CYP52X1, representing new cytochrome P450 subfamily, displays fatty acid hydroxylase activity and contributes to virulence and growth on insect cuticular substrates in entomopathogenic fungus Beauveria bassiana

Infection of insects by the entomopathogenic fungus Beauveria bassiana proceeds via attachment and penetration of the host cuticle. The outermost epicuticular layer or waxy layer of the insect represents a structure rich in lipids including abundant amounts of hydrocarbons and fatty acids. A member of a novel cytochrome P450 subfamily, CYP52X1, implicated in fatty acid assimilation by B. bassiana was characterized. B. bassiana targeted gene knockouts lacking Bbcyp52x1 displayed reduced virulence when topically applied to Galleria mellonella, but no reduction in virulence was noted when the insect cuticle was bypassed using an intrahemoceol injection assay. No significant growth defects were noted in the mutant as compared with the wild-type parent on any lipids substrates tested including alkanes and fatty acids. Insect epicuticle germination assays, however, showed reduced germination of ΔBbcyp52x1 conidia on grasshopper wings as compared with the wild-type parent. Complementation of the gene-knock with the full-length gene restored virulence and insect epicuticle germination to wild-type levels. Heterologous expression of CYP52X1 in yeast was used to characterize the substrate specificity of the enzyme. CYP52X1 displayed the highest activity against midrange fatty acids (C12:0 and C14:0) and epoxy stearic acid, 4–8-fold lower activity against C16:0, C18:1, and C18:2, and little to no activity against C9:0 and C18:0. Analyses of the products of the C12:0 and C18:1 reactions confirmed NADPH-dependent regioselective addition of a terminal hydroxyl to the substrates (ω-hydroxylase). These data implicate CYP52X1 as contributing to the penetration of the host cuticle via facilitating the assimilation of insect epicuticle lipids.Fil: Zhang, Shizhu. Nanjing Normal University; China. University of Florida; Estados UnidosFil: Widemann, Emilie. Université de Strasbourg; FranciaFil: Bernard, Grausem. Université de Strasbourg; FranciaFil: Lesot, Agnes. Université de Strasbourg; FranciaFil: Pinot, Franck. Université de Strasbourg; FranciaFil: Pedrini, Nicolás. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Bioquímicas de La Plata "Prof. Dr. Rodolfo R. Brenner". Universidad Nacional de la Plata. Facultad de Ciencias Médicas. Instituto de Investigaciones Bioquímicas de La Plata "Prof. Dr. Rodolfo R. Brenner"; ArgentinaFil: Keyhani, Nemat O.. University of Florida; Estados Unido

Cephalic Tetanus from Penetrating Orbital Wound

Tetanus is a neurologic disorder caused by tetanospasmin, a protein toxin elaborated by Clostridium tetani. Cephalic tetanus is a localized form of the disease causing trismus and dysfunction of cranial nerves. We report the case of a man who presented with facial trauma, complete ophthalmoplegia, exophthalmos, areactive mydriasis, and periorbital hematoma. An orbital CT revealed air bubbles in the right orbital apex. The patient was given a tetanus toxoid booster and antibiotherapy. After extraction of a wooden foreign body, the patient developed right facial nerve palsy, disorders of swallowing, contralateral III cranial nerve palsy, and trismus. Only one case of cephalic tetanus from penetrating orbital wound has been reported in literature 20 years ago. When a patient presents with an orbital wound with ophthalmoplegia and signs of anaerobic infection, cephalic tetanus should be ruled out

