The C-terminus is essential for the stability of the mycobacterial channel protein MspA.

Outer membrane proteins perform essential functions in uptake and secretion processes in bacteria. MspA is an octameric channel protein in the outer membrane of Mycobacterium smegmatis and is structurally distinct from any other known outer membrane protein. MspA is the founding member of a family with more than 3000 homologs and is one of the most widely used proteins in nanotechnological applications due to its advantageous pore structure and extraordinary stability. While a conserved C-terminal signal sequence is essential for folding and protein assembly in the outer membrane of Gram-negative bacteria, the molecular determinants of these processes are unknown for MspA. In this study, we show that mutation and deletion of methionine 183 in the highly conserved C-terminus of MspA and mutation of the conserved tryptophan 40 lead to a complete loss of protein in heat extracts of M. smegmatis. Swapping these residues partially restores the heat stability of MspA indicating that methionine 183 and tryptophan 40 form a conserved sulfur-π electron interaction, which stabilizes the MspA monomer. Flow cytometry showed that all MspA mutants are surface-accessible demonstrating that oligomerization and membrane integration in M. smegmatis are not affected. Thus, the conserved C-terminus of MspA is essential for its thermal stability, but it is not required for protein assembly in its native membrane, indicating that this process is mediated by a mechanism distinct from that in Gram-negative bacteria. These findings will benefit the rational design of MspA-like pores to tailor their properties in current and future applications.

Changes to virus taxonomy and the ICTV Statutes ratified by the International Committee on Taxonomy of Viruses (2023).

This article reports changes to virus taxonomy and taxon nomenclature that were approved and ratified by the International Committee on Taxonomy of Viruses (ICTV) in April 2023. The entire ICTV membership was invited to vote on 174 taxonomic proposals that had been approved by the ICTV Executive Committee in July 2022, as well as a proposed revision of the ICTV Statutes. All proposals and the revised ICTV Statutes were approved by a majority of the voting membership. Of note, the ICTV continued the process of renaming existing species in accordance with the recently mandated binomial format and included gene transfer agents (GTAs) in the classification framework by classifying them as viriforms. In total, one class, seven orders, 31 families, 214 genera, and 858 species were created.

Intrinsic IL-2 production by effector CD8 T cells affects IL-2 signaling and promotes fate decisions, stemness, and protection.

Here, we show that the capacity to manufacture IL-2 identifies constituents of the expanded CD8 T cell effector pool that display stem-like features, preferentially survive, rapidly attain memory traits, resist exhaustion, and control chronic viral challenges. The cell-intrinsic synthesis of IL-2 by CD8 T cells attenuates the ability to receive IL-2-dependent STAT5 signals, thereby limiting terminal effector formation, endowing the IL-2-producing effector subset with superior protective powers. In contrast, the non-IL-2-producing effector cells respond to IL-2 signals and gain effector traits at the expense of memory formation. Despite having distinct properties during the effector phase, IL-2-producing and nonproducing CD8 T cells appear to converge transcriptionally as memory matures to form populations with equal recall abilities. Therefore, the potential to produce IL-2 during the effector, but not memory stage, is a consequential feature that dictates the protective capabilities of the response.

Changes to virus taxonomy and to the International Code of Virus Classification and Nomenclature ratified by the International Committee on Taxonomy of Viruses (2021).

This article reports the changes to virus taxonomy approved and ratified by the International Committee on Taxonomy of Viruses (ICTV) in March 2021. The entire ICTV was invited to vote on 290 taxonomic proposals approved by the ICTV Executive Committee at its meeting in October 2020, as well as on the proposed revision of the International Code of Virus Classification and Nomenclature (ICVCN). All proposals and the revision were ratified by an absolute majority of the ICTV members. Of note, ICTV mandated a uniform rule for virus species naming, which will follow the binomial 'genus-species' format with or without Latinized species epithets. The Study Groups are requested to convert all previously established species names to the new format. ICTV has also abolished the notion of a type species, i.e., a species chosen to serve as a name-bearing type of a virus genus. The remit of ICTV has been clarified through an official definition of 'virus' and several other types of mobile genetic elements. The ICVCN and ICTV Statutes have been amended to reflect these changes.

Sex-based differences in the activation of peripheral blood monocytes in early Parkinson disease.

Increasing evidence supports the role of brain and systemic inflammation in the etiology of Parkinson disease (PD). We used gene expression profiling to examine the activation state of peripheral blood monocytes in 18 patients with early, untreated PD and 16 healthy control (HC) subjects. Monocytes were isolated by negative selection, and gene expression studied by RNA-seq and gene set enrichment analysis. A computational model that incorporated case/control status, sex, and the interaction between case/control status and sex was utilized. We found that there was a striking effect of sex on monocyte gene expression. There was inflammatory activation of monocytes in females with PD, with enrichment of gene sets associated with interferon gamma stimulation. In males, the activation patterns were more heterogeneous. These data point to the importance of systemic monocyte activation in PD, and the importance of studies which examine the differential effects of sex on pathophysiology of the disease.

Changes to virus taxonomy and the Statutes ratified by the International Committee on Taxonomy of Viruses (2020).

This article reports the changes to virus classification and taxonomy approved and ratified by the International Committee on Taxonomy of Viruses (ICTV) in March 2020. The entire ICTV was invited to vote on 206 taxonomic proposals approved by the ICTV Executive Committee at its meeting in July 2019, as well as on the proposed revision of the ICTV Statutes. All proposals and the revision of the Statutes were approved by an absolute majority of the ICTV voting membership. Of note, ICTV has approved a proposal that extends the previously established realm Riboviria to encompass nearly all RNA viruses and reverse-transcribing viruses, and approved three separate proposals to establish three realms for viruses with DNA genomes.

