Distinct roles of the three Akt isoforms in lactogenic differentiation and involution

The three Akt isoforms differ in their ability to transduce oncogenic signals initiated by the Neu and PyMT oncogenes in mammary epithelia. As a result, ablation of Akt1 inhibits and ablation of Akt2 accelerates mammary tumor development by both oncogenes, while ablation of Akt3 is phenotypically almost neutral. Since the risk of breast cancer development in humans correlates with multiple late pregnancies, we embarked on a study to determine whether individual Akt isoforms also differ in their ability to transduce hormonal and growth factor signals during pregnancy, lactation and post-lactation involution. The results showed that the ablation of Akt1 delays the differentiation of the mammary epithelia during pregnancy and lactation, and that the ablation of Akt2 has the opposite effect. Finally, ablation of Akt3 results in minor defects, but its phenotype is closer to that of the wild type mice. Whereas the phenotype of the Akt1 ablation is cell autonomous, that of Akt2 is not. The ablation of Akt1 promotes apoptosis and accelerates involution, whereas the ablation of Akt2 inhibits apoptosis and delays involution. Mammary gland differentiation during pregnancy depends on the phosphorylation of Stat5a, which is induced by prolactin, a hormone that generates signals transduced via Akt. Here we show that the ablation of Akt1, but not the ablation of Akt2 or Akt3 interferes with the phosphorylation of Stat5a during late pregnancy and lactation. We conclude that the three Akt isoforms have different roles in mammary gland differentiation during pregnancy and this may reflect differences in hormonal signaling.     

Synthesis and evaluation of novel 3,4,6-substituted 2-quinolones as FMS kinase inhibitors

A series of 3,4,6-substituted 2-quinolones has been synthesized and evaluated as inhibitors of the kinase domain of macrophage colony-stimulating factor-1 receptor (FMS). The fully optimized compound, 4-(4-ethyl-phenyl)-3-(2-methyl-3H-imidazol-4-yl)-2-quinolone-6-carbonitrile 21b, has an IC(50) of 2.5 nM in an in vitro assay and 5.0 nM in a bone marrow-derived macrophage cellular assay. Inhibition of FMS signaling in vivo was also demonstrated in a mouse pharmacodynamic model.     

Complete genome sequence of Treponema succinifaciens type strain (6091)

Treponema succinifaciens Cwyk and Canale-Parola 1981 is of interest because this strictly anaerobic, apathogenic member of the genus Treponema oxidizes carbohydrates and couples the Embden-Meyerhof pathway via activity of a pyruvate-formate lyase to the production of acetyl-coenzyme A and formate. This feature separates this species from most other anaerobic spirochetes. The genome of T. succinifaciens 6091(T) is only the second completed and published type strain genome from the genus Treponema in the family Spirochaetaceae. The 2,897,425 bp long genome with one plasmid harbors 2,723 protein-coding and 63 RNA genes and is a part of the Genomic Encyclopedia of Bacteria and Archaea project.     

iLIR: A web resource for prediction of Atg8-family interacting proteins

Macroautophagy was initially considered to be a nonselective process for bulk breakdown of cytosolic material. However, recent evidence points toward a selective mode of autophagy mediated by the so-called selective autophagy receptors (SARs). SARs act by recognizing and sorting diverse cargo substrates (e.g., proteins, organelles, pathogens) to the autophagic machinery. Known SARs are characterized by a short linear sequence motif (LIR-, LRS-, or AIM-motif) responsible for the interaction between SARs and proteins of the Atg8 family. Interestingly, many LIR-containing proteins (LIRCPs) are also involved in autophagosome formation and maturation and a few of them in regulating signaling pathways. Despite recent research efforts to experimentally identify LIRCPs, only a few dozen of this class of—often unrelated—proteins have been characterized so far using tedious cell biological, biochemical, and crystallographic approaches. The availability of an ever-increasing number of complete eukaryotic genomes provides a grand challenge for characterizing novel LIRCPs throughout the eukaryotes. Along these lines, we developed iLIR, a freely available web resource, which provides in silico tools for assisting the identification of novel LIRCPs. Given an amino acid sequence as input, iLIR searches for instances of short sequences compliant with a refined sensitive regular expression pattern of the extended LIR motif (xLIR-motif) and retrieves characterized protein domains from the SMART database for the query. Additionally, iLIR scores xLIRs against a custom position-specific scoring matrix (PSSM) and identifies potentially disordered subsequences with protein interaction potential overlapping with detected xLIR-motifs. Here we demonstrate that proteins satisfying these criteria make good LIRCP candidates for further experimental verification. Domain architecture is displayed in an informative graphic, and detailed results are also available in tabular form. We anticipate that iLIR will assist with elucidating the full complement of LIRCPs in eukaryotes.     

