Marcos Pires

ABSTRACT

Burkholderia pseudomallei and Burkholderia mallei are the causative agents of melioidosis and glanders, respectively. There is no vaccine to protect against these highly pathogenic bacteria, and there is concern regarding their emergence as global public health (B. pseudomallei) and biosecurity (B. mallei) threats. In this issue of mSphere, an article by Khakhum and colleagues (N. Khakhum, P. Bharaj, J. N. Myers, D. Tapia, et al., mSphere 4:e00570-18, 2019, https://doi.org/10.1128/mSphere.00570-18) describes a novel vaccination platform with excellent potential for cross-protection against both Burkholderia species. The report also highlights the importance of antibodies in immunity against these facultative intracellular organisms.

m⁶A modification controls the innate immune response to infection by targeting type I interferons

m⁶A modification controls the innate immune response to infection by targeting type I interferons, Published online: 17 December 2018; doi:10.1038/s41590-018-0275-z

RNAs can be dynamically modified by N6-methylation of adenosine (m6A), which leads to their destabilization. Stern-Ginossar and colleagues demonstrate a role for m6A modification of host transcripts encoding type I interferons during viral infection.

Superresolution, single-particle tracking reveals effects of the cationic antimicrobial peptide LL-37 on the Escherichia coli cytoplasm. Seconds after LL-37 penetrates the cytoplasmic membrane, the chromosomal DNA becomes rigidified on a length scale of ∼30 nm, evidenced by the loss of jiggling motion of specific DNA markers. The diffusive motion of...

Mycobacteria and related organisms in the Corynebacterineae suborder are characterized by a distinctive outer membrane referred to as the mycomembrane. Biosynthesis of the mycomembrane occurs through an essential process called mycoloylation, which involves antigen 85 (Ag85)‐catalyzed transfer of mycolic acids from the mycoloyl donor trehalose monomycolate (TMM) to acceptor carbohydrates and, in some organisms, proteins. We recently described an alkyne‐modified TMM analogue (O‐AlkTMM‐C7) which, in conjunction with click chemistry, acted as a chemical reporter for mycoloylation in intact cells and allowed metabolic labeling of mycoloylated components of the mycomembrane. Here, we describe the synthesis and evaluation of a toolbox of TMM‐based reporters bearing alkyne, azide, trans‐cyclooctene, and fluorescent tags. These compounds gave further insight into the substrate tolerance of mycoloyltransferases (e.g., Ag85s) in a cellular context and they provide significantly expanded experimental versatility by allowing one‐ or two‐step cell labeling, live cell labeling, and rapid cell labeling via tetrazine ligation. Such capabilities will facilitate research on mycomembrane composition, biosynthesis, and dynamics. Moreover, because TMM is exclusively metabolized by Corynebacterineae, the described probes may be valuable for the specific detection and cell‐surface engineering of Mycobacterium tuberculosis and related pathogens.

Staple reaction: A multicomponent method allows the late‐stage derivatization of peptides with the incorporation of sugars, fluorescent and affinity labels, PEGs, steroids, and lipids at both the peptide N‐terminus and Lys side‐chains. Stapled peptides with rigid aromatic linkages can also be produced, proving the versatility of this multicomponent reaction in most areas of peptide chemistry.

Abstract

For the first time, the Petasis (borono‐Mannich) reaction is employed for the multicomponent labeling and stapling of peptides. The report includes the solid‐phase derivatization of peptides at the N‐terminus, Lys, and N ^ϵ ‐MeLys side‐chains by an on‐resin Petasis reaction with variation of the carbonyl and boronic acid components. Peptides were simultaneously functionalized with aryl/vinyl substituents bearing fluorescent/affinity tags and oxo components such as dihydroxyacetone, glyceraldehyde, glyoxylic acid, and aldoses, thus encompassing a powerful complexity‐generating approach without changing the charge of the peptides. The multicomponent stapling was conducted in solution by linking N ^ϵ ‐MeLys or Orn side‐chains, positioned at i, i+7 and i, i+4, with aryl tethers, while hydroxy carbonyl moieties were introduced as exocyclic fragments. The good efficiency and diversity oriented character of these methods show prospects for peptide drug discovery and chemical biology.

Weapon of choice: Penicillin‐binding proteins and serine β‐lactamases, which employ nucleophilic serines, hydrolyze β‐lactam antibiotics. However, the l,d‐transpeptidases, which employ nucleophilic cysteines, also fragment some β‐lactams. Serine β‐lactamases where the nucleophilic serine is substituted with cysteine also catalyze β‐lactam fragmentation, consistent with a mechanism involving the formation of thioester‐enolate intermediates.

