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Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/145387/1/cpmia2200.pd

Structural Insights into Rational Design of Single-Domain Antibody-Based Antitoxins against Botulinum Neurotoxins.

Botulinum neurotoxin (BoNT) is one of the most acutely lethal toxins known to humans, and effective treatment for BoNT intoxication is urgently needed. Single-domain antibodies (VHH) have been examined as a countermeasure for BoNT because of their high stability and ease of production. Here, we investigate the structures and the neutralization mechanisms for six unique VHHs targeting BoNT/A1 or BoNT/B1. These studies reveal diverse neutralizing mechanisms by which VHHs prevent host receptor binding or block transmembrane delivery of the BoNT protease domain. Guided by this knowledge, we design heterodimeric VHHs by connecting two neutralizing VHHs via a flexible spacer so they can bind simultaneously to the toxin. These bifunctional VHHs display much greater potency in a mouse co-intoxication model than similar heterodimers unable to bind simultaneously. Taken together, our studies offer insight into antibody neutralization of BoNTs and advance our ability to design multivalent anti-pathogen VHHs with improved therapeutic properties

Polymicrobial periodontal disease triggers a wide radius of effect and unique virome.

Periodontal disease is a microbially-mediated inflammatory disease of tooth-supporting tissues that leads to bone and tissue loss around teeth. Although bacterially-mediated mechanisms of alveolar bone destruction have been widely studied, the effects of a polymicrobial infection on the periodontal ligament and microbiome/virome have not been well explored. Therefore, the current investigation introduced a new mouse model of periodontal disease to examine the effects of a polymicrobial infection on periodontal ligament (PDL) properties, changes in bone loss, the host immune response, and the microbiome/virome using shotgun sequencing. Periodontal pathogens, namely Porphyromonas gingivalis, Treponema denticola, Tannerella forsythia, and Fusobacterium nucleatum were used as the polymicrobial oral inoculum in BALB/cByJ mice. The polymicrobial infection triggered significant alveolar bone loss, a heightened antibody response, an elevated cytokine immune response, a significant shift in viral diversity and virome composition, and a widening of the PDL space; the latter two findings have not been previously reported in periodontal disease models. Changes in the PDL space were present at sites far away from the site of insult, indicating that the polymicrobial radius of effect extends beyond the bone loss areas and site of initial infection and wider than previously appreciated. Associations were found between bone loss, specific viral and bacterial species, immune genes, and PDL space changes. These findings may have significant implications for the pathogenesis of periodontal disease and biomechanical properties of the periodontium. This new polymicrobial mouse model of periodontal disease in a common mouse strain is useful for evaluating the features of periodontal disease

Superfluid to solid crossover in a rotating Bose-Einstein condensed gas

The properties of a rotating Bose-Einstein condensate confined in a prolate cylindrically symmetric trap are explored both analytically and numerically. As the rotation frequency increases, an ever greater number of vortices are energetically favored. Though the cloud anisotropy and moment of inertia approach those of a classical fluid at high frequencies, the observed vortex density is consistently lower than the solid-body estimate. Furthermore, the vortices are found to arrange themselves in highly regular triangular arrays, with little distortion even near the condensate surface. These results are shown to be a direct consequence of the inhomogeneous confining potential.Comment: 4+e pages, 5 embedded figures, revte

A multi-wavelength study of Supernova Remnants in six nearby galaxies. I: Detection of new X-ray selected Supernova Remnants with Chandra

