The Pkn22 Ser/Thr kinase in Nostoc PCC 7120: role of FurA and NtcA regulators and transcript profiling under nitrogen starvation and oxidative stress

International audienceBackground: The filamentous cyanobacterium Nostoc sp. strain PCC 7120 can fix N2 when combined nitrogen is not available. Furthermore, it has to cope with reactive oxygen species generated as byproducts of photosynthesis and respiration. We have previously demonstrated the synthesis of Ser/Thr kinase Pkn22 as an important survival response of Nostoc to oxidative damage. In this study we wished to investigate the possible involvement of this kinase in signalling peroxide stress and nitrogen deprivation. Results: Quantitative RT-PCR experiments revealed that the pkn22 gene is induced in response to peroxide stress and to combined nitrogen starvation. Electrophoretic motility assays indicated that the pkn22 promoter is recognized by the global transcriptional regulators FurA and NtcA. Transcriptomic analysis comparing a pkn22-insertion mutant and the wild type strain indicated that this kinase regulates genes involved in important cellular functions such as photosynthesis, carbon metabolism and iron acquisition. Since metabolic changes may lead to oxidative stress, we investigated whether this is the case with nitrogen starvation. Our results rather invalidate this hypothesis thereby suggesting that the function of Pkn22 under nitrogen starvation is independent of its role in response to peroxide stress. Conclusions: Our analyses have permitted a more complete functional description of Ser/Thr kinase in Nostoc. We have decrypted the transcriptional regulation of the pkn22 gene, and analysed the whole set of genes under the control of this kinase in response to the two environmental changes often encountered by cyanobacteria in their natural habitat: oxidative stress and nitrogen deprivation

Prediction of recurrence of ischemic stroke within 1 year of discharge based on machine learning MRI radiomics

PurposeThis study aimed to investigate the value of a machine learning-based magnetic resonance imaging (MRI) radiomics model in predicting the risk of recurrence within 1 year following an acute ischemic stroke (AIS).MethodsThe MRI and clinical data of 612 patients diagnosed with AIS at the Second Affiliated Hospital of Nanchang University from March 1, 2019, to March 5, 2021, were obtained. The patients were divided into recurrence and non-recurrence groups according to whether they had a recurrent stroke within 1 year after discharge. Randomized splitting was used to divide the data into training and validation sets using a ratio of 7:3. Two radiologists used the 3D-slicer software to label the lesions on brain diffusion-weighted (DWI) MRI sequences. Radiomics features were extracted from the annotated images using the pyradiomics software package, and the features were filtered using the Least Absolute Shrinkage and Selection Operator (LASSO) regression analysis. Four machine learning algorithms, logistic regression (LR), Support Vector Classification (SVC), LightGBM, and Random forest (RF), were used to construct a recurrence prediction model. For each algorithm, three models were constructed based on the MRI radiomics features, clinical features, and combined MRI radiomics and clinical features. The sensitivity, specificity, and area under the receiver operating characteristic (ROC) curve (AUC) were used to compare the predictive efficacy of the models.ResultsTwenty features were selected from 1,037 radiomics features extracted from DWI images. The LightGBM model based on data with three different features achieved the best prediction accuracy from all 4 models in the validation set. The LightGBM model based solely on radiomics features achieved a sensitivity, specificity, and AUC of 0.65, 0.671, and 0.647, respectively, and the model based on clinical data achieved a sensitivity, specificity, and AUC of 0.7, 0.799, 0.735, respectively. The sensitivity, specificity, and AUC of the LightGBM model base on both radiomics and clinical features achieved the best performance with a sensitivity, specificity, and AUC of 0.85, 0.805, 0.789, respectively.ConclusionThe ischemic stroke recurrence prediction model based on LightGBM achieved the best prediction of recurrence within 1 year following an AIS. The combination of MRI radiomics features and clinical data improved the prediction performance of the model

Elucidation of molecular mechanisms involved diazotrophic cyanobacteria Anabaena sp. PCC 7120 in response to oxidative stress and combined nitrogen starvation

