Tissue-colonizing disseminated tumor cells secrete prostaglandin E2 to promote NK cell dysfunction and evade anti-metastatic immunity.

Natural killer (NK) cells are critical for anti-metastatic immunity and can eliminate metastasizing tumor cells within circulation and sites of metastatic seeding. Here, we show that disseminated tumor cells (DTCs) colonizing the mouse lung secrete prostaglandin E2 (PGE) to locally induce NK cell dysfunction, allowing outgrowing metastases to escape immune control and establish metastatic disease. Mechanistically, PGE signaling through its receptors EP2 and EP4 mediates NK cell dysfunction, which leads to reprogramming of NK cell gene expression and results in impaired production of anti-metastatic cytokines.

Orthogonal translation with 5-cyanotryptophan as an infrared probe for local structural information, electrostatics, and hydrogen bonding.

Orthogonal translation is an efficient tool that provides many valuable spectral probes capable of covering different parts of the electromagnetic spectrum and thus enabling parameterization of various structural and dynamic phenomena in proteins. In this context, nitrile-containing tryptophan analogs are very useful probes to study local electrostatics and hydrogen bonding in both rigid and dynamic environments. Here, we report a semi-rational approach to engineer a tyrosyl-tRNA synthetase (TyrRS) variant of Methanocaldococcus jannaschii capable of incorporating 5-cyanotryptophan (5CNW) via orthogonal translation.

Engineered bacterial host for genetic encoding of physiologically stable protein nitration.

Across scales, many biological phenomena, such as protein folding or bioadhesion and cohesion, rely on synergistic effects of different amino acid side chains at multiple positions in the protein sequence. These are often fine-tuned by post-translational modifications that introduce additional chemical properties. Several PTMs can now be genetically encoded and precisely installed at single and multiple sites by genetic code expansion.

Efficient Unnatural Protein Production by Pyrrolysyl-tRNA Synthetase With Genetically Fused Solubility Tags.

Introducing non-canonical amino acids (ncAAs) by engineered orthogonal pairs of aminoacyl-tRNA synthetases and tRNAs has proven to be a highly useful tool for the expansion of the genetic code. Pyrrolysyl-tRNA synthetase (PylRS) from methanogenic archaeal and bacterial species is particularly attractive due to its natural orthogonal reactivity in bacterial and eukaryotic cells. However, the scope of such a reprogrammed translation is often limited, due to low yields of chemically modified target protein.

Using advanced racial and ethnic identity demographics to improve surveillance of work-related conditions in an occupational clinic setting.

Background: Although racial and ethnic identities are associated with a multitude of disparate medical outcomes, surveillance of these subpopulations in the occupational clinic setting could benefit enormously from a more detailed and nuanced recognition of racial and ethnic identity.

Methods: The research group designed a brief questionnaire to capture several dimensions of this identity and collected data from patients seen for work-related conditions in four occupational medicine clinics from May 2019 through March 2020. Responses were used to calculate the sensitivity and specificity of extant racial/ethnic identity data within our electronic health records system, and were compared to participants' self-reported industry and occupation, coded according to North American Industry Classification System and Standard Occupational Classification System listings.

IL-6-induced FOXO1 activity determines the dynamics of metabolism in CD8 T cells cross-primed by liver sinusoidal endothelial cells.

Liver sinusoidal endothelial cells (LSECs) are liver-resident antigen (cross)-presenting cells that generate memory CD8 T cells, but metabolic properties of LSECs and LSEC-primed CD8 T cells remain understudied. Here, we report that high-level mitochondrial respiration and constitutive low-level glycolysis support LSEC scavenger and sentinel functions. LSECs fail to increase glycolysis and co-stimulation after TLR4 activation, indicating absence of metabolic and functional maturation compared with immunogenic dendritic cells.

Expanding the Scope of Orthogonal Translation with Pyrrolysyl-tRNA Synthetases Dedicated to Aromatic Amino Acids.

In protein engineering and synthetic biology, pyrrolysyl-tRNA synthetase (PylRS), with its cognate tRNA, is one of the most popular tools for site-specific incorporation of non-canonical amino acids (ncAAs). Numerous orthogonal pairs based on engineered PylRS variants have been developed during the last decade, enabling a substantial genetic code expansion, mainly with aliphatic pyrrolysine analogs. However, comparatively less progress has been made to expand the substrate range of PylRS towards aromatic amino acid residues.

Virtual Quality Control in Middle Ear Surgery: Is Image-guided Tympanoscopy a Valuable Tool for Depicting Borderline Situations?

