Colonization of nasal cavities by Staphylococcus epidermidis mitigates SARS-CoV-2 nucleocapsid phosphoprotein-induced interleukin (IL)-6 in the lung

Infection by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can trigger excessive interleukin (IL)-6 signalling, leading to a myriad of biological effects including a cytokine storm that contributes to multiple organ failure in severe coronavirus disease 2019 (COVID-19). Using a mouse model, we demonstrated that nasal inoculation of nucleocapsid phosphoprotein (NPP) of SARS-CoV-2 increased IL-6 content in bronchoalveolar lavage fluid (BALF). Nasal administration of liquid coco-caprylate/caprate (LCC) onto Staphylococcus epidermidis (S. epidermidis)-colonized mice significantly attenuated NPP-induced IL-6. Furthermore, S. epidermidis-mediated LCC fermentation to generate electricity and butyric acid that promoted bacterial colonization and activated free fatty acid receptor 2 (Ffar2) respectively. Inhibition of Ffar2 impeded the effect of S. epidermidis plus LCC on the reduction of NPP-induced IL-6. Collectively, these results suggest that nasal S. epidermidis is part of the first line of defence in ameliorating a cytokine storm induced by airway infection of SARS-CoV-2.     

Histone deacetylase complexes: functional entities or molecular reservoirs

Using the dextran-binding domain (DBD) of a type of glucosyltransferase (GTF) from Streptococcus sobrinus, we have developed a novel method for purifying recombinant proteins. DBD-tagged green and red fluorescent proteins as well as the parent GTF and DBD moiety were adsorbed well to commercially available cross-linked dextran (such as Sephadex beads and Sephacryl beads), and eluted efficiently with water-soluble dextran. The purity of the eluted proteins after this one-step affinity purification was 90% or better. The results suggest that DBD can be used as a powerful carrier for purification of various recombinant proteins.     

Relative importance of Na+, Cl−, and abscisic acid in NaCl induced inhibition of root growth of rice seedlings

The relative importance of endogenous abscisic acid (ABA), as well as Na⁺ and Cl^– in NaCl-induced responses related to growth in roots of rice seedlings were investigated. The increase in ammonium, proline and H₂O₂ levels, and cell wall peroxidase (POD) activity has been shown to be related to NaCl-inhibited root growth of rice seedlings. Increasing concentrations of NaCl from 50 to 150 mM progressively decreased root growth and increased both Na⁺ and Cl^–. Treatment with NaCl in the presence of 4,4-diisothiocyano-2,2-disulfonic acid (DIDS, a nonpermeating amino-reactive disulfonic acid known to inhibit the uptake of Cl^–) had less Cl^– level in roots than that in the absence of DIDS, but did not affect the levels of Na⁺, and responses related to growth in roots. Treatment with 50 mM Na-gluconate (the anion of which is not permeable to membrane) had similar Na⁺ level in roots as that with 100 mM NaCl. It was found that treatment with 50 mM Na-gluconate effected growth reduction and growth-related responses in roots in the same way as 100 mM NaCl. All these results suggest that Cl^– is not required for NaCl-induced responses in root of rice seedlings. Endogenous ABA level showed no increase in roots of rice seedlings exposed to 150 mM NaCl. It is unlikely that ABA is associated with NaCl-inhibited root growth of rice seedlings.     

Molecular dissection of mitogillin reveals that the fungal ribotoxins are a family of natural genetically engineered ribonucleases

Mitogillin and the related fungal ribotoxins are highly specific ribonucleases which inactivate the ribosome enzymatically by cleaving the 23-28 S RNA of the large ribosomal subunit at a single phosphodiester bond. The site of cleavage occurs between G4325 and A4326 (rat ribosome numbering) which are present in one of the most conserved sequences (the alpha-sarcin loop) among the large subunit ribosomal RNAs of all living species. Amino acid sequence comparison of ribotoxins and guanyl/purine ribonucleases have identified domains or residues likely involved in ribonucleolytic activity or cleavage specificity. Fifteen deletion mutants (each 4 to 8 amino acid deletions) in motifs of mitogillin showing little amino acid sequence homology with guanyl/purine ribonucleases were constructed by site-directed mutagenesis. Analyses of the purified mutant proteins identified those regions in fungal ribotoxins contributing to ribosome targeting and modulating the catalytic activity of the toxin; some of the identified motifs are homologous to sequences in ribosomal proteins and elongation factors. This mutational study of mitogillin together with the recently published x-ray structure of restrictocin (a close relative of mitogillin) supports the hypothesis that the specific cleavage properties of ribotoxins are the result of natural genetic engineering in which the ribosomal targeting elements of ribosome-associated proteins were inserted into nonessential regions of T1-like ribonucleases.     

