Sphingomonas yanoikuyae B1 is able to utilize toluene, m-xylene, p-xylene, biphenyl, naphthalene, phenanthrene, and anthracene as sole sources of carbon and energy for growth. A forty kilobase region of DNA containing most of the genes for the degradation of these aromatic compounds was previously cloned and sequenced. Insertional inactivation of bphC results in the inability of B1 to grow on both polycyclic and monocyclic compounds. Complementation experiments indicate that the metabolic block is actually due to a polar effect on the expression of bphA3, coding for a ferredoxin component of a dioxygenase. Lack of the ferredoxin results in a nonfunctional polycyclic aromatic hydrocarbon dioxygenase and a nonfunctional toluate dioxygenase indicating that the electron transfer components are capable of interacting with multiple oxygenase components. Insertional inactivation of a gene for a dioxygenase oxygenase component downstream of bphA3 had no apparent effect on growth besides a polar effect on nahD which is only needed for growth of B1 on naphthalene. Insertional inactivation of either xylE or xylG in the meta-cleavage operon results in a polar effect on bphB, the last gene in the operon. However, insertional inactivation of xylX at the beginning of this cluster of genes does not result in a polar effect suggesting that the genes for the meta-cleavage pathway, although colinear, are organized in at least two operons. These experiments confirm the biological role of several genes involved in metabolism of aromatic compounds by S. yanoikuyae B1 and demonstrate the interdependency of the metabolic pathways for polycyclic and monocyclic aromatic hydrocarbon degradation.
Extracts of heterotrophically grown cells ofPseudomonas carboxydovorans were found to contain an inhibitor of carbon monoxide dehydrogenase (CO-DH). The inhibitor activity was not detected in CO-autotrophically grown cells. The inhibitor was extremely stable to heat treatment based on the extent of inhibition of CO-DH activity. The extent of inhibition was proportional to the amount of cell extract added to the reaction mixture.The inhibition was independent of a prior incubation period of the extracts with CO-DH. The inhibitor was precipitable with ammonium sulfate, phenol, and trichloroacetic acid. It was passed through benzoylated dialysis tubing and Amicon ultrafiltration membrane YM2. Denaturing and