We present results from a study of the Supernova Remnant (SNR) population in a sample of six nearby galaxies (NGC 2403, NGC 3077, NGC 4214, NGC 4449, NGC 4395 and NGC 5204) based on Chandra archival data. We have detected 244 discrete X-ray sources down to a limiting flux of 10^{-15} erg/s. We identify 37 X-ray selected thermal SNRs based on their X-ray colors or spectra, 30 of which are new discoveries. In many cases the X-ray classification is confirmed based on counterparts with SNRs identified in other wavelengths. Three of the galaxies in our sample (NGC 4214, NGC 4395 and NGC 5204) are studied for the first time, resulting in the discovery of 13 thermal SNRs. We discuss the properties (luminosity, temperature, density) of the X-ray detected SNRs in the galaxies of our sample in order to address their dependence on their environment. We find that X-ray selected SNRs in irregular galaxies appear to be more luminous than those in spirals. We attribute this to the lower metalicities and therefore more massive progenitor stars of irregular galaxies or the higher local densities of the ISM. We also discuss the X-ray selected SNR populations in the context of the Star Formation Rate of their host galaxies. A comparison of the numbers of observed luminous X-ray selected SNRs with those expected based on the luminosity functions of X-ray SNRs in the MCs and M33 suggest different luminosity distributions between the SNRs in spiral and irregular galaxies with the latter tending to have flatter distributions.Comment: 56 pages, 14 figures, 26 tables. Accepted for publication in Ap

Accurate classification of 29 objects detected in the 39 months Palermo Swift/BAT hard X-ray catalogue

Through an optical campaign performed at 4 telescopes located in the northern and the southern hemispheres, plus archival data from two on-line sky surveys, we have obtained optical spectroscopy for 29 counterparts of unclassified or poorly studied hard X-ray emitting objects detected with Swift/BAT and listed in the 39 months Palermo catalogue. All these objects have also observations taken with Swift/XRT or XMM-EPIC which not only allow us to pinpoint their optical counterpart, but also to study their X-ray spectral properties (column density, power law photon index and F2-10 keV flux). We find that 28 sources in our sample are AGN; 7 are classified as type 1 while 21 are of type 2; the remaining object is a galactic cataclysmic variable. Among our type 1 AGN, we find 5 objects of intermediate Seyfert type (1.2-1.9) and one Narrow Line Seyfert 1 galaxy; for 4 out of 7 sources, we have been able to estimate the central black hole mass. Three of the type 2 AGN of our sample display optical features typical of the LINER class and one is a likely Compton thick AGN. All galaxies classified in this work are relatively nearby objects since their redshifts lie in the range 0.008-0.075; the only galactic object found lies at an estimated distance of 90 pc. We have also investigated the optical versus X-ray emission ratio of the galaxies of our sample to test the AGN unified model. For them, we have also compared the X-ray absorption (due to gas) with the optical reddening (due to dust): we find that for most of our sources, specifically those of type 1.9-2.0 the former is higher than the latter confirming early results by Maiolino et al. (2001); this is possibly due to the properties of dust in the circumnuclear obscuring torus of the AGN.Comment: 15 pages, 4 figures, 8 tables, accepted for publication on Astronomy and Astrophysic

Unveiling the nature of INTEGRAL objects through optical spectroscopy. VIII. Identification of 44 newly detected hard X-ray sources

(abridged) Hard X-ray surveys performed by the INTEGRAL satellite have discovered a conspicuous fraction (up to 30%) of unidentified objects among the detected sources. Here we continue our identification program by selecting probable optical candidates using positional cross-correlation with soft X-ray, radio, and/or optical archives, and performing optical spectroscopy on them. As a result, we identified or more accurately characterized 44 counterparts of INTEGRAL sources: 32 active galactic nuclei, with redshift 0.019 < z < 0.6058, 6 cataclysmic variables (CVs), 5 high-mass X-ray binaries (2 of which in the Small Magellanic Cloud), and 1 low-mass X-ray binary. This was achieved by using 7 telescopes of various sizes and archival data from two online spectroscopic surveys. The main physical parameters of these hard X-ray sources were also determined using the available multiwavelength information. AGNs are the most abundant population among hard X-ray objects, and our results confirm this tendency when optical spectroscopy is used as an identification tool. The deeper sensitivity of recent INTEGRAL surveys enables one to begin detecting hard X-ray emission above 20 keV from sources such as LINER-type AGNs and non-magnetic CVs.Comment: 22 pages, 14 figures, 6 tables, accepted for publication on A&A, main journa