Outbreaks of SARS-CoV-2 in naturally infected mink farms: Impact,transmission dynamics,genetic patterns,and environmental contamination

SARS-CoV-2 infection outbreaks in minks have serious implications associated with animal health and welfare, and public health. In two naturally infected mink farms (A and B) located in Greece, we investigated the outbreaks and assessed parameters associated with virus transmission, immunity, pathology, and environmental contamination. Symptoms ranged from anorexia and mild depression to respiratory signs of varying intensity. Although the farms were at different breeding stages, mortality was similarly high (8.4% and 10.0%). The viral strains belonged to lineages B.1.1.218 and B.1.1.305, possessing the mink-specific S-Y453F substitution. Lung histopathology identified necrosis of smooth muscle and connective tissue elements of vascular walls, and vasculitis as the main early key events of the acute SARS-CoV-2-induced broncho-interstitial pneumonia. Molecular investigation in two dead minks indicated a consistently higher (0.3–1.3 log₁₀ RNA copies/g) viral load in organs of the male mink compared to the female. In farm A, the infected farmers were responsible for the significant initial infection of 229 out of 1,000 handled minks, suggesting a very efficient human-to-mink transmission. Subsequent infections across the sheds wherein animals were being housed occurred due to airborne transmission. Based on a R₀ of 2.90 and a growth rate equal to 0.293, the generation time was estimated to be 3.6 days, indicative of the massive SARS-CoV-2 dispersal among minks. After the end of the outbreaks, a similar percentage of animals were immune in the two farms (93.0% and 93.3%), preventing further virus transmission whereas, viral RNA was detected in samples collected from shed surfaces and air. Consequently, strict biosecurity is imperative during the occurrence of clinical signs. Environmental viral load monitoring, in conjunction with NGS should be adopted in mink farm surveillance. The minimum proportion of minks that need to be immunized to avoid outbreaks in farms was calculated at 65.5%, which is important for future vaccination campaigns.     

A Solanum tuberosum inositol phosphate kinase (StITPK1) displaying inositol phosphate-inositol phosphate and inositol phosphate-ADP phosphotransferase activities

We describe a multifunctional inositol polyphosphate kinase/phosphotransferase from Solanum tuberosum, StITPKalpha (GenBank accession number: EF362784), hereafter called StITPK1. StITPK1 displays inositol 3,4,5,6-tetrakisphosphate 1-kinase activity: K(m) = 27 microM, and V(max) = 19 nmol min(-1) mg(-1). The enzyme displays inositol 1,3,4,5,6-pentakisphosphate 1-phosphatase activity in the absence of a nucleotide acceptor and inositol 1,3,4,5,6-pentakisphosphate-ADP phosphotransferase activity in the presence of physiological concentrations of ADP. Additionally, StITPK1 shows inositol phosphate-inositol phosphate phosphotransferase activity. Homology modelling provides a structural rationale of the catalytic abilities of StITPK1. Inter-substrate transfer of phosphate groups between inositol phosphates is an evolutionarily conserved function of enzymes of this class.     