Hypothesis: Before modern imaging was introduced, revision surgery was the only way to evaluate possible reasons for inadequate improvement in hearing after ossicular replacement during reconstructive middle ear surgery.

Background: The aim of this study was to evaluate freely navigable virtual tympanoscopy using different computed tomographic modalities. We compared cone-beam computed tomography (CBCT), flat panel computed tomography (FPCT), and conventional computed tomography in helical mode (CTH), volume mode (CTV), and ultra high resolution mode (CTD).

Regulatory myeloid cells paralyze T cells through cell-cell transfer of the metabolite methylglyoxal.

Regulatory myeloid immune cells, such as myeloid-derived suppressor cells (MDSCs), populate inflamed or cancerous tissue and block immune cell effector functions. The lack of mechanistic insight into MDSC suppressive activity and a marker for their identification has hampered attempts to overcome T cell inhibition and unleash anti-cancer immunity. Here, we report that human MDSCs were characterized by strongly reduced metabolism and conferred this compromised metabolic state to CD8 T cells, thereby paralyzing their effector functions.

Red, Orange, Green: Light- and Temperature-Dependent Color Tuning in a Cyanobacteriochrome.

Cyanobacteriochromes (CBCRs) are photoreceptor proteins that photoconvert between two parent states and thereby regulate various biological processes. An intriguing property is their variable ultraviolet-visible (UV-vis) absorption that covers the entire spectral range from the far-red to the near-UV region and thus makes CBCRs promising candidates for optogenetic applications. Here, we have studied Slr1393, a CBCR that photoswitches between red- and green-absorbing states (Pr and Pg, respectively).

rAAV Engineering for Capsid-Protein Enzyme Insertions and Mosaicism Reveals Resilience to Mutational, Structural and Thermal Perturbations.

Recombinant adeno-associated viruses (rAAV) provide outstanding options for customization and superior capabilities for gene therapy. To access their full potential, facile genetic manipulation is pivotal, including capsid loop modifications. Therefore, we assessed capsid tolerance to modifications of the structural VP proteins in terms of stability and plasticity.

Computational Aminoacyl-tRNA Synthetase Library Design for Photocaged Tyrosine.

Engineering aminoacyl-tRNA synthetases (aaRSs) provides access to the ribosomal incorporation of noncanonical amino acids via genetic code expansion. Conventional targeted mutagenesis libraries with 5-7 positions randomized cover only marginal fractions of the vast sequence space formed by up to 30 active site residues. This frequently results in selection of weakly active enzymes.

Site-Resolved Observation of Vibrational Energy Transfer Using a Genetically Encoded Ultrafast Heater.

Allosteric information transfer in proteins has been linked to distinct vibrational energy transfer (VET) pathways in a number of theoretical studies. Experimental evidence for such pathways, however, is sparse because site-selective injection of vibrational energy into a protein, that is, localized heating, is required for their investigation. Here, we solved this problem by the site-specific incorporation of the non-canonical amino acid β-(1-azulenyl)-l-alanine (AzAla) through genetic code expansion.

Antimicrobial Peptides Produced by Selective Pressure Incorporation of Non-canonical Amino Acids.

Nature has a variety of possibilities to create new protein functions by modifying the sequence of the individual amino acid building blocks. However, all variations are based on the 20 canonical amino acids (cAAs). As a way to introduce additional physicochemical properties into polypeptides, the incorporation of non-canonical amino acids (ncAAs) is increasingly used in protein engineering.

Engineering 'Golden' Fluorescence by Selective Pressure Incorporation of Non-canonical Amino Acids and Protein Analysis by Mass Spectrometry and Fluorescence.

Fluorescent proteins are fundamental tools for the life sciences, in particular for fluorescence microscopy of living cells. While wild-type and engineered variants of the green fluorescent protein from Aequorea victoria (avGFP) as well as homologs from other species already cover large parts of the optical spectrum, a spectral gap remains in the near-infrared region, for which avGFP-based fluorophores are not available. Red-shifted fluorescent protein (FP) variants would substantially expand the toolkit for spectral unmixing of multiple molecular species, but the naturally occurring red-shifted FPs derived from corals or sea anemones have lower fluorescence quantum yield and inferior photo-stability compared to the avGFP variants.

Expanding the Genetic Code of and to Incorporate Non-canonical Amino Acids for Production of Modified Lantibiotics.

The incorporation of non-canonical amino acids (ncAAs) into ribosomally synthesized and post-translationally modified peptides, e.g., nisin from the Gram-positive bacterium , bears great potential to expand the chemical space of various antimicrobials.