Aurora A contributes to p150ᵍⁱᵘᵉᵈ phosphorylation and function during mitosis

Aurora A is a spindle pole–associated protein kinase required for mitotic spindle assembly and chromosome segregation. In this study, we show that Drosophila melanogaster aurora A phosphorylates the dynactin subunit p150ᵍⁱᵘᵉᵈ on sites required for its association with the mitotic spindle. Dynactin strongly accumulates on microtubules during prophase but disappears as soon as the nuclear envelope breaks down, suggesting that its spindle localization is tightly regulated. If aurora A's function is compromised, dynactin and dynein become enriched on mitotic spindle microtubules. Phosphorylation sites are localized within the conserved microtubule-binding domain (MBD) of the p150ᵍⁱᵘᵉᵈ. Although wild-type p150ᵍⁱᵘᵉᵈ binds weakly to spindle microtubules, a variant that can no longer be phosphorylated by aurora A remains associated with spindle microtubules and fails to rescue depletion of endogenous p150ᵍⁱᵘᵉᵈ. Our results suggest that aurora A kinase participates in vivo to the phosphoregulation of the p150ᵍⁱᵘᵉᵈ MBD to limit the microtubule binding of the dynein–dynactin complex and thus regulates spindle assembly

Effects of Age and Task Load on Drivers’ Response Accuracy and Reaction Time When Responding to Traffic Lights

International audienceDue to population aging, elderly drivers represent an increasing proportion of car drivers. Yet, how aging alters sensorimotor functions and impacts driving safety remains poorly understood. This paper aimed at assessing to which extent elderly drivers are sensitive to various task loads and how this affects the reaction time (RT) in a driving context. Old and middle-aged people completed RT tasks which reproduced cognitive demands encountered while driving. Participants had to detect and respond to traffic lights or traffic light arrows as quickly as possible, under three experimental conditions of incremental difficulty. In both groups, we hypothesized that decision-making would be impacted by the number of cues to be processed. The first test was a simple measure of RT. The second and third tests were choice RT tasks requiring the processing of 3 and 5 cues, respectively. Responses were collected within a 2 s time-window. Otherwise, the trial was considered a no-response. In both groups, the data revealed that RT, error rate (incorrect answers), and no-response rate increased along with task difficulty. However, the middle-aged group outperformed the elderly group. The RT difference between the two groups increased drastically along with task difficulty. In the third test, the rate of no-response suggested that elderly drivers needed more than 2 s to process complex information and respond accurately. Both prolonged RT and increased no-response rate, especially for difficult tasks, might attest an impairment of cognitive abilities in relation to aging. Accordingly, casual driving conditions for young drivers may be particularly complex and stressful for elderly people who should thus be informed about the effects of normal aging upon driving

Aurora A contributes to p150ᵍⁱᵘᵉᵈ phosphorylation and function during mitosis

Aurora A is a spindle pole–associated protein kinase required for mitotic spindle assembly and chromosome segregation. In this study, we show that Drosophila melanogaster aurora A phosphorylates the dynactin subunit p150ᵍⁱᵘᵉᵈ on sites required for its association with the mitotic spindle. Dynactin strongly accumulates on microtubules during prophase but disappears as soon as the nuclear envelope breaks down, suggesting that its spindle localization is tightly regulated. If aurora A's function is compromised, dynactin and dynein become enriched on mitotic spindle microtubules. Phosphorylation sites are localized within the conserved microtubule-binding domain (MBD) of the p150ᵍⁱᵘᵉᵈ. Although wild-type p150ᵍⁱᵘᵉᵈ binds weakly to spindle microtubules, a variant that can no longer be phosphorylated by aurora A remains associated with spindle microtubules and fails to rescue depletion of endogenous p150ᵍⁱᵘᵉᵈ. Our results suggest that aurora A kinase participates in vivo to the phosphoregulation of the p150ᵍⁱᵘᵉᵈ MBD to limit the microtubule binding of the dynein–dynactin complex and thus regulates spindle assembly

Approche variationnelle pour la déconvolution rapide de données 3D en microscopie biphotonique

