Effect of Flagella on Initial Attachment of Listeria monocytogenes to Stainless Steel

At 22°C a flagellin mutant of Listeria monocytogenes was found to attach to stainless steel at levels 10-fold lower than wild-type cells, even under conditions preventing active motility. At 37°C, when flagella are not produced, attachment of both strains was identical. Therefore, flagella per se facilitate the early stage of attachment.     

Programmed cell death 4: A novel player in the pathogenesis of polycystic ovary syndrome

Polycystic ovary syndrome (PCOS) is a pathological condition recognized by menstrual cycle irregularities, androgen excess, and polycystic ovarian morphology, affecting a significant proportion of women of childbearing age and accounting for the most prevalent cause of anovulatory sterility. In addition, PCOS is frequently accompanied by metabolic and endocrine disturbances such as obesity, dyslipidemia, insulin resistance, and hyperinsulinemia, indicating the multiplicity of mechanisms implicated in the progression of PCOS. However, the exact pathogenesis of PCOS is yet to be elucidated. Programmed cell death 4 (PDCD4) is a ubiquitously expressed protein that contributes to the regulation of various cellular processes, including gene expression, cell cycle progression, proliferation, and apoptosis. Despite some disparities concerning its exact cellular effects, PDCD4 is generally characterized as a protein that inhibits cell cycle progression and proliferation and instead drives the cell into apoptosis. The apoptosis of granulosa cells (GCs) is speculated to take a major part in the occurrence and progression of PCOS by ceasing antral follicle development and compromising oocyte competence. Given the possible involvement of GC apoptosis in the progression of PCOS, as well as the contribution of PDCD4 to the regulation of cell apoptosis and the development of metabolic diseases, the current review aimed to discuss whether or how PDCD4 can play a role in the pathogenesis of PCOS by affecting GC apoptosis.     

Genetic determinants of phenotypic diversity in humans

New technologies for rapidly assaying DNA sequences have revealed that the degree and nature of human genetic variation is far more complex then previously realized. These same technologies have also resulted in the identification of common genetic variants associated with more than 30 human diseases and traits.     

Neural Representation of Ambiguous Visual Objects in the Inferior Temporal Cortex

Inferior temporal (IT) cortex as the final stage of the ventral visual pathway is involved in visual object recognition. In our everyday life we need to recognize visual objects that are degraded by noise. Psychophysical studies have shown that the accuracy and speed of the object recognition decreases as the amount of visual noise increases. However, the neural representation of ambiguous visual objects and the underlying neural mechanisms of such changes in the behavior are not known. Here, by recording the neuronal spiking activity of macaque monkeys’ IT we explored the relationship between stimulus ambiguity and the IT neural activity. We found smaller amplitude, later onset, earlier offset and shorter duration of the response as visual ambiguity increased. All of these modulations were gradual and correlated with the level of stimulus ambiguity. We found that while category selectivity of IT neurons decreased with noise, it was preserved for a large extent of visual ambiguity. This noise tolerance for category selectivity in IT was lost at 60% noise level. Interestingly, while the response of the IT neurons to visual stimuli at 60% noise level was significantly larger than their baseline activity and full (100%) noise, it was not category selective anymore. The latter finding shows a neural representation that signals the presence of visual stimulus without signaling what it is. In general these findings, in the context of a drift diffusion model, explain the neural mechanisms of perceptual accuracy and speed changes in the process of recognizing ambiguous objects.     

A conditional yeast E1 mutant blocks the ubiquitin-proteasome pathway and reveals a role for ubiquitin conjugates in targeting Rad23 to the proteasome

E1 ubiquitin activating enzyme catalyzes the initial step in all ubiquitin-dependent processes. We report the isolation of uba1-204, a temperature-sensitive allele of the essential Saccharomyces cerevisiae E1 gene, UBA1. Uba1-204 cells exhibit dramatic inhibition of the ubiquitin-proteasome system, resulting in rapid depletion of cellular ubiquitin conjugates and stabilization of multiple substrates. We have employed the tight phenotype of this mutant to investigate the role ubiquitin conjugates play in the dynamic interaction of the UbL/UBA adaptor proteins Rad23 and Dsk2 with the proteasome. Although proteasomes purified from mutant cells are intact and proteolytically active, they are depleted of ubiquitin conjugates, Rad23, and Dsk2. Binding of Rad23 to these proteasomes in vitro is enhanced by addition of either free or substrate-linked ubiquitin chains. Moreover, association of Rad23 with proteasomes in mutant and wild-type cells is improved upon stabilizing ubiquitin conjugates with proteasome inhibitor. We propose that recognition of polyubiquitin chains by Rad23 promotes its shuttling to the proteasome in vivo.     

