Novel intranasal vaccine targeting SARS-CoV-2 receptor binding domain to mucosal microfold cells and adjuvanted with TLR3 agonist Riboxxim™ elicits strong antibody and T-cell responses in mice

SARS-CoV-2 continues to circulate in the human population necessitating regular booster immunization for its long-term control. Ideally, vaccines should ideally not only protect against symptomatic disease, but also prevent transmission via asymptomatic shedding and cover existing and future variants of the virus. This may ultimately only be possible through induction of potent and long-lasting immune responses in the nasopharyngeal tract, the initial entry site of SARS-CoV-2. To this end, we have designed a vaccine based on recombinantly expressed receptor binding domain (RBD) of SARS-CoV-2, fused to the C-terminus of C. perfringens enterotoxin, which is known to target Claudin-4, a matrix molecule highly expressed on mucosal microfold (M) cells of the nasal and bronchial-associated lymphoid tissues. To further enhance immune responses, the vaccine was adjuvanted with a novel toll-like receptor 3/RIG-I agonist (Riboxxim™), consisting of synthetic short double stranded RNA. Intranasal prime-boost immunization of mice induced robust mucosal and systemic anti-SARS-CoV-2 neutralizing antibody responses against SARS-CoV-2 strains Wuhan-Hu-1, and several variants (B.1.351/beta, B.1.1.7/alpha, B.1.617.2/delta), as well as systemic T-cell responses. A combination vaccine with M-cell targeted recombinant HA1 from an H1N1 G4 influenza strain also induced mucosal and systemic antibodies against influenza. Taken together, the data show that development of an intranasal SARS-CoV-2 vaccine based on recombinant RBD adjuvanted with a TLR3 agonist is feasible, also as a combination vaccine against influenza

Evolutionary games on graphs

Game theory is one of the key paradigms behind many scientific disciplines from biology to behavioral sciences to economics. In its evolutionary form and especially when the interacting agents are linked in a specific social network the underlying solution concepts and methods are very similar to those applied in non-equilibrium statistical physics. This review gives a tutorial-type overview of the field for physicists. The first three sections introduce the necessary background in classical and evolutionary game theory from the basic definitions to the most important results. The fourth section surveys the topological complications implied by non-mean-field-type social network structures in general. The last three sections discuss in detail the dynamic behavior of three prominent classes of models: the Prisoner's Dilemma, the Rock-Scissors-Paper game, and Competing Associations. The major theme of the review is in what sense and how the graph structure of interactions can modify and enrich the picture of long term behavioral patterns emerging in evolutionary games.Comment: Review, final version, 133 pages, 65 figure

Evolution of Cooperation in Social Dilemmas: Signaling Internalized Norms

Economists have a long tradition in identifying the evolution of cooperation in large, unstructured societies as a puzzle. We suggest a new explanation for cooperation which avoids restrictions of most previous attempts. Our explanation deals with the role of internalized norms for cooperation in large unstructured populations. Even internalized norms, i.e. norms which alter the perceived utility from acting in a cooperative or in an uncooperative way, will not help to overcome a dilemma in an unstructured society, unless and this is the thrust of the current paper individuals are able to signal their property of being a norm bearer. Only when internalization of the norm may be communicated in a reliable way, the picture may change. We derive necessary and sufficient condition for cooperation to be part of an evolutionary stable equilibrium. These conditions relate signaling cost of norm-adopters and non-adopters, the strength of the social norm and parameter measuring the cost of cooperation

Dictator Games: A Meta Study

Over the last 25 years, more than a hundred dictator game experiments have been published. This meta study summarizes the evidence. Exploiting the fact that most experiments had to fix parameters they did not intend to test, the meta study explores a rich set of control variables for multivariate analysis. It shows that Tobit models (assuming that dictators would even want to take money) and hurdle models (assuming that the decision to give a positive amount is separate from the choice of amount, conditional on giving) outperform mere meta-regression and OLS

Röntgenkristallographische Analyse dreier Membranproteine: Nikotinischer Acetylcholinrezeptor aus Torpedo californica, Omp85 und TtoA aus Thermus thermophilus HB27