Interleukin-1 polymorphisms associated with increased risk of gastric cancer

Helicobacter pylori infection is associated with a variety of clinical outcomes including gastric cancer and duodenal ulcer disease. The reasons for this variation are not clear, but the gastric physiological response is influenced by the severity and anatomical distribution of gastritis induced by H. pylori. Thus, individuals with gastritis predominantly localized to the antrum retain normal (or even high) acid secretion, whereas individuals with extensive corpus gastritis develop hypochlorhydria and gastric atrophy, which are presumptive precursors of gastric cancer. Here we report that interleukin-1 gene cluster polymorphisms suspected of enhancing production of interleukin-1-beta are associated with an increased risk of both hypochlorhydria induced by H. pylori and gastric cancer. Two of these polymorphism are in near-complete linkage disequilibrium and one is a TATA-box polymorphism that markedly affects DNA-protein interactions in vitro. The association with disease may be explained by the biological properties of interleukin-1-beta, which is an important pro-inflammatory cytokine and a powerful inhibitor of gastric acid secretion. Host genetic factors that affect interleukin-1-beta may determine why some individuals infected with H. pylori develop gastric cancer while others do no

The Fusion of CLEC12A and MIR223HG Arises from a trans-Splicing Event in Normal and Transformed Human Cells

Chimeric RNAs are often associated with chromosomal rearrangements in cancer. In addition, they are also widely detected in normal tissues, contributing to transcriptomic complexity. Despite their prevalence, little is known about the characteristics and functions of chimeric RNAs. Here, we examine the genetic structure and biological roles of CLEC12A-MIR223HG, a novel chimeric transcript produced by the fusion of the cell surface receptor CLEC12A and the miRNA-223 host gene (MIR223HG), first identified in chronic myeloid leukemia (CML) patients. Surprisingly, we observed that CLEC12A-MIR223HG is not just expressed in CML, but also in a variety of normal tissues and cell lines. CLEC12A-MIR223HG expression is elevated in pro-monocytic cells resistant to chemotherapy and during monocyte-to-macrophage differentiation. We observed that CLEC12A-MIR223HG is a product of trans-splicing rather than a chromosomal rearrangement and that transcriptional activation of CLEC12A with the CRISPR/Cas9 Synergistic Activation Mediator (SAM) system increases CLEC12A-MIR223HG expression. CLEC12A-MIR223HG translates into a chimeric protein, which largely resembles CLEC12A but harbours an altered C-type lectin domain altering key disulphide bonds. These alterations result in differences in post-translational modifications, cellular localization, and protein–protein interactions. Taken together, our observations support a possible involvement of CLEC12A-MIR223HG in the regulation of CLEC12A function. Our workflow also serves as a template to study other uncharacterized chimeric RNAs

The Fusion of CLEC12A and MIR223HG Arises from a trans-Splicing Event in Normal and Transformed Human Cells

Chimeric RNAs are often associated with chromosomal rearrangements in cancer. In addition, they are also widely detected in normal tissues, contributing to transcriptomic complexity. Despite their prevalence, little is known about the characteristics and functions of chimeric RNAs. Here, we examine the genetic structure and biological roles of CLEC12A-MIR223HG, a novel chimeric transcript produced by the fusion of the cell surface receptor CLEC12A and the miRNA-223 host gene (MIR223HG), first identified in chronic myeloid leukemia (CML) patients. Surprisingly, we observed that CLEC12A-MIR223HG is not just expressed in CML, but also in a variety of normal tissues and cell lines. CLEC12A-MIR223HG expression is elevated in pro-monocytic cells resistant to chemotherapy and during monocyte-to-macrophage differentiation. We observed that CLEC12A-MIR223HG is a product of trans-splicing rather than a chromosomal rearrangement and that transcriptional activation of CLEC12A with the CRISPR/Cas9 Synergistic Activation Mediator (SAM) system increases CLEC12A-MIR223HG expression. CLEC12A-MIR223HG translates into a chimeric protein, which largely resembles CLEC12A but harbours an altered C-type lectin domain altering key disulphide bonds. These alterations result in differences in post-translational modifications, cellular localization, and protein–protein interactions. Taken together, our observations support a possible involvement of CLEC12A-MIR223HG in the regulation of CLEC12A function. Our workflow also serves as a template to study other uncharacterized chimeric RNAs