A minimal RNA promoter for minus-strand RNA synthesis by the brome mosaic virus polymerase complex

The approximately 150 nt tRNA-like structure present at the 3' end of each of the brome mosaic virus (BMV) genomic RNAs is sufficient to direct minus-strand RNA synthesis. RNAs containing mutations in the tRNA-like structure that decrease minus-strand synthesis were tested for their ability to interact with RdRp (RNA-dependent RNA polymerase) using a template competition assay. Mutations that are predicted to disrupt the pseudoknot and stem B1 do not affect the ability of the tRNA-like structure to interact with RdRp. Similarly, the +1 and +2 nucleotides are not required for stable template-RdRp interaction. Mutations in the bulge and hairpin loops of stem C decreased the ability of the tRNA-like structure to interact with RdRp. Furthermore, in the absence of the rest of the BMV tRNA, stem C is able to interact with RdRp. The addition of an accessible initiation sequence containing ACCA3' to stem C created an RNA capable of directing RNA synthesis. Synthesis from this minimal minus-strand template is dependent on sequences in the hairpin and bulged loops.     

Autoantibodies define a family of proteins with conserved double-stranded RNA-binding domains as well as DNA binding activity

Cellular responses to viral infection are signaled by double-stranded (ds) RNA, which is not found in substantial amounts in uninfected cells. Although cellular dsRNA-binding proteins have been described, their characterization is incomplete. We show that dsRNA-binding proteins are prominent autoantigens. Sera from B6 and B10.S mice with pristane-induced lupus and human autoimmune sera immunoprecipitated a novel set of 130-, 110-, 90-, 80-, and 45-kDa proteins. The proteins were all major cellular poly(IC)-binding factors. N-terminal amino acid sequences of p110 and p90 were identical and matched nuclear factor (NF) 90 and M phase phosphoprotein 4. p45 and p90 were identified as the NF45.NF90 complex, which binds the interleukin-2 promoter as well as certain highly structured viral RNAs. NF90.NF45 and M phase phosphoprotein 4 belong to a large group of proteins with conserved dsRNA-binding motifs. Besides binding dsRNA, NF90.NF45, p110, and p130 had single-stranded and dsDNA binding activity. Some sera contained autoantibodies whose binding was inhibited by poly(IC) but not single-stranded DNA or vice versa, suggesting that the DNA- and RNA-binding sites are different. These autoantibodies will be useful probes of the function of dsRNA-binding proteins. Their interaction with dsRNA, an immunological adjuvant, also could promote autoimmunity.     

A prevalent POLG CAG microsatellite length allele in humans and African great apes

The human nuclear gene for the catalytic subunit of mitochondrial DNA polymerase (POLG) contains within its coding region a CAG microsatellite encoding a polyglutamine repeat. Previous studies demonstrated an association between length variation at this repeat and male infertility, suggesting a mechanism whereby the prevalent (CAG)₁₀ allele, which occurs at a frequency of >80% in different populations, could be maintained by selection. Sequence analysis of the POLG CAG microsatellite region of more than 1000 human chromosomes reveals that virtually all allelic variation at the locus is accounted for by length variation of the CAG repeat. Analysis of POLG from African great apes shows that a prevalent length allele is present in each species, although its exact length is species-specific. In common chimpanzee (Pan troglodytes) a number of different sequence variants contribute to the prevalent length allele, strongly supporting the idea that the length of the POLG microsatellite region, rather than its exact nucleotide or amino acid sequence, is what is maintained. Analysis of POLG in other primates indicates that the repeat has expanded from a shorter, glutamine-rich sequence, present in the common ancestor of Old and New World monkeys.