Allosteric modulation of the human P-glycoprotein involves conformational changes mimicking catalytic transition intermediates

The drug transport function of human P-glycoprotein (Pgp, ABCB1) can be inhibited by a number of pharmacological agents collectively referred to as modulators or reversing agents. In this study, we demonstrate that certain thioxanthene-based Pgp modulators with an allosteric mode of action induce a distinct conformational change in the cytosolic domain of Pgp, which alters susceptibility to proteolytic digestion. Both cis and trans-isomers of the Pgp modulator flupentixol confer considerable protection of an 80 kDa Pgp fragment against trypsin digestion, that is recognized by a polyclonal antibody specific for the NH(2)-terminal half to Pgp. The protection by flupentixol is abolished in the Pgp F983A mutant that is impaired in modulation by flupentixols, indicating involvement of the allosteric site in generating the conformational change. A similar protection to an 80 kDa fragment is conferred by ATP, its nonhydrolyzable analog ATPgammaS, and by trapping of ADP-vanadate at the catalytic domain, but not by transport substrate vinblastine or by the competitive modulator cyclosporin A, suggesting different outcomes from modulator interaction at the allosteric site and at the substrate site. In summary, we demonstrate that allosteric interaction of flupentixols with Pgp generates conformational changes that mimic catalytic transition intermediates induced by nucleotide binding and hydrolysis, which may play a crucial role in allosteric inhibition of Pgp-mediated drug transport.     

Clustered epitopes within a new poly-epitopic HIV-1 DNA vaccine shows immunogenicity in BALB/c mice

Despite a huge number of studies towards vaccine development against human immunodeficiency virus-1, no effective vaccine has been approved yet. Thus, new vaccines should be provided with new formulations. Herein, a new DNA vaccine candidate encoding conserved and immunogenic epitopes from HIV-1 antigens of tat, pol, gag and env is designed and constructed. After bioinformatics analyses to find the best epitopes and their tandem, nucleotide sequence corresponding to the designed multiepitope was synthesized and cloned into pcDNA3.1+ vector. Expression of pcDNA3.1-tat/pol/gag/env plasmid was evaluated in HEK293T cells by RT-PCR and western-blotting. Seven groups of BALB/c mice were intramuscularly immunized three times either with 50, 100, 200 µg of plasmid in 2-week intervals or with similar doses of insert-free plasmid. Two weeks after the last injection, proliferation of T cells and secretion of IL4 and IFN-γ cytokines were evaluated using Brdu and ELISA methods, respectively. Results showed the proper expression of the plasmid in protein and mRNA levels. Moreover, the designed multiepitope plasmid was capable of induction of both proliferation responses as well as IFN-γ and IL-4 cytokine production in a considerable level compared to the control groups. Overall, our primary data warranted further detailed studies on the potency of this vaccine.     

Bile acid-independent protection against Clostridioides difficile infection

Clostridioides difficile infections occur upon ecological / metabolic disruptions to the normal colonic microbiota, commonly due to broad-spectrum antibiotic use. Metabolism of bile acids through a 7α-dehydroxylation pathway found in select members of the healthy microbiota is regarded to be the protective mechanism by which C. difficile is excluded. These 7α-dehydroxylated secondary bile acids are highly toxic to C. difficile vegetative growth, and antibiotic treatment abolishes the bacteria that perform this metabolism. However, the data that supports the hypothesis that secondary bile acids protect against C. difficile infection is supported only by in vitro data and correlative studies. Here we show that bacteria that 7α-dehydroxylate primary bile acids protect against C. difficile infection in a bile acid-independent manner. We monoassociated germ-free, wildtype or Cyp8b1^-/- (cholic acid-deficient) mutant mice and infected them with C. difficile spores. We show that 7α-dehydroxylation (i.e., secondary bile acid generation) is dispensable for protection against C. difficile infection and provide evidence that Stickland metabolism by these organisms consumes nutrients essential for C. difficile growth. Our findings indicate secondary bile acid production by the microbiome is a useful biomarker for a C. difficile-resistant environment but the microbiome protects against C. difficile infection in bile acid-independent mechanisms.