The work presented here was focused on the purification, crystallization and X-ray structure determination of three different transmembrane proteins - the nicotinic acetylcholine receptor (nAChR) from Torpedo californica and two outer membrane proteins from Thermus thermophilus HB27, TtOmp85, and one of its potential substrates, TtoA.The nAChR is a ligand-gated ion channel which belongs to the superfamily of Cys-loop receptors and plays a keyrole in synaptic transmission of neuronal signals. nAChRs consist of five homologous subunits composed as tissue-specific homo- or heteropentamers. So far, only a structure of 4.0 Å resolution derived from electron microscopy on tubular 2D-crystals is available for whole nAChR pentamers from Torpedo marmorata. Albeit two high resolution structures of prokaryotic homologues were published recently, a high-resolution structure of the nAChR is still missing. Microcrystals obtained from preparations of detergent-solubilized nAChR-enriched membranes from the electric organ of the pacific ray Torpedo california were reported already in 1988, but the properties of those crystals remained uncharacterized. In the work presented here, such crystals were examined by synchroton radiation and diffraction patterns typical for protein crystals were recorded. Diffraction of those crystals was limited to 20 Å resolution, possible spacegroup and unit-cell parameters were determined with the recorded data. Neither refinement of the crystallization conditions nor optimization of the purification protocol allowed to grow crystals of solubilized nAChR suitable for high resolution X-ray structure determination.Proteins belonging to the Omp85 family are involved in the assembly of beta-barrel outer membrane proteins or in the translocation of proteins across the outer membrane in bacteria, mitochondria and chloroplasts. The thermophilic bacterium Thermus thermophilus represents an intermediate between Gram-positive and Gram-negative bacteria with an only roughly characterized outer membrane. It encodes one Omp85-like protein, TtOmp85, which represents an ancestral type of this family. TtOmp85 was overexpressed in T. thermophilus strain HB27 and natively purified from preparations of the outer membrane. In the presence of detergent, purified TtOmp85 existed mainly as a monomer, composed of two stable protease-resistant modules. Detergent solubilized TtOmp85 was used for crystallization experiments and microcrystals appeared under crystallization conditions including PEG400 as precipitant. Crystals diffracted to 5.0 Å resolution and belonged to spacegroup C222 with unit cell dimensions of a = 96.3 Å, b = 416.1 Å, c = 94.3 Å; alpha = beta = gamma = 90°. Only a partial dataset could be collected because diffraction was anisotropic, depending on the orientation of the plate shaped microcrystals in the X-ray beam, and crystals rapidly accumulated radiation damage during proceeding datacollection. The structure of TtOmp85 could not be solved with the available data. Preparations of detergent solubilized TtOmp85 were provided for accompanying studies in collaborating working groups to further characterize TtOmp85.Only a few further outer membrane proteins from T. thermophilus are described. TtoA, a new beta-barrel outer membrane protein, was identified by bioinformatic analysis of the genome sequence of strain HB27. Results of an in vitro binding assay where TtoA bound to TtOmp85 suggest that insertion of proteins in the outer membrane in T. thermophilus might be similar to the mechanisms found in modern Gram-negative bacteria. TtoA was successfully crystallized and its structure was determined to a resolution of 2.8 Å, representing the first crystal structure of an outer membrane protein from a thermophilic bacterium. Crystals belonged to space group P3(1)21 with unit cell parameters a = b = 166.7 Å, c = 97.5 Å; alpha = beta = 90°, gamma = 120°. TtoA consists of an eight-stranded beta-barrel with a large extracellular part to which a divalent cation is bound. A five-stranded extracellular beta-sheet protrudes out of the membrane-embedded transmembrane barrel and is stabilized by a disulfide bridge. The edge of this beta-sheet forms crystal contacts with neighbouring TtoA molecules that could mimic physiologic interactions with other proteins

Expression, crystallization and preliminary X-ray analysis of an outer membrane protein from Thermus thermophilus HB27

The cell envelope of the thermophilic bacterium Thermus thermophilus is multilayered and includes an outer membrane with integral outer membrane proteins that are not well characterized. The hypothetical protein TTC0834 from T. thermophilus HB27 was identified as a 22 kDa outer membrane protein containing eight predicted -strands. TTC0834 was expressed with an N-terminal His tag in T. thermophilus HB8 and detected in the S-layer/outer membrane envelope fraction. His-TTC0834 was purified and crystallized under various conditions. Native data sets were collected to 3.2 Å resolution and the best diffracting crystals belonged to space group P3121 or P3221, with unit-cell parameters a = b = 166.67, c = 97.53 Å

Crystal Structure of a Major Outer Membrane Protein from Thermus thermophilus HB27

The thermophilic eubacterium Thermus thermophilus belongs to one of the oldest branches of evolution and has a multilayered cell envelope that differs from that of modern Gram-negative bacteria. Its outer membrane contains integral outer membrane proteins (OMPs), of which only a few are characterized. TtoA, a new p-barrel OMP, was identified by searching the genome sequence of strain HB27 for the presence of a C-terminal signature sequence. The structure of TtoA was determined to a resolution of 2.8 Å, representing the first crystal structure of an OMP from a thermophilic bacterium. TtoA consists of an eight-stranded p-barrel with a large extracellular part to which a divalent cation is bound. A five-stranded extracellular β-sheet protrudes out of the membrane-embedded transmembrane barrel and is stabilized by a disulfide bridge. The edge of this β-sheet forms crystal contacts that could mimic interactions with other proteins. In modem Gram-negative bacteria, the C-terminal signature sequence of OMPs is required for binding to an Omp85 family protein as a prerequisite for its assembly. We present hints that a similar assembly pathway exists in T. thermophilus by an in vitro binding assay, where unfolded TtoA binds to the Thermus Omp85 family protein TtOmp85, while a mutant without the signature sequence does not