLA-DRB1*01 alleles was shown to have a protective effect against this disease in the Russian population. In this work, we compared the binding of the encephalitogenic fragment of the myelin basic protein (MBP) to two HLA-DR complexes that provide protection against and predisposition to MS: HLA-DR1 (HLA-DRB1*0101) and HLA-DR15 (HLA-DRB1*1501), respectively. We found that the myelin peptide MBP88-100 binds to HLA-DR1 at a rate almost an order of magnitude lower than the viral peptide of hemagglutinin (HA). The same was true for the binding of MBP85-97 to HLA-DR15 in comparison with viral pp65. The structure of the C-terminal part of the peptide plays a key role in the binding to HLA-DR1 for equally high-affinity N-terminal regions of the peptides. The IC50 of the myelin peptide MBP88-100 competing with viral HA for binding to HLA-DR1 is almost an order of magnitude higher than that of HA. As for HA, the same was also true for the binding of MBP85-97 to HLA-DR15 in comparison with viral pp65. Thus, autoantigenic MBP cannot compete with the viral peptide for binding to protective HLA-DR1. However, it is more competitive than viral peptide for HLA-DR15.

Indication of Neurodegenerative Cascade Initiation by Amyloid-like Aggregate-Forming EBV Proteins and Peptide in Alzheimer's Disease.

The neurotropic potential of the Epstein-Barr virus (EBV) was demonstrated quite recently; however, the mechanistic details are yet to be explored. Therefore, the effects of EBV infection in the neural milieu remain underexplored. Previous reports have suggested the potential role of virus-derived peptides in seeding the amyloid-ß aggregation cascade, which lies at the center of Alzheimer's disease (AD) pathophysiology. However, no such study has been undertaken to explore the role of EBV peptides in AD. In our research, ∼100 EBV proteins were analyzed for their aggregation proclivity in silico using bioinformatic tools, followed by the prediction of 20S proteasomal cleavage sites using online algorithms NetChop ver. 3.1 and Pcleavage, thereby mimicking the cellular proteasomal cleavage activity generating short antigenic peptides of viral origin. Our study reports a high aggregate-forming tendency of a 12-amino-acid-long (146SYKHVFLSAFVY157) peptide derived from EBV glycoprotein M (EBV-gM). The in vitro analysis of aggregate formation done using Congo red and Thioflavin-S assays demonstrated dose- and time-dependent kinetics. Thereafter, Raman spectroscopy was used to validate the formation of secondary structures (α helix, ß sheets) in the aggregates. Additionally, cytotoxicity assay revealed that even a low concentration of these aggregates has a lethal effect on neuroblastoma cells. The findings of this study provide insights into the mechanistic role of EBV in AD and open up new avenues to explore in the future.

Immune Reactivity of a 20-mer Peptide Representing the Zika E Glycan Loop Involves the Antigenic Determinants E-152/156/158.

Mosquito-borne Zika virus (ZIKV) causes a severe congenital syndrome and neurological disorders in humans. With the aim to develop a live-attenuated ZIKV strain, we generated a chimeric viral clone ZIKALIVax with African MR766-NIID strain as backbone and the envelope E protein of epidemic Brazilian BeH810915 strain. The MR766-NIID residues E-T152/I156/Y158 were introduced into BeH810915 E protein leading to a nonglycosylated ZIKALIVax. Recently, we reported that the residues E-152/156/158 that are part of ZIKV glycan loop (GL) region might have an impact on the availability of neutralizing antibody epitopes on ZIKV surface. In the present study, we evaluated the antigenic reactivity of a synthetic 20-mer peptide representing the ZIKALIVax GL region. The GL-related peptide was effective for the detection of GL-reactive antibody in mouse anti-ZIKALIVax immune serum. We showed that the residue E-158 influences the antigenic reactivity of GL-related peptide. The ZIKALIVax peptide was effective in generating mouse antibodies with reactivity against a recombinant E domain I that encompasses the GL region. The GL peptide-reactive antibodies revealed that antigenic reactivity of E-domain I may be impacted by both residues E-152 and E-156. In conclusion, we proposed a role for the residues E-152/156/158 as key antigenic determinants of ZIKV glycan loop region.

A Novel Human Radixin Peptide Inhibits Hepatitis C Virus Infection at the Level of Cell Entry.

Hepatitis C virus infection of hepatocytes is a multistep process involving the interaction between viral and host cell molecules. Recently, we identified ezrin-moesin-radixin proteins and spleen tyrosine kinase (SYK) as important host therapeutic targets for HCV treatment development. Previously, an ezrin hinge region peptide (Hep1) has been shown to exert anti-HCV properties in vivo, though its mechanism of action remains limited. In search of potential novel inhibitors of HCV infection and their functional mechanism we analyzed the anti-HCV properties of different human derived radixin peptides. Sixteen different radixin peptides were derived, synthesized and tested. Real-time quantitative PCR, cell toxicity assay, immuno-precipitation/western blot analysis and computational resource for drug discovery software were used for experimental analysis. We found that a human radixin hinge region peptide (Peptide1) can specifically block HCV J6/JFH-1 infection of Huh7.5 cells. Peptide 1 had no cell toxicity or intracellular uptake into Huh7.5 cells. Mechanistically, the anti-HCV activity of Peptide 1 extended to disruption of HCV engagement of CD81 thereby blocking downstream SYK activation, which we have recently demonstrated to be important for effective HCV infection of target hepatocytes. Our findings highlight a novel functional class of anti-HCV agents that can inhibit HCV infection, most likely by disrupting vital viral-host signaling interactions at the level of virus entry.

pMHC/scFv fusion protein mediates killing of tumor cells by viral specific cytotoxic T cells / 中华微生物学和免疫学杂志

Objective To study whether the HBC-A2/scFv fusion protein mediates killing of tumor cells by viral specific cytotoxic T cells. Methods The fusion protein was attached to the CD71-expressing, HLA class Ⅰ negative tumor cells. And then, cytolysis by viral peptide-specific CTLs which were generated by co-culture of peripheral blood lymphocytes from HLA-A2 positive donors with inactivated T2 cells pulsed with the viral peptide were tested by lactate dehydrogenase (LDH) releasing. Results The fusion protein can attach the active viral peptide/HLA-A2 complex to K562, HepG2 and U937 cells through binding of CD71 scFv to CD71 (37.30% ±8.25%, 27.20% ±3.88%, 21.80% ±6.49% ) and mediate cytotoxicity of viral peptide-specific CTLs against those cells in vitro ( K562: 42.08% ± 1.14% vs 8.07%± 1.39%; HepG2: 49.72% ± 1.59% vs 12.46% ± 1.26%; U937: 39.72% ± 3.26% vs 7.13% ±1.48% ). Conclusion This viral peptide/HLA-A2 complex targeted by CD71 scFv is able to redirect viral peptide-specific T-cell mediated immune responses against tumor cells.