Experimental Simulation of Clinical Borderline Situations in Temporal Bone Specimens After Ossiculoplasty.

Objectives: One reason for insufficient hearing improvement with a distinct air-bone gap after ossiculoplasty with implantation of partial or total ossicular replacement prostheses can be the dislocation or minimal shifting of the prosthesis. The aim of this study was the simulation of common clinical borderline situations with minimal shifting of the prosthesis in temporal bone specimens after ossiculoplasty. It was furthermore the goal to identify these specific situations through imaging by cone beam computed tomography (cbCT) and direct visual inspection using the operation microscope.

Photoactivatable Mussel-Based Underwater Adhesive Proteins by an Expanded Genetic Code.

Marine mussels exhibit potent underwater adhesion abilities under hostile conditions by employing 3,4-dihydroxyphenylalanine (DOPA)-rich mussel adhesive proteins (MAPs). However, their recombinant production is a major biotechnological challenge. Herein, a novel strategy based on genetic code expansion has been developed by engineering efficient aminoacyl-transfer RNA synthetases (aaRSs) for the photocaged noncanonical amino acid ortho-nitrobenzyl DOPA (ONB-DOPA).

Prospects of Incorporation of Non-canonical Amino Acids for the Chemical Diversification of Antimicrobial Peptides.

The incorporation of non-canonical amino acids (ncAA) is an elegant way for the chemical diversification of recombinantly produced antimicrobial peptides (AMPs). Residue- and site-specific installation methods in several bacterial production hosts hold great promise for the generation of new-to-nature AMPs, and can contribute to tackle the ongoing emergence of antibiotic resistance in pathogens. Especially from a pharmacological point of view, desirable improvements span pH and protease resistance, solubility, oral availability and circulation half-life.

Discovery and Investigation of Natural Editing Function against Artificial Amino Acids in Protein Translation.

Fluorine being not substantially present in the chemistry of living beings is an attractive element in tailoring novel chemical, biophysical, and pharmacokinetic properties of peptides and proteins. The hallmark of ribosome-mediated artificial amino acid incorporation into peptides and proteins is a broad substrate tolerance, which is assumed to rely on the absence of evolutionary pressure for efficient editing of artificial amino acids. We used the well-characterized editing proficient isoleucyl-tRNA synthetase (IleRS) from to investigate the crosstalk of aminoacylation and editing activities against fluorinated amino acids.

Rapid laccolith intrusion driven by explosive volcanic eruption.

Magmatic intrusions and volcanic eruptions are intimately related phenomena. Shallow magma intrusion builds subsurface reservoirs that are drained by volcanic eruptions. Thus, the long-held view is that intrusions must precede and feed eruptions.

Orthogonal Protein Translation Using Pyrrolysyl-tRNA Synthetases for Single- and Multiple-Noncanonical Amino Acid Mutagenesis.

To date, the two systems most extensively used for noncanonical amino acid (ncAA) incorporation via orthogonal translation are based on the Methanococcus jannaschii TyrRS/tRNA and the Methanosarcina barkeri/Methanosarcina mazei PylRS/tRNA pairs. Here, we summarize the development and usage of the pyrrolysine-based system for orthogonal translation, a process that allows for the recombinant production of site-specifically labeled proteins and peptides. Via stop codon suppression in Escherichia coli and mammalian cells, genetically encoded biomolecules can be equipped with a great diversity of chemical functionalities including click chemistry handles, post-translational modifications, and photocaged sidechains.

Echocardiographic B-mode evaluation of the right heart in older people: The ActiFE Study.

Purpose: Right heart B-mode echocardiography reference values have been predominantly obtained in younger age cohorts (<65years). Aims of the study were to describe and evaluate the feasibility of right heart B-mode echocardiography in a large geriatric cohort and to analyze standard parameters in defined subgroups with or without any evidence for structural cardiac disease.

Methods: ActiFE-Ulm study is a population-based cohort study including a sample of people≥65years old living in the region of Ulm, Germany.

Molecular Epidemiology of Carbapenem-Resistant Acinetobacter Baumannii Complex Isolates from Patients that were Injured During the Eastern Ukrainian Conflict.

This study addressed carbapenem-resistant Acinetobacter baumannii complex (ABC) isolates from patients that were injured during the military conflict in the Eastern Ukraine and treated at German Armed Forces Hospitals in 2014 and 2015. Clonal diversity of the strains and potential ways of transmission were analyzed. Patients with one or several isolation events of carbapenem-resistant ABC were included.