Molecular Probing of the HPV-16 E6 Protein Alpha Helix Binding Groove with Small Molecule Inhibitors

The human papillomavirus (HPV) HPV E6 protein has emerged as a central oncoprotein in HPV-associated cancers in which sustained expression is required for tumor progression. A majority of the E6 protein interactions within the human proteome use an alpha-helix groove interface for binding. The UBE3A/E6AP HECT domain ubiquitin ligase binds E6 at this helix-groove interface. This enables formation of a trimeric complex with p53, resulting in destruction of this tumor suppressor. While recent x-ray crystal structures are useful, examples of small molecule probes that can modulate protein interactions at this interface are limited. To develop insights useful for potential structure-based design of ligands for HPV E6, a series of 2,6-disubstituted benzopyranones were prepared and tested as competitive antagonists of E6-E6AP helix-groove interactions. These small molecule probes were used in both binding and functional assays to evaluate recognition features of the E6 protein. Evidence for an ionic functional group interaction within the helix groove was implicated by the structure-activity among the highest affinity ligands. The molecular topographies of these protein-ligand interactions were evaluated by comparing the binding and activities of single amino acid E6 mutants with the results of molecular dynamic simulations. A group of arginine residues that form a rim-cap over the E6 helix groove offer compensatory roles in binding and recognition of the small molecule probes. The flexibility and impact on the overall helix-groove shape dictated by these residues offer new insights for structure-based targeting of HPV E6.     

The Power of the Picture: How Narrative Film Captures Attention and Disrupts Goal Pursuit

Narrative transportation is described as a state of detachment that arises when one becomes immersed in the narrative of a story. Participants viewed either an intact version of an engaging 20 min film, “Bang You’re Dead!,” (1961) by Alfred Hitchcock (contiguous condition), or a version of the same film with scenes presented out of order (noncontiguous condition). In this latter condition, the individual scenes were intact but were presented out of chronological order. Participants were told a cover story that we were interested in the amount of gun violence depicted in films. Both groups were given the goal to remember to lift their hand every time they heard the word “gun” spoken during the film. Results revealed that participants were significantly less likely to remember to execute their goal in the contiguous condition, presumably because this narrative transported viewers’ attention and thereby “hijacked” processing resources away from internal goals.     

Dasatinib Inhibits CXCR4 Signaling in Chronic Lymphocytic Leukaemia Cells and Impairs Migration Towards CXCL12

Chemokines and their ligands play a critical role in enabling chronic lymphocytic leukaemia (CLL) cells access to protective microenvironmental niches within tissues, ultimately resulting in chemoresistance and relapse: disruption of these signaling pathways has become a novel therapeutic approach in CLL. The tyrosine kinase inhibitor dasatinib inhibits migration of several cell lines from solid-organ tumours, but effects on CLL cells have not been reported. We studied the effect of clinically achievable concentrations of dasatinib on signaling induced by the chemokine CXCL12 through its'' receptor CXCR4, which is highly expressed on CLL cells. Dasatinib pre-treatment inhibited Akt and ERK phosphorylation in CLL cells upon stimulation with CXCL12. Dasatinib also significantly diminished the rapid increase in actin polymerisation observed in CLL cells following CXCL12 stimulation. Moreover, the drug significantly inhibited chemotaxis in a transwell assay, and reduced the percentage of cells able to migrate beneath a CXCL12-expressing murine stromal cell line. Dasatinib also abrogated the anti-apoptotic effect of prolonged CXCL12 stimulation on cultured CLL cells. These data suggest that dasatinib, akin to other small molecule kinase inhibitors targeting the B-cell receptor signaling pathway, may redistribute CLL cells from protective tissue niches to the peripheral blood, and support the investigation of dasatinib in combination strategies.     

Intermittent illumination increases biophotolytic hydrogen yield by Anabaena cylindrica.

Intermittent illumination increased H2 and C2H4 yields per unit of light from growing cells and from nitrogren-starved cells by 1.7- and 1.35-fold, respectively, as compared with continuous illumination.     

Alteration of the Pathogenicity of Pasteurella pneumotropica for the Murine Lung Caused by Changes in Pulmonary Antibacterial Activity

Pasteurella pneumotropica is a potential pulmonary pathogen in mice. In healthy animals, this organism was killed rapidly by the normal function of the intrapulmonary phagocytic defense mechanisms. Impairment of this bactericidal activity by the acute renal failure of nephrectomy resulted in multiplication of the Pasteurella in the lung, both when the animals were nephrectomized first and then infected, and when the animals were infected first and nephrectomized several hours after the infection. The study demonstrates that the pathogenicity of the Pasteurella organisms is governed by the functional state of these pulmonary antibacterial mechanisms.     

Gene regulation during dedifferentiation in Dictyostelium discoideum

During development of Dictyostelium discoideum, cells acquire the capacity to rapidly recapitulate morphogenesis. Therefore, when cells at the loose aggregate stage are disaggregated and challenged to reaggregate, they do so in a tenth of the original time. If loose aggregate cells are disaggregated and resuspended in buffered dextrose solution (erasure medium), they retain the capacity of rapid recapitulation for 80 min, then completely lose this capacity in a single, synchronous step referred to as the "erasure event." The erasure event sets in motion a program of dedifferentiation during which cells lose developmentally acquired characteristics at different times. The erasure event is inhibited by the addition of 10(-4) M cAMP to erasure medium. The synthesis of 33 growth-associated polypeptides, the synthesis of 53 development-associated polypeptides, and the level of 2 development-associated RNAs have been monitored during the erasure program and in cultures inhibited from erasing by the addition of 10(-4) M cAMP. Growth-associated polypeptides begin to be resynthesized and development-associated polypeptides exhibit dramatic decreases in rate of synthesis at different times throughout the first 240 min in erasure medium. Inhibiting the erasure event with cAMP has no major effect in the resynthesis of the majority of growth-associated polypeptides. Only one growth-associated polypeptide, V28, is completely inhibited by cAMP, suggesting that it may play a role in the erasure process. In contrast, inhibiting the erasure event with cAMP has a marked effect on the synthesis of development-associated polypeptides, causing a dramatic reduction in the rate at which synthesis decreases for 6 polypeptides, and completely inhibits the decrease in the synthetic rate of 8 polypeptides. The two development-associated RNAs, 16G1 and 10C3, exhibit two distinctly different patterns of loss during erasure, but in both cases cAMP added at time zero of the erasure process dramatically retards or inhibits loss. In addition, when cAMP is added just prior to the erasure event, it inhibits the erasure event and stimulates a rapid increase in the level of 16G1 RNA back to the developmental level. The level of 16G1 RNA then remains at this level for at least 400 min. When cAMP is added after the erasure event, it causes a low, transient increase in the level of 16G1 RNA. These results are considered both in relation to the program of erasure, and in relation to the role of cAMP in the expression of developmental genes during the forward program of development.