Allergy and risk of acute lymphoblastic leukemia among children: A nationwide case control study in Greece

Background: Several reports point to inverse associations between allergies and ALL; yet, no study has explored this link using both self-reported-data on allergic history and biomarkers of atopic sensitization. Methods: Clinical information for the variables of interest was available for 252 out of 292 cases of childhood (0–14 years) ALL, newly diagnosed across Greece over a 4.5 year period as well as for 294 hospital controls. Allergen-specific-IgEs, as markers of allergic predisposition, against 24 most prevalent respiratory and food allergens, were determined, using an enzyme immunoassay procedure for 199 children with ALL and 113 controls. Cases were compared with controls through frequency distributions and unconditional multiple logistic regression models to estimate odds ratios (ORs) and 95% confidence-intervals (CIs) regarding associations of allergy with childhood ALL. Results: Self-reported-allergic history overall (OR: 0.49, 95%CI: 0.34–0.72) and practically each one of its main components (respiratory, food, any other clinical allergy) were strongly and inversely associated with ALL. Likewise, the serum IgE inverse association was of the same magnitude (OR: 0.43, 95%CI: 0.22–0.84) mainly contributed by food IgE (OR: 0.39, 95%CI: 0.18–0.83). Conclusion: Beyond the already established inverse association of allergic history with childhood ALL, a same magnitude association is evident when serologic markers of allergic predisposition are used as an alternative measure of allergy. Further research with more appropriate study designs is needed to better understand possible associations between prior allergy and childhood ALL risk.     

Evidence for the early emergence of piperaquine-resistant Plasmodium falciparum malaria and modeling strategies to mitigate resistance

Multidrug-resistant Plasmodium falciparum parasites have emerged in Cambodia and neighboring countries in Southeast Asia, compromising the efficacy of first-line antimalarial combinations. Dihydroartemisinin + piperaquine (PPQ) treatment failure rates have risen to as high as 50% in some areas in this region. For PPQ, resistance is driven primarily by a series of mutant alleles of the P. falciparum chloroquine resistance transporter (PfCRT). PPQ resistance was reported in China three decades earlier, but the molecular driver remained unknown. Herein, we identify a PPQ-resistant pfcrt allele (China C) from Yunnan Province, China, whose genotypic lineage is distinct from the PPQ-resistant pfcrt alleles currently observed in Cambodia. Combining gene editing and competitive growth assays, we report that PfCRT China C confers moderate PPQ resistance while re-sensitizing parasites to chloroquine (CQ) and incurring a fitness cost that manifests as a reduced rate of parasite growth. PPQ transport assays using purified PfCRT isoforms, combined with molecular dynamics simulations, highlight differences in drug transport kinetics and in this transporter’s central cavity conformation between China C and the current Southeast Asian PPQ-resistant isoforms. We also report a novel computational model that incorporates empirically determined fitness landscapes at varying drug concentrations, combined with antimalarial susceptibility profiles, mutation rates, and drug pharmacokinetics. Our simulations with PPQ-resistant or -sensitive parasite lines predict that a three-day regimen of PPQ combined with CQ can effectively clear infections and prevent the evolution of PfCRT variants. This work suggests that including CQ in combination therapies could be effective in suppressing the evolution of PfCRT-mediated multidrug resistance in regions where PPQ has lost efficacy.     

Fluorescence characterisation of multiply-loaded anti-HER2 single chain Fv-photosensitizer conjugates suitable for photodynamic therapy.

We report the synthesis, spectroscopic properties and intracellular imaging of recombinant antibody single chain fragment (scFv) conjugates with photosensitizers used for photodynamic therapy of cancer (PDT). Two widely-studied photosensitizers have been selected: preclinical pyropheophorbide-a (PPa) and verteporfin (VP), which has been clinically approved for the treatment of acute macular degeneration (Visudyne). Pyropheophorbide-a and verteporfin have been conjugated to an anti-HER2 scFv containing on average ten photosensitizer molecules per scFv with a small contribution (相似文献