Abstract

Enzymes often use nucleophilic serine, threonine, and cysteine residues to achieve the same type of reaction; the underlying reasons for this are not understood. While bacterial d,d‐transpeptidases (penicillin‐binding proteins) employ a nucleophilic serine, l,d‐transpeptidases use a nucleophilic cysteine. The covalent complexes formed by l,d‐transpeptidases with some β‐lactam antibiotics undergo non‐hydrolytic fragmentation. This is not usually observed for penicillin‐binding proteins, or for the related serine β‐lactamases. Replacement of the nucleophilic serine of serine β‐lactamases with cysteine yields enzymes which fragment β‐lactams via a similar mechanism as the l,d‐transpeptidases, implying the different reaction outcomes are principally due to the formation of thioester versus ester intermediates. The results highlight fundamental differences in the reactivity of nucleophilic serine and cysteine enzymes, and imply new possibilities for the inhibition of nucleophilic enzymes.

The composition of the gut microbiota is largely determined by environmental factors including the host diet. Dietary components are believed to influence the composition of the gut microbiota by serving as nutrients to a subset of microbes, thereby favoring their expansion. However, we now report that dietary fructose and glucose,...

In PNAS, Kaplanov et al. (1) report on how IL-1β orchestrates recruitment of immunosuppressive monocytes and their polarization in IL-10–producing macrophages during tumor development. This mechanism consequently inhibits and limits CD8+ T cell-driven immune responses and enables tumor outgrowth. Therapeutic blockade of IL-1β and inhibition of monocyte recruitment synergized and...

The role of the microbiota in infectious diseases

The role of the microbiota in infectious diseases, Published online: 13 December 2018; doi:10.1038/s41564-018-0278-4

This Review discusses the role of the gut, respiratory tract and vaginal microbiota in susceptibility and resistance to infectious diseases.

Bacteriophages can “listen in” on their host bacterium’s quorum-sensing systems, shifting their lysis-lysogeny decisions based on host cell density.

e^‐‐‐lucidation of structures: The microcrystal electron diffraction (MicroED) method promises to significantly accelerate the ability of synthetic chemists to gain structural information about small organic molecules and might be crucial for the acceleration of innovations across many fields.

Abstract

The development of new methods to analyze and determine molecular structures parallels the ability to accelerate synthetic research. For many decades, single‐crystal analysis by X‐ray diffraction (SXRD) has been the definitive tool for structural analysis at the atomic level; the drawback, however, is that a suitable single crystal of the analyte needs to be grown. The recent innovation of the crystalline sponge (CS) method allows the microanalysis of compounds simply soaked in a readily prepared CS crystal, thus circumventing the need to screen crystallization conditions while also using only a trace amount of the sample. In this context, electron diffraction for the structure determination of small molecules is discussed as potentially the next big development in this field.

Gut microbiome structure and metabolic activity in inflammatory bowel disease

Gut microbiome structure and metabolic activity in inflammatory bowel disease, Published online: 10 December 2018; doi:10.1038/s41564-018-0306-4

Using metabolomics and shotgun metagenomics on stool samples from individuals with and without inflammatory bowel disease, metabolites, microbial species and genes associated with disease were identified and validated in an independent cohort.

Intracellular lipopolysaccharide (LPS) triggers the non-canonical inflammasome pathway, resulting in pyroptosis of innate immune cells. In addition to its well-known proinflammatory role, LPS can directly cause regression of some tumors, although the underlying mechanism has remained unknown. Here we show that secretoglobin(SCGB)3A2, a small protein predominantly secreted in airways, chaperones LPS to the cytosol through the cell surface receptor syndecan-1; this leads to pyroptotic cell death driven by caspase-11. SCGB3A2 and LPS co-treatment significantly induced pyroptosis of macrophage RAW264.7 cells and decreased cancer cell proliferation in vitro, while SCGB3A2 treatment resulted in reduced progression of xenograft tumors in mice. These data suggest a conserved function for SCGB3A2 in the innate immune system and cancer cells. These findings demonstrate a critical role for SCGB3A2 as an LPS delivery vehicle; they reveal one mechanism whereby LPS enters innate immune cells leading to pyroptosis, and they clarify the direct effect of LPS on cancer cells.

Host-targeted niclosamide inhibits C. difficile virulence and prevents disease in mice without disrupting the gut microbiota

Host-targeted niclosamide inhibits <i>C. difficile</i> virulence and prevents disease in mice without disrupting the gut microbiota, Published online: 07 December 2018; doi:10.1038/s41467-018-07705-w

Clostridium difficile causes diarrhea and colitis by producing up to three different protein toxins. Here, Tam et al. show that an anthelmintic drug, niclosamide, inhibits the pathogenesis of all three toxins by targeting a host process required for toxin entry into host cells, without disrupting the gut microbiota.

