Cross-reactive memory T cells associate with protection against SARS-CoV-2 infection in COVID-19 contacts

Cross-reactive immune responses to SARS-CoV-2 have been observed in pre-pandemic cohorts and proposed to contribute to host protection. Here we assess 52 COVID-19 household contacts to capture immune responses at the earliest timepoints after SARS-CoV-2 exposure. Using a dual cytokine FLISpot assay on peripheral blood mononuclear cells, we enumerate the frequency of T cells specific for spike, nucleocapsid, membrane, envelope and ORF1 SARS-CoV-2 epitopes that cross-react with human endemic coronaviruses. We observe higher frequencies of cross-reactive (p = 0.0139), and nucleocapsid-specific (p = 0.0355) IL-2-secreting memory T cells in contacts who remained PCR-negative despite exposure (n = 26), when compared with those who convert to PCR-positive (n = 26); no significant difference in the frequency of responses to spike is observed, hinting at a limited protective function of spike-cross-reactive T cells. Our results are thus consistent with pre-existing non-spike cross-reactive memory T cells protecting SARS-CoV-2-naïve contacts from infection, thereby supporting the inclusion of non-spike antigens in second-generation vaccines.

Structural basis of mitochondrial protein import by the TIM complex

Mitochondria import nearly all their ~1,000-2,000 constituent proteins from the cytosol across their double membrane envelope. Genetic and biochemical studies have shown that the conserved protein translocase, termed the TIM complex (also known as TIM23 complex), mediates import of presequence-containing proteins into the mitochondrial matrix and inner membrane. Among ~10 different subunits of the complex, the essential multi-pass membrane protein Tim23, together with the evolutionarily related protein Tim17, has long been postulated to form a protein-conducting channel. However, the mechanism of TIM-mediated protein import remains uncertain due to a lack of structural information on the complex. Here, we have determined the cryo-EM structure of the core TIM complex (Tim17-Tim23-Tim44) from Saccharomyces cerevisiae. We show that, contrary to the prevailing model, Tim23 and Tim17 do not form a water-filled channel, but instead have separate, lipid-exposed concave cavities that face in opposite directions. Remarkably, our data suggest that the cavity of Tim17 itself forms the protein translocation path whereas Tim23 plays a structural role. We also show how the Tim17-Tim23 heterodimer associates with the scaffold protein Tim44 and J-domain proteins to mediate Hsp70-driven polypeptide transport into the matrix. Our work provides the structural foundation to understand the mechanism of TIM-mediated protein import and sorting, a central pathway in mitochondrial biogenesis.

Colony-like Protocell Superstructures

We report the formation, growth, and dynamics of model protocell superstructures on solid surfaces, resembling single cell colonies. These structures, consisting of several layers of lipidic compartments enveloped in a dome-shaped outer lipid bilayer, emerged as a result of spontaneous shape transformation of lipid agglomerates deposited on thin film aluminum surfaces. Collective protocell structures were observed to be mechanically more stable compared to isolated spherical compartments. We show that the model colonies encapsulate DNA and accommodate non-enzymatic, strand displacement DNA reactions. The membrane envelope is able to disassemble and expose individual daughter protocells, which can migrate and attach via nano-tethers to distant surface locations, while maintaining their encapsulated contents. Some colonies feature ‘exo-compartments’, which spontaneously extend out of the enveloping bilayer, internalize DNA, and merge again with the superstructure. A continuum elastohydrodynamic theory that we developed reveals that the subcompartment formation must be governed by attractive van der Waals (vdW) interactions between the membrane and surface. The balance between membrane bending and vdW interactions yields a critical length scale of 273 nm, above which the membrane invaginations can form subcompartments. The findings support our hypotheses that in extension of the ‘lipid world hypothesis’, protocells may have existed in the form of colonies, potentially benefiting from the increased mechanical stability provided by a superstructure.

