Maturation and persistence of the anti-SARS-CoV-2 memory B cell response

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In vitro and in vivo characterization of an interleukin‐15 antagonist peptide by metabolic stability, 99mTc‐labeling,and biological activity assays

Interleukin (IL)–15 is an inflammatory cytokine that constitutes a validated therapeutic target in some immunopathologies, including rheumatoid arthritis (RA). Previously, we identified an IL‐15 antagonist peptide named [K6T]P8, with potential therapeutic application in RA. In the current work, the metabolic stability of this peptide in synovial fluids from RA patients was studied. Moreover, [K6T]P8 peptide was labeled with ^99mTc to investigate its stability in human plasma and its biodistribution pattern in healthy rats. The biological activity of [K6T]P8 peptide and its dimer was evaluated in CTLL‐2 cells, using 3 different additives to improve the solubility of these peptides. The half‐life of [K6T]P8 in human synovial fluid was 5.88 ± 1.73 minutes, and the major chemical modifications included peptide dimerization, cysteinylation, and methionine oxidation. Radiolabeling of [K6T]P8 with ^99mTc showed a yield of approximately 99.8%. The ^99mTc‐labeled peptide was stable in a 30‐fold molar excess of cysteine and in human plasma, displaying a low affinity to plasma proteins. Preliminary biodistribution studies in healthy Wistar rats suggested a slow elimination of the peptide through the renal and hepatic pathways. Although citric acid, sucrose, and Tween 80 enhanced the solubility of [K6T]P8 peptide and its dimer, only the sucrose did not interfere with the in vitro proliferation assay used to assess their biological activity. The results here presented, reinforce nonclinical characterization of the [K6T]P8 peptide, a potential agent for the treatment of RA and other diseases associated with IL‐15 overexpression.     

Structures of P. falciparum protein kinase 7 identify an activation motif and leads for inhibitor design

Malaria is a major threat to world health. The identification of parasite targets for drug development is a priority and parasitic protein kinases suggest themselves as suitable targets as many display profound structural and functional divergences from their host counterparts. In this paper, we describe the structure of the orphan protein kinase, Plasmodium falciparum protein kinase 7 (PFPK7). Several Plasmodium protein kinases contain extensive insertions, and the structure of PFPK7 reveals how these may be accommodated as excursions from the canonical eukaryotic protein kinase fold. The constitutively active conformation of PFPK7 is stabilized by a structural motif in which the role of the conserved phosphorylated residue that assists in structuring the activation loop of many protein kinases is played by an arginine residue. We identify two series of PFPK7 ATP-competitive inhibitors and suggest further developments for the design of selective and potent PFPK7 lead compounds as potential antimalarials.     

Analysis of the polymerization initiation and activity of Pasteurella multocida heparosan synthase PmHS2, an enzyme with glycosyltransferase and UDP-sugar hydrolase activity

Heparosan synthase catalyzes the polymerization of heparosan (-4GlcUAβ1-4GlcNAcα1-)(n) by transferring alternatively the monosaccharide units from UDP-GlcUA and UDP-GlcNAc to an acceptor molecule. Details on the heparosan chain initiation by Pasteurella multocida heparosan synthase PmHS2 and its influence on the polymerization process have not been reported yet. By site-directed mutagenesis of PmHS2, the single action transferases PmHS2-GlcUA(+) and PmHS2-GlcNAc(+) were obtained. When incubated together in the standard polymerization conditions, the PmHS2-GlcUA(+)/PmHS2-GlcNAc(+) showed comparable polymerization properties as determined for PmHS2. We investigated the first step occurring in heparosan chain initiation by the use of the single action transferases and by studying the PmHS2 polymerization process in the presence of heparosan templates and various UDP-sugar concentrations. We observed that PmHS2 favored the initiation of the heparosan chains when incubated in the presence of an excess of UDP-GlcNAc. It resulted in a higher number of heparosan chains with a lower average molecular weight or in the synthesis of two distinct groups of heparosan chain length, in the absence or in the presence of heparosan templates, respectively. These data suggest that PmHS2 transfers GlcUA from UDP-GlcUA moiety to a UDP-GlcNAc acceptor molecule to initiate the heparosan polymerization; as a consequence, not only the UDP-sugar concentration but also the amount of each UDP-sugar is influencing the PmHS2 polymerization process. In addition, it was shown that PmHS2 hydrolyzes the UDP-sugars, UDP-GlcUA being more degraded than UDP-GlcNAc. However, PmHS2 incubated in the presence of both UDP-sugars favors the synthesis of heparosan polymers over the hydrolysis of UDP-sugars.     

Evolution and Molecular Epidemiology of Citrus tristeza virus on Crete: Recent Introduction of a Severe Strain

Sporadic incidences of Citrus tristeza virus (CTV) in western Crete resulting from the introduction of a mild strain (Spanish isolate T385) have been reported previously. Further analysis within this region has identified an emerging second CTV strain with minimal genetic divergence, sharing 99% nucleotide identity with the severe stem‐pitting isolate Taiwan‐Pum/SP/T1. Other severe isolates from the Mediterranean region appear in the same phylogenetic cluster, indicating movement or new introductions and the need for targeted control actions and improved phytosanitary measures in this area.     