Many bacterial infections are hard to treat and tend to relapse, possibly due to the presence of antibiotic-tolerant persisters. In vitro, persister cells appear to be dormant. After uptake of Salmonella species by macrophages, nongrowing persisters also occur, but their physiological state is poorly understood. In this work, we show that Salmonella persisters arising during macrophage infection maintain a metabolically active state. Persisters reprogram macrophages by means of effectors secreted by the Salmonella pathogenicity island 2 type 3 secretion system. These effectors dampened proinflammatory innate immune responses and induced anti-inflammatory macrophage polarization. Such reprogramming allowed nongrowing Salmonella cells to survive for extended periods in their host. Persisters undermining host immune defenses might confer an advantage to the pathogen during relapse once antibiotic pressure is relieved.

Many discoveries in cell biology rely on making specific proteins visible within their native cellular environment. There are various genetically encoded tags, such as fluorescent proteins, developed for fluorescence microscopy (FM). However, there are almost no genetically encoded tags that enable cellular proteins to be observed by both FM and...

Therapeutic faecal microbiota transplantation controls intestinal inflammation through IL10 secretion by immune cells

Therapeutic faecal microbiota transplantation controls intestinal inflammation through IL10 secretion by immune cells, Published online: 05 December 2018; doi:10.1038/s41467-018-07359-8

Faecal microbiota transplantation (FMT) is becoming a therapeutic option in several gastrointestinal disorders. Here, Burrello et al. study the immunological mechanisms by which FMT reduces colonic inflammation and initiates the restoration of intestinal homeostasis in a mouse model of colitis.

Honey bee Royalactin unlocks conserved pluripotency pathway in mammals

Honey bee Royalactin unlocks conserved pluripotency pathway in mammals, Published online: 04 December 2018; doi:10.1038/s41467-018-06256-4

Royal jelly is the queen-maker for the honey bee that also has effects on longevity, fertility, and regeneration in mammals. Here the authors provide evidence that its major protein component Royalactin, and the mammalian structural analog Regina, maintain pluripotency in mouse ESCs by activating a ground-state pluripotency-like gene network.

Programmable and printable Bacillus subtilis biofilms as engineered living materials

Programmable and printable <i>Bacillus subtilis</i> biofilms as engineered living materials, Published online: 03 December 2018; doi:10.1038/s41589-018-0169-2

Co-opting the amyloid machinery from Bacillus subtilis, engineering of TasA fusion proteins enables the assembly of functionalized biofilms with tunable physicochemical properties that are amenable to 3D printing and microencapsulation techniques.

C−H activation meets metathesis: Bioorthogonal C−H activation of peptides was accomplished by using air‐and moisture‐stable catalysts based on Earth‐abundant cobalt. This reaction set the stage for the combination of C−H functionalization and olefin metathesis to construct complex cyclic peptides.

Abstract

Bioorthogonal late‐stage diversification of structurally complex peptides has enormous potential for drug discovery and molecular imaging. In recent years, transition‐metal‐catalyzed C−H activation has emerged as an increasingly viable tool for peptide modification. Despite major accomplishments, these strategies largely rely on expensive palladium catalysts. We herein report an unprecedented cobalt(III)‐catalyzed peptide C−H activation, which enables the direct C−H functionalization of structurally complex peptides, and sets the stage for a multicatalytic C−H activation/alkene metathesis/hydrogenation strategy for the assembly of novel cyclic peptides.

In bacteria, nascent proteins bear the pretranslationally generated N-terminal (Nt) formyl-methionine (fMet) residue. Nt-fMet of bacterial proteins is a degradation signal, termed fMet/N-degron. By contrast, proteins synthesized by cytosolic ribosomes of eukaryotes were presumed to bear unformylated Nt-Met. Here we found that the yeast formyltransferase Fmt1, although imported into mitochondria, could also produce Nt-formylated proteins in the cytosol. Nt-formylated proteins were strongly up-regulated in stationary phase or upon starvation for specific amino acids. This up-regulation strictly required the Gcn2 kinase, which phosphorylates Fmt1 and mediates its retention in the cytosol. We also found that the Nt-fMet residues of Nt-formylated proteins act as fMet/N-degrons and identified the Psh1 ubiquitin ligase as the recognition component of the eukaryotic fMet/N-end rule pathway, which destroys Nt-formylated proteins.

TRIM5 self-assembles into a lattice that envelopes retroviral capsids, stimulating virion destruction and innate immune signaling. Fletcher et al. demonstrate that self-assembly induces K63-linked ubiquitination (K63-Ub) of the TRIM5 N terminus that drives innate immune stimulation and degradation. Thus, ordered capsid binding allows TRIM5 to couple virus recognition with antiviral responses.