La photosynthèse oxygénique peut être le lieu de formation des Formes Réactives de l’Oxygéne (FROs). Les FROs altèrent toutes les macromolécules de la cellule, générant ainsi un stress oxydant. Toute perturbation du métabolisme cellulaire peut conduire à ce type de stress. Les cyanobactérie sétant les premiers organismes à avoir émis de l’oxygène sur terre, elles ont du développer très tôt au cours de l’évolution des mécanismes de perception et de défense pour lutter contre ce stress. Nous nous sommes intéressés à l’étude des mécanismes qui permettent à la cyanobactérie filamentuse et diazotrophe Anabaena PCC 7120 de s’adapter à diverses conditions de stress et de carence : stress oxydant, carence en fer et en azote combiné. En réponse à une carence en azote combiné, elle différencie en 24 h des hétérocystes : cellules spécialisées dans la fixation de l’azote atmosphérique. Nous avons étudié la réponse transcriptomique globale de cette bactérie à la fois au stress oxydant et à la carence en fer et nous avons établit la connection existant entre ces deux stress. Nous avons pu identifier le régulateur transcriptionnel pleiotrope impliqué dans la perception et la signalisation du stress peroxyde et nous en avons élucidé le mécanisme d’action. Nous avons également étudié une Ser/Thr kinase qui joue un rôle important à la fois dans la réponse au stress oxydant et à la carence en azote combiné. Notre étude a montré que cette kinase pourrait être le lien moléculaire entre ces deux conditions, puisque une cible potentielle de cette kinase semble être la protéine HetR qui est le régulateur clé du processus de différenciation cellulaire.Oxygenic photosynthesis may generate Reactive forms of Oxygéne (ROS). These reactive oxygen species can damage all the macromolecules of the cell, inducing oxidative stress. Any disruption of cellular metabolism can lead to oxidant damage. Cyanobacteria were the first organisms producing oxygen on Earth, they therefore had to develop very early during evolution the mechanisms of perception and defence to cope with this tstress. We are interested in studying the mechanisms that allow the diazotrophic filamentous cyanobacterium Anabaena PCC 7120 to adapt to various conditions of stress and stravations: oxidative stress, iron and combined nitrogen starvations. Anabaena PCC 7120 is a simple model for the study of cell differentiation. In response to combined nitrogen stravation it can differentiate heterocysts, cell specialized in molecular nitrogen fixation. We studied the global transcriptomic response of this bacterium to both oxidative stress and iron deficiency and we establish the crosstalk between these two stresses. We were able to identify the global transcriptional regulator involved in the perception and in the signaling of a peroxide stress. Its mechanism of action was elucidated. We also studied a Ser / Thr kinase that plays an important role both in the response to oxidative stress and combined nitrogen stravation. Our study showed that this kinase may be the molecular link between these two conditions, as a potential target of this kinase appears to be the HetR protein which is the key regulator of cellular differentiation process

Cellular apoptosis of hemocytes from Dendrolimus tabulaeformis Tsai et Liu larvae induced with the secondary metabolites of Beauveria brongniartii (Sacc.) Petch.