Comprehensive analysis of iron utilization by Mycobacterium tuberculosis.

Iron is essential for nearly all bacterial pathogens, including Mycobacterium tuberculosis (Mtb), but is severely limited in the human host. To meet its iron needs, Mtb secretes siderophores, small molecules with high affinity for iron, and takes up iron-loaded mycobactins (MBT) and carboxymycobactins (cMBT), from the environment. Mtb is also capable of utilizing heme and hemoglobin which contain more than 70% of the iron in the human body. However, many components of these iron acquisition pathways are still unknown. In this study, a high-density transposon mutagenesis coupled with deep sequencing (TnSeq) showed that Mtb exhibits nearly opposite requirements for 165 genes in the presence of heme and hemoglobin versus MBT and cMBT as iron sources. The ESX-3 secretion system was assessed as essential for siderophore-mediated iron uptake and, surprisingly, also for heme utilization by Mtb. Predictions derived from the TnSeq analysis were validated by growth experiments with isogenic Mtb mutants. These results showed that (i) the efflux pump MmpL5 plays a dominant role in siderophore secretion, (ii) the Rv2047c protein is essential for growth of Mtb in the presence of mycobactin, and (iii) the transcriptional repressor Zur is required for heme utilization by Mtb. The novel genetic determinants of iron utilization revealed in this study will stimulate further experiments in this important area of Mtb physiology.

Changes to virus taxonomy and the International Code of Virus Classification and Nomenclature ratified by the International Committee on Taxonomy of Viruses (2019).

This article reports the changes to virus taxonomy approved and ratified by the International Committee on Taxonomy of Viruses (ICTV) in February 2019. Of note, in addition to seven new virus families, the ICTV has approved, by an absolute majority, the creation of the realm Riboviria, a likely monophyletic group encompassing all viruses with positive-strand, negative-strand and double-strand genomic RNA that use cognate RNA-directed RNA polymerases for replication.

Federated Aggregate Cohort Estimator (FACE): an easy to deploy, vendor neutral, multi-institutional cohort query architecture.

Cross-institutional data sharing for cohort discovery is critical to enabling future research. While particularly useful in rare diseases, the ability to target enrollment and to determine if an institution has a sufficient number of patients is valuable in all research, particularly in the initiation of projects and collaborations. An optimal technology solution would work with any source database with minimal resource investment for deployment and would meet all necessary security and confidentiality requirements of participating organizations. We describe a platform-neutral reference implementation to meet these requirements: the Federated Aggregate Cohort Estimator (FACE). FACE was developed and implemented through a collaboration of The University of Alabama at Birmingham (UAB), The Ohio State University (OSU), the University of Massachusetts Medical School (UMMS), and the Denver Health and Hospital Authority (DHHA) a clinical affiliate of the Colorado Clinical and Translational Sciences Institute. The reference implementation of FACE federated diverse SQL data sources and an i2b2 instance to estimate combined research subject availability from three institutions. It used easily-deployed virtual machines and addressed privacy and security concerns for data sharing.

Poxvirus protein evolution: family wide assessment of possible horizontal gene transfer events.

To investigate the evolutionary origins of proteins encoded by the Poxviridae family of viruses, we examined all poxvirus protein coding genes using a method of characterizing and visualizing the similarity between these proteins and taxonomic subsets of proteins in GenBank. Our analysis divides poxvirus proteins into categories based on their relative degree of similarity to two different taxonomic subsets of proteins such as all eukaryote vs. all virus (except poxvirus) proteins. As an example, this allows us to identify, based on high similarity to only eukaryote proteins, poxvirus proteins that may have been obtained by horizontal transfer from their hosts. Although this method alone does not definitively prove horizontal gene transfer, it allows us to provide an assessment of the possibility of horizontal gene transfer for every poxvirus protein. Potential candidates can then be individually studied in more detail during subsequent investigation. Results of our analysis demonstrate that in general, proteins encoded by members of the subfamily Chordopoxvirinae exhibit greater similarity to eukaryote proteins than to proteins of other virus families. In addition, our results reiterate the important role played by host gene capture in poxvirus evolution; highlight the functions of many genes poxviruses share with their hosts; and illustrate which host-like genes are present uniquely in poxviruses and which are also present in other virus families.

Kinetic analysis of a complete poxvirus transcriptome reveals an immediate-early class of genes.

Vaccinia virus is the prototypic orthopoxvirus and was the vaccine used to eradicate smallpox, yet the expression profiles of many of its genes remain unknown. Using a genome tiling array approach, we simultaneously measured the expression levels of all 223 annotated vaccinia virus genes during infection and determined their kinetics. For 95% of these genes, significant transcript levels were detected. Most remarkably, classification of the genes by their expression profiles revealed 35 genes exhibiting immediate-early expression. Although a similar kinetic class has been described for other virus families, to our knowledge, this is the first demonstration of its existence in orthopoxviruses. Despite expression levels higher than for genes in the other three kinetic classes, the functions of more than half of these remain unknown. Additionally, genes within each kinetic class were spatially grouped together in the genome. This genome-wide picture of transcription alters our understanding of how orthopoxviruses regulate gene expression.