Combining detection and reconstruction of correlational and periodic motifs in viral genomic sequences with transitional genome mapping: Application to COVID-19

A method of Transitional Automorphic Mapping of the Genome on Itself (TAMGI) is aimed at combining detection and reconstruction of correlational and quasi-periodic motifs in the viral genomic RNA/DNA sequences. The motifs reconstructed by TAMGI are robust with respect to indels and point mutations and can be tried as putative therapeutic targets. We developed and tested the relevant theory and statistical criteria for TAMGI applications. The applications of TAMGI are illustrated by the study of motifs in the genomes of the severe acute respiratory syndrome coronaviruses SARS-CoV and SARS-CoV-2 (the latter coronavirus SARS-CoV-2 being responsible for the COVID-19 pandemic) packaged within filament-like helical capsid. Such ribonucleocapsid is transported into spherical membrane envelope with incorporated spike glycoproteins. Two other examples concern the genomes of viruses with icosahedral capsids, satellite tobacco mosaic virus (STMV) and bacteriophage PHIX174. A part of the quasi-periodic motifs in these viral genomes was evolved due to weakly specific cooperative interaction between genomic ssRNA/ssDNA and nucleocapsid proteins. The symmetry of the capsids leads to the natural selection of specific quasi-periodic motifs in the related genomic sequences. Generally, TAMGI provides a convenient tool for the study of numerous molecular mechanisms with participation of both quasi-periodic motifs and complete repeats, the genome organization, contextual analysis of cis/trans regulatory elements, data mining, and correlations in the genomic sequences.

Features of immunopathogenesis of a new coronavirus infection

The main biological characteristics of viruses of the Coronaviridae family are presented. The features of the immunopathogenesis of these infections are analyzed. It was found that the structural proteins of the spine, membrane, envelope and nucleocapsid play an important role in the immunopathogenesis of COVID-19 infection. They are associated with hyperactivation of neutrophils and monocytes-macrophages, secreting large amounts of pro-inflammatory cytokines and chemokines. This contributes to the development of a cytokine storm and an unfavorable prognosis of the disease. A particularly high risk of developing pneumonia exists against the background of an increase in the production of: macrophage inflammatory protein-1 alpha, macrophage chemotactic protein, interleukin 8. At the height of infection in some patients, macrophages and dendritic cells infected with SARS-CoV-2 lose their ability to produce type I interferons and pro-inflammatory cytokines. On the part of cellular immunity, a significant decrease in the number of CD4+ and CD8+-lymphocytes was noted. Among IgG sub-isotypes, IgG3 antibodies had the highest reactivity, and IgG1 antibodies were less reactive. Antibodies to spike protein with low specificity or low titer do not neutralize the virus and contribute to the contamination of immunocompetent cells via Fc receptors. Low-affinity antibodies or their low levels can contribute to increased cell sensitivity to SARS-CoV-2 and the development of severe forms of COVID-19 disease.

Circulating CD4 T cells elicited by endemic coronaviruses display vast disparities in abundance and functional potential linked to both antigen specificity and age

Repeated infections with endemic human coronaviruses are thought to reflect lack of long-lasting protective immunity. Here, we evaluate circulating human CD4 T cells collected prior to 2020 for reactivity towards hCoV spike proteins, probing for the ability to produce IFN-γ, IL-2 or granzyme B. We find robust reactivity to spike-derived epitopes, comparable to influenza, but highly variable abundance and functional potential across subjects, depending on age and viral antigen specificity. To explore the potential of these memory cells to be recruited in SARS-CoV-2 infection, we examined the same subjects for cross-reactive recognition of epitopes from SARS-CoV-2 nucleocapsid, membrane/envelope, and spike. The functional potential of these cross-reactive CD4 T cells was highly variable, with nucleocapsid-specific CD4 T cells, but not spike-reactive cells showing exceptionally high levels of granzyme production upon stimulation. These results are considered in light of recruitment of hCoV-reactive cells into responses of humans to SARS-CoV infections or vaccinations.

Genomic analysis of Helicobacter himalayensis sp. nov. isolated from Marmota himalayana

Background: Helicobacter himalayensis was isolated from Marmota himalayana in the Qinghai-Tibet Plateau, China, and is a new non-H. pylori species, with unclear taxonomy, phylogeny, and pathogenicity. Results: A comparative genomic analysis was performed between the H. himalayensis type strain 80(YS1)T and other the genomes of Helicobacter species present in the National Center for Biotechnology Information (NCBI) database to explore the molecular evolution and potential pathogenicity of H. himalayensis. H. himalayensis 80(YS1)T formed a clade with H. cinaedi and H. hepaticus that was phylogenetically distant from H. pylori. The H. himalayensis genome showed extensive collinearity with H. hepaticus and H. cinaedi. However, it also revealed a low degree of genome collinearity with H. pylori. The genome of 80(YS1)T comprised 1,829,936 bp, with a 39.89% GC content, a predicted genomic island, and 1,769 genes. Comparatively, H. himalayensis has more genes for functions in “cell wall/membrane/envelope biogenesis” and “coenzyme transport and metabolism” sub-branches than the other compared helicobacters, and its genome contained 42 virulence factors genes, including that encoding cytolethal distending toxin (CDT). Conclusions: We characterized the H. himalayensis 80(YS1)T genome, its phylogenetic position, and its potential pathogenicity. However, further understanding of the pathogenesis of this potentially pathogenic bacterium is required, which might help to manage H. himalayensis-induced diseases.