To investigate the effect of the secondary metabolites of entomopathogenic fungus on the hemocyte immunity of host insect, the secondary metabolite complex (SMC) of Beauveriabrongniartii was used in three concentrations (5.5, 55, and 550 µg/mL), and the 4(th) instar larvae of the pine caterpillar Dendrolimustabulaeformis were employed as host insects. The larvae were inoculated with the SMC solutions by injection in bioassays. Apoptosis of the larval hemocytes was observed using fluorescence microscopy (FM), transmission electron microscopy (TEM), and flow cytometry (FCM). The FM results showed that in the treated groups, larval hemocytes exhibited symptoms of early apoptosis at 6 h post-treatment by radiating a non-uniform kelly fluorescence and exhibited symptoms of late apoptosis at 12 h post-treatment by radiating a non-uniform orange fluorescence. Under TEM, the following ultra-structural changes associated with apoptosis of the larval hemocytes were observed in the treated groups: the nuclei were hypertrophied, slight folds were on the nuclear envelope, the chromatin became concentrated, the mitochondrial cristae disappeared or were disorderly, most cells developed blebs, and fibrillar aggregation appeared and accumulated in the cytoplasm. Apoptosis of the larval hemocytes was detected by FCM at 6 h post-treatment; the percentage of early apoptotic cells in the SMC 5.5, 55, and 550 µg/mL treatment groups were 11.93%, 13.10%, and 18.42%, respectively. Late apoptosis first occurred at 12 h post-treatment; the highest rate of apoptosis was 36.54 ± 4.37% at 24 h post-treatment in the SMC 55 µg/mL treatment group. In general, the cellular apoptosis rate was positively correlated with the SMC concentration and the time post-treatment. These results indicate that secondary metabolites of B. brongniartii are able to attack the hemocytes of D. tabulaeformis larvae and induce cellular apoptosis, thereby providing new evidence that secondary metabolites of mycopathogens can act on host immune systems

The Pkn22 Kinase of Nostoc PCC 7120 Is Required for Cell Differentiation via the Phosphorylation of HetR on a Residue Highly Conserved in Genomes of Heterocyst-Forming Cyanobacteria

International audienceHanks-type kinases encoding genes are present in most cyanobacterial genomes. Despite their widespread pattern of conservation, little is known so far about their role because their substrates and the conditions triggering their activation are poorly known. Here we report that under diazotrophic conditions, normal heterocyst differentiation and growth of the filamentous cyanobacterium Nostoc PCC 7120 require the presence of the Pkn22 kinase, which is induced under combined nitrogen starvation conditions. By analyzing the phenotype of pkn22 mutant overexpressing genes belonging to the regulatory cascade initiating the development program, an epistatic relationship was found to exist between this kinase and the master regulator of differentiation, HetR. The results obtained using a bacterial two hybrid approach indicated that Pkn22 and HetR interact, and the use of a genetic screen inducing the loss of this interaction showed that residues of HetR which are essential for this interaction to occur are also crucial to HetR activity both in vitro and in vivo. Mass spectrometry showed that HetR co-produced with the Pkn22 kinase in Escherichia coli is phosphorylated on Serine 130 residue. Phosphoablative substitution of this residue impaired the ability of the strain to undergo cell differentiation, while its phosphomimetic substitution increased the number of heterocysts formed. The Serine 130 residue is part of a highly conserved sequence in filamentous cyanobacterial strains differentiating heterocysts. Heterologous complementation assays showed that the presence of this domain is necessary for heterocyst induction. We propose that the phosphorylation of HetR might have been acquired to control heterocyst differentiation

Effect of acidic and red-ox sites over modified ZSM-5 surface on selectivity in oxidation of toluene

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A new method of “one blow and two washes” for the analysis of stain categories and content of down raw materials

The down raw materials collected from the slaughterhouses generally adhered with an excessive amount of dust and complex stains which hinder them to be utilized directly for down textiles processing. Water washing is an important core process of down raw material processing for dust removal, decontamination, and sterilization of down fibers. However, there is no standard method to analyze the stain category and content of down raw materials, so it can only rely on traditional experience to wash down, which may cause problems including insufficient or excessive washing, and serious damage to the quality of down fibers. Thus, this study has been specifically designed and developed a method dedicated to categorizing stain types and content of down raw materials. This novel design process is named as “one blow and two washes”, which is composed of three steps to analyze dust content, water-soluble stains, and grease stains in down raw material more accurately. In this experiment, the stain analysis of six different sources of down raw materials was carried out, scanning electron microscope (SEM) and super depth of field 3D microscope system were used to characterize down samples. The results showed that the “one blow and two washes” method could accurately be used to distinguish stain categories of different sources of down raw material with a small average variance value and a coefficient of variation less than 12%, indicating that this experimental method has high reliability and good reproducibility. These research results will provide an important basis for the design of the washing agent dosage and process parameters for raw down, which is expected to achieve the precise washing process of raw down and improve the quality of down