Biochemical Characterization of the Bi-lobe Reveals a Continuous Structural Network Linking the Bi-lobe to Other Single-copied Organelles in Trypanosoma brucei

Trypanosoma brucei, a unicellular parasite, contains several single-copied organelles that duplicate and segregate in a highly coordinated fashion during the cell cycle. In the procyclic stage, a bi-lobed structure is found adjacent to the single ER exit site and Golgi apparatus, forming both stable and dynamic association with other cytoskeletal components including the basal bodies that seed the flagellum and the flagellar pocket collar that is critical for flagellar pocket biogenesis. To further understand the bi-lobe and its association with adjacent organelles, we performed proteomic analyses on the immunoisolated bi-lobe complex. Candidate proteins were localized to the flagellar pocket, the basal bodies, a tripartite attachment complex linking the basal bodies to the kinetoplast, and a segment of microtubule quartet linking the flagellar pocket collar and bi-lobe to the basal bodies. These results supported an extensive connection among the single-copied organelles in T. brucei, a strategy employed by the parasite for orderly organelle assembly and inheritance during the cell cycle.     

Walking Performance: Correlation between Energy Cost of Walking and Walking Participation. New Statistical Approach Concerning Outcome Measurement

Walking ability, though important for quality of life and participation in social and economic activities, can be adversely affected by neurological disorders, such as Spinal Cord Injury, Stroke, Multiple Sclerosis or Traumatic Brain Injury. The aim of this study is to evaluate if the energy cost of walking (CW), in a mixed group of chronic patients with neurological diseases almost 6 months after discharge from rehabilitation wards, can predict the walking performance and any walking restriction on community activities, as indicated by Walking Handicap Scale categories (WHS). One hundred and seven subjects were included in the study, 31 suffering from Stroke, 26 from Spinal Cord Injury and 50 from Multiple Sclerosis. The multivariable binary logistical regression analysis has produced a statistical model with good characteristics of fit and good predictability. This model generated a cut-off value of.40, which enabled us to classify correctly the cases with a percentage of 85.0%. Our research reveal that, in our subjects, CW is the only predictor of the walking performance of in the community, to be compared with the score of WHS. We have been also identifying a cut-off value of CW cost, which makes a distinction between those who can walk in the community and those who cannot do it. In particular, these values could be used to predict the ability to walk in the community when discharged from the rehabilitation units, and to adjust the rehabilitative treatment to improve the performance.     

Morphological and Molecular Characterization of Dietary-Induced Pseudo-Albinism during Post-Embryonic Development of Solea senegalensis (Kaup, 1858)

The appearance of the pseudo-albino phenotype was investigated in developing Senegalese sole (Solea senegalensis, Kaup 1858) larvae at morphological and molecular levels. In order to induce the development of pseudo-albinos, Senegalese sole larvae were fed Artemia enriched with high levels of arachidonic acid (ARA). The development of their skin pigmentation was compared to that of a control group fed Artemia enriched with a reference commercial product. The relative amount of skin melanophores, xanthophores and iridophores revealed that larval pigmentation developed similarly in both groups. However, results from different relative proportions, allocation patterns, shapes and sizes of skin chromatophores revealed changes in the pigmentation pattern between ARA and control groups from 33 days post hatching onwards. The new populations of chromatophores that should appear at post-metamorphosis were not formed in the ARA group. Further, spatial patterns of distribution between the already present larval xanthophores and melanophores were suggestive of short-range interaction that seemed to be implicated in the degradation of these chromatophores, leading to the appearance of the pseudo-albino phenotype. The expression profile of several key pigmentation-related genes revealed that melanophore development was promoted in pseudo-albinos without a sufficient degree of terminal differentiation, thus preventing melanogenesis. Present results suggest the potential roles of asip1 and slc24a5 genes on the down-regulation of trp1 expression, leading to defects in melanin production. Moreover, gene expression data supports the involvement of pax3, mitf and asip1 genes in the developmental disruption of the new post-metamorphic populations of melanophores, xanthophores and iridophores.     

Structure and regulation of the <Emphasis Type="Italic">Asr</Emphasis> gene family in banana

Abscisic acid, stress, ripening proteins (ASR) are a family of plant-specific small hydrophilic proteins. Studies in various plant species have highlighted their role in increased resistance to abiotic stress, including drought, but their specific function remains unknown. As a first step toward their potential use in crop improvement, we investigated the structure and regulation of the Asr gene family in Musa species (bananas and plantains). We determined that the Musa Asr gene family contained at least four members, all of which exhibited the typical two exons, one intron structure of Asr genes and the “ABA/WDS” (abscisic acid/water deficit stress) domain characteristic of Asr genes. Phylogenetic analyses determined that the Musa Asr genes were closely related to each other, probably as the product of recent duplication events. For two of the four members, two versions corresponding to the two sub-genomes of Musa, acuminata and balbisiana were identified. Gene expression and protein analyses were performed and Asr expression could be detected in meristem cultures, root, pseudostem, leaf and cormus. In meristem cultures, mAsr1 and mAsr3 were induced by osmotic stress and wounding, while mAsr3 and mAsr4 were induced by exposure to ABA. mASR3 exhibited the most variation both in terms of amino acid sequence and expression pattern, making it the most promising candidate for further functional study and use in crop improvement.