A new gene involved in coenzyme Q biosynthesis in Escherichia coli: UbiI functions in aerobic C5-hydroxylation

International audienceCoenzyme Q (ubiquinone or Q) is a redox-active lipid found in organisms ranging from bacteria to mammals in which it plays a crucial role in energy-generating processes. Q biosynthesis is a complex pathway that involves multiple proteins. In this work, we show that the uncharacterized conserved visC gene is involved in Q biosynthesis in Escherichia coli, and we have renamed it ubiI. Based on genetic and biochemical experiments, we establish that the UbiI protein functions in the C5-hydroxylation reaction. A strain deficient in ubiI has a low level of Q and accumulates a compound derived from the Q biosynthetic pathway, which we purified and characterized. We also demonstrate that UbiI is only implicated in aerobic Q biosynthesis and that an alternative enzyme catalyzes the C5-hydroxylation reaction in the absence of oxygen. We have solved the crystal structure of a truncated form of UbiI. This structure shares many features with the canonical FAD-dependent para-hydroxybenzoate hydroxylase and represents the first structural characterization of a monooxygenase involved in Q biosynthesis. Site-directed mutagenesis confirms that residues of the flavin binding pocket of UbiI are important for activity. With our identification of UbiI, the three monooxygenases necessary for aerobic Q biosynthesis in E. coli are known

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival

Les personnalités publiques : entre droit à la vie privée et liberté d'expression

Master [120] en droit, Université catholique de Louvain, 200

La capture de l'attention par des expressions faciales schématiques de colère est-elle la conséquence d'un traitement holistique ou de bas niveau ?

Nous avons étudié l'effet de capture attentionnelle de visages schématiques menaçants. Pour cela, six visages sont présentés autour d'un point de fixation mais un seul possède un nez orienté différemment. Les participants doivent rapporter son orientation. Un des visages neutres peut être remplacé aléatoirement par un visage distracteur émotionnel de colère. Ces derniers peuvent être présentés à l'endroit ou à l'envers, nous permettant ainsi de tester l'impact de bas niveau des stimuli. L'effet de capture a été étudié grâce à la composante N2pc, reflétant le traitement attentionnel en direction d'un stimulus latéralisé, ainsi que des temps de réaction. Nous prédisons une capture de l'attention en direction des visages de colère, effet qui serait aboli par l'inversion du visage. Nos résultats électrophysiologiques indiquent une capture par le visage cible uniquement. Grâce à une analyse post-hoc, nous observons un effet d'habituation dans la condition à l'endroit

Approche électrophysiologique de la suppression des distracteurs

Nos capacités attentionnelles étant limitées, il est important de pouvoir sélectionner l'information pertinente. Nous avons analysé différentes situations dans lesquelles un distracteur visuel saillant est présenté alors qu'il est non-pertinent. Nous avons donc mené une série d'expériences sur la base de deux paradigmes: le paradigme d'indiçage spatial modifié ; l'additional singleton paradigm. Afin d'aller plus loin dans la compréhension des résultats, nous avons mené ces expériences en électrophysiologie pour comprendre les mécanismes de traitement visuel impliqués dans la suppression du distracteur. Nous nous sommes focalisés sur deux composantes, la N2pc reflétant une capture attentionnelle, et la PD reflétant une suppression du distracteur. Nos résultats montrent que les distracteurs saillants sont supprimés lorsque la tâche est facile. De plus, suite à une phase d'entrainement, une PD est observée en direction du distracteur dans une recherche d'élément unique. À l'inverse, nous sommes incapables d'ignorer un distracteur saillant dans une recherche difficile

Attentional suppression is delayed for threatening distractors

According to the threat-capture hypothesis, fear-related stimuli have a high attentional priority. As a result, irrelevant-but-salient stimuli interfere more with a visual search task when they are perceived as threatening. We investigated the neural basis for behavioral interference in conditions that promote attentional suppression of distracting stimuli (i.e., easy search with fixed target/distractor roles). In Experiment 1, participants discriminated the shape of a neutral target (a flower), which competed for selection with a threat-related (spider) or neutral (leaf) distractor. In line with prior results, we observed larger interference from spider than leaf distractors. At an electrophysiological level, we found that participants actively suppressed both distractors as evidenced by the presence of a posterior positivity between 200-300 ms, the PD. Critically, the PD was delayed with spider compared to leaf distractors. Further, in the spider distractor condition, the offset of the PD component correlated with response time to complete the search task when the spider was present. Experiment 2 was a control experiment where we confirmed that the results depended on the execution of the peripheral search task. When participants performed a localization task on the fixation cross, the decisive results from Experiment 1 were not replicated despite equal peripheral stimulation. Our results indicate that the behavioral delay incurred by threatening stimuli is accompanied by a delay of suppressive mechanisms. In contrast, we found no evidence for initial capture followed by suppression that may be predicted by hypervigilance-avoidance theory

Attentional suppression is delayed for threatening distractors

According to the threat-capture hypothesis, fear-related stimuli have a high attentional priority. As a result, irrelevant-but-salient stimuli interfere more with a visual search task when they are perceived as threatening. We investigated the neural basis for behavioral interference in conditions that promote attentional suppression of distracting stimuli (i.e., easy search with fixed target/distractor roles). In Experiment 1, participants discriminated the shape of a neutral target (a flower), which competed for selection with a threat-related (spider) or neutral (leaf) distractor. In line with prior results, we observed larger interference from spider than leaf distractors. At an electrophysiological level, we found that participants actively suppressed both distractors as evidenced by the presence of a posterior positivity between 200-300 ms, the PD. Critically, the PD was delayed with spider compared to leaf distractors. Further, in the spider distractor condition, the offset of the PD component correlated with response time to complete the search task when the spider was present. Experiment 2 was a control experiment where we confirmed that the results depended on the execution of the peripheral search task. When participants performed a localization task on the fixation cross, the decisive results from Experiment 1 were not replicated despite equal peripheral stimulation. Our results indicate that the behavioral delay incurred by threatening stimuli is accompanied by a delay of suppressive mechanisms. In contrast, we found no evidence for initial capture followed by suppression that may be predicted by hypervigilance-avoidance theory

Attentional suppression is delayed for threatening distractors

According to the threat-capture hypothesis, fear-related stimuli have a high attentional priority. As a result, irrelevant-but-salient stimuli interfere more with a visual search task when they are perceived as threatening. We investigated the neural basis for behavioral interference in conditions that promote attentional suppression of distracting stimuli (i.e., easy search with fixed target/distractor roles). In Experiment 1, participants discriminated the shape of a neutral target (a flower), which competed for selection with a threat-related (spider) or neutral (leaf) distractor. In line with prior results, we observed larger interference from spider than leaf distractors. At an electrophysiological level, we found that participants actively suppressed both distractors as evidenced by the presence of a posterior positivity between 200-300 ms, the PD. Critically, the PD was delayed with spider compared to leaf distractors. Further, in the spider distractor condition, the offset of the PD component correlated with response time to complete the search task when the spider was present. Experiment 2 was a control experiment where we confirmed that the results depended on the execution of the peripheral search task. When participants performed a localization task on the fixation cross, the decisive results from Experiment 1 were not replicated despite equal peripheral stimulation. Our results indicate that the behavioral delay incurred by threatening stimuli is accompanied by a delay of suppressive mechanisms. In contrast, we found no evidence for initial capture followed by suppression that may be predicted by hypervigilance-avoidance theory