An in silico LLPS perturbation approach in the design of a novel SARS-CoV-2 spike receptor-binding domain inhibitor

The reversible process where a homogenous fluid de-mixes into two distinctively separate liquid phases is referred to as LLPS (Liquid-liquid phase separation). The resulting liquid is made up of one dilute phase and one condensed phase. An increasing number of studies have shown that the liquid-liquid phase separation is an important principle that underlies intracellular organization in biological systems, forming liquid condensates without a membrane envelope, otherwise known as MLOs (membraneless organelles). Such organelles include the P bodies, nucleolus and stress granules. Moreover, the regulation of many other biological processes such as signal transduction, chromatin rearrangement and RNA metabolism have been linked to the liquid-liquid phase separation.

Sequence and Structural Analysis of COVID-19 E and M Protein With MERS Virus E and M Protein – A Comparative Study

The outbreak of SARS in 2003, MERS in 2012, and now COVID-19 in 2019 have demonstrated that Coronaviruses are capable of causing primary lethal infections in humans, and the pandemic is now a global concern. The COVID-19 belongs to the beta coronavirus family encoding 29 proteins, of which 4 are structural, the Spike, Membrane, Envelope, and Nucleocapsid proteins. Here we have analyzed and compared the Membrane (M) and Envelope (E) proteins of COVID-19 and MERS with SARS and Bat viruses. The sequence analysis of conserved regions of both E and M protein revealed that many regions of COVID-19 are similar to Bat and SARS viruses while the MERS virus showed variations. The essential binding motifs found in SARS-CoV appeared in COVID-19. Besides, the M protein of COVID- 19 showed a distinct serine phosphorylation site in the C-terminal domain, which looked like a catalytic triad seen in serine proteases. A Dileucine motif occurred many times in the sequence of the M protein of all the four viruses compared. Concerning the structural part, the COVID-19 E protein showed more similarity to Bat while MERS shared similarity with the SARS virus. The M protein of both COVID-19 and MERS displayed variations in the structure. The interaction between M and E protein was also studied to know the additional binding regions. Our study highlights the critical motifs and structural regions to be considered for further research to design better inhibitors for the infection caused by these viruses.

Genomic analysis of Helicobacter himalayensis sp. nov. isolated from Marmota himalayana

Background Helicobacter himalayensis was isolated from Marmota himalayana in the Qinghai-Tibet Plateau, China, and is a new non-H. pylori species, with unclear taxonomy, phylogeny, and pathogenicity. Results A comparative genomic analysis was performed between the H. himalayensis type strain 80(YS1)T and other the genomes of Helicobacter species present in the National Center for Biotechnology Information (NCBI) database to explore the molecular evolution and potential pathogenicity of H. himalayensis. H. himalayensis 80(YS1)T formed a clade with H. cinaedi and H. hepaticus that was phylogenetically distant from H. pylori. The H. himalayensis genome showed extensive collinearity with H. hepaticus and H. cinaedi. However, it also revealed a low degree of genome collinearity with H. pylori. The genome of 80(YS1)T comprised 1,829,936 bp, with a 39.89% GC content, a predicted genomic island, and 1769 genes. Comparatively, H. himalayensis has more genes for functions in “cell wall/membrane/envelope biogenesis” and “coenzyme transport and metabolism” sub-branches than the other compared helicobacters, and its genome contained 42 virulence factors genes, including that encoding cytolethal distending toxin (CDT). Conclusions We characterized the H. himalayensis 80(YS1)T genome, its phylogenetic position, and its potential pathogenicity. However, further understanding of the pathogenesis of this potentially pathogenic bacterium is required, which might help to manage H. himalayensis-induced diseases.