X-linked inheritances recessive of congenital nystagmus and autosomal dominant inheritances of congenital cataracts coexist in a Chinese family: a case report and literature review

Abstract Background Congenital nystagmus (CN) and congenital cataracts are distinct eye diseases and are usually isolated. Cases with CN and congenital cataracts caused by different genes in one family have been rarely reported. Case presentation A 27-year-old man presented with CN and congenital cataracts and he underwent cataract extraction 2 weeks after birth. Three years later, he had posterior chamber intraocular lens implantation. The proband’s mother was only afflicted by bilateral lens opacities. Lensectomy was performed in both eyes at age 15. The proband’s daughter had bilateral central cataracts and no nystagmus. She had undergone cataract extraction when she was two months old. In this family, 8 affected individuals were affected by bilateral cataracts, and three of them presented with CN. The genetic analysis was performed using a specific Hereditary Ophthalmological Disease Gene Panel on proband and his parents (one of which was a patient). PCR and Sanger sequencing verified the presence of these variants in all members of the family. The novel mutation, c.498-3C > T, in FRMD7 explains why X-Linked recessive inheritance of CN was found in a subset of patients. A heterozygous mutation of the GJA8 gene (c.139G > C), was identified in all patients and thus explains the autosomal dominant pattern of inheritance of congenital cataracts within the family. Conclusions This is the first time that FRMD7 and GJA8 gene mutations have been linked to the pathogenesis of a family with both CN and congenital cataracts. The phenomenon of two different genetic patterns coexisting in one family is rare

Dietary Bacitracin Methylene Disalicylate Improves Growth Performance by Mediating the Gut Microbiota in Broilers

The growth performance of livestock and poultry has always been a concern. However, much work is currently focused on the selection of breeds and diets to improve the growth performance of livestock and poultry. Furthermore, numerous studies have shown that the gut microbiota is closely related to the growth performance of livestock and poultry. At present, there are many reports on the impact of antibiotic intervention on the structure of gut microbiota. However, there are few reports on the influence of antibiotic intervention on the structure of intestinal microbes and the effect of this change on growth performance. Bacitracin methylene disalicylate (BMD) intervention changes the microbial structure in the caecum of broilers at different growth stages, as shown in this study. To further reveal the potential relationship between gut microbiota changes and growth performance caused by BMD intervention, correlation analysis was used for analysis. A total of 144 1-day-old male Cobb-Vantress were randomly divided into two groups. In addition to antibiotic-free starter mash diets, starter mash diets supplemented with 55 mg/kg BMD were also used, called the CON group and the BMD group, and lasted 28 days. (1) These study results showed that adding BMD to the diet had a significant effect on the growth performance of broilers. Compared with the CON group, the body weight of the BMD group increased significantly by 11.08% and 20.13% on Days 14 and 28, respectively (p &lt; 0.05). Similarly, at 0&ndash;14, 14&ndash;28 and 0&ndash;28 days of age, the average daily gain of the BMD group increased significantly by 12.28%, 24.49% and 20.80%, respectively. The average daily feed intake of the BMD group increased significantly by 18.28%, 27.39% and 24.97% (p &lt; 0.05). In addition, at 0&ndash;28 days of age, the feed conversion ratio increased significantly by 5.5% (p &lt; 0.05). (2) Alpha diversity results show that BMD intervention has an impact on gut microbiota at different growth stages. (3) The early intervention significantly affected 7 taxa by Day 14, followed by 22 taxa by Day 28, which is similar to the results in the caecal flora. Compared with the CON group, the Christensenellaceae R-7 group had the highest linear discriminant analysis (LDA) score on Day 28. In addition, Pearson&rsquo;s correlation analysis showed that the Lachnospiraceae FCS020 group was significantly negatively correlated with growth performance. In general, these results indicate that dietary supplementation of BMD has an effect on broiler gut microbiota structure and growth performance. However, changes in growth performance may be caused by the gut microbiota structure