National audienceBetter understanding of biological processes requires new, improved, high resolution imagery techniques. The present work concerns the restoration of data acquired with two-photon microscopy in biological tissue, in-vivo in particular. Two main challenges to tackle are: the large dimensionality of the acquired data and the incomplete knowledge of the impulse response of the system. We propose here an experimental setting to estimate it based on the observation of fluorescent micro-beads. The non-blind formulation of the related inverse problem of image restoration is then solved by minimizing a penalized criterion using an efficient convex optimization strategy based on the Majoration-Minimization approach. The effectiveness of the proposed approach is not only shown on simulated data but also on real data.Les technologies d'imagerie permettant d'obtenir des images à l'échelle cellulaire sont devenues incontournables pour mieux comprendre les processus biologiques. Nous nous intéressons ici à l'imagerie biphotonique et plus particulièrement, à la reconstruction des images acquises par un tel système dans un contexte in-vivo. Deux difficultés majeures sont : les gros volumes de données que représentent les acquisitions et la mauvaise connaissance de la réponse impulsionnelle de l'instrument. Nous proposons dans ce travail d'estimer cette dernière à l'aide d'un montage expérimental basé sur l'observation de micro-billes fluorescentes. Le problème inverse de restauration d'image est ensuite résolu dans un cadre non aveugle, en le formulant sous la forme d'un critère pénalisé minimisé à l'aide d'une méthode d'optimisation convexe efficace fondée sur la technique de Majoration-Minimisation. L'efficacité de l'approche proposée est démontrée sur données simulées et réelles

CDK11p58 Is Required for Centriole Duplication and Plk4 Recruitment to Mitotic Centrosomes

BACKGROUND: CDK11(p58) is a mitotic protein kinase, which has been shown to be required for different mitotic events such as centrosome maturation, chromatid cohesion and cytokinesis. METHODOLOGY/PRINCIPAL FINDINGS: In addition to these previously described roles, our study shows that CDK11(p58) inhibition induces a failure in the centriole duplication process in different human cell lines. We propose that this effect is mediated by the defective centrosomal recruitment of proteins at the onset of mitosis. Indeed, Plk4 protein kinase and the centrosomal protein Cep192, which are key components of the centriole duplication machinery, showed reduced levels at centrosomes of mitotic CDK11-depleted cells. CDK11(p58), which accumulates only in the vicinity of mitotic centrosomes, directly interacts with the centriole-associated protein kinase Plk4 that regulates centriole number in cells. In addition, we show that centriole from CDK11 defective cells are not able to be over duplicated following Plk4 overexpression. CONCLUSION/SIGNIFICANCE: We thus propose that CDK11 is required for centriole duplication by two non-mutually-exclusive mechanisms. On one hand, the observed duplication defect could be caused indirectly by a failure of the centrosome to fully maturate during mitosis. On the other hand, CDK11(p58) could also directly regulate key centriole components such as Plk4 during mitosis to trigger essential mitotic centriole modifications, required for centriole duplication during subsequent interphase

Label-Free Quantification of Nanoencapsulated Piperonyl Esters in Cosmetic Hydrogels Using Raman Spectroscopy

Raman spectroscopy is a well-established technique for the molecular characterisation of samples and does not require extensive pre-analytical processing for complex cosmetic products. As an illustration of its potential, this study investigates the quantitative performance of Raman spectroscopy coupled with partial least squares regression (PLSR) for the analysis of Alginate nanoencapsulated Piperonyl Esters (ANC-PE) incorporated into a hydrogel. A total of 96 ANC-PE samples covering a 0.4% w/w–8.3% w/w PE concentration range have been prepared and analysed. Despite the complex formulation of the sample, the spectral features of the PE can be detected and used to quantify the concentrations. Using a leave-K-out cross-validation approach, samples were divided into a training set (n = 64) and a test set, samples that were previously unknown to the PLSR model (n = 32). The root mean square error of cross-validation (RMSECV) and prediction (RMSEP) was evaluated to be 0.142% (w/w PE) and 0.148% (w/w PE), respectively. The accuracy of the prediction model was further evaluated by the percent relative error calculated from the predicted concentration compared to the true value, yielding values of 3.58% for the training set and 3.67% for the test set. The outcome of the analysis demonstrated the analytical power of Raman to obtain label-free, non-destructive quantification of the active cosmetic ingredient, presently PE, in complex formulations, holding promise for future analytical quality control (AQC) applications in the cosmetics industry with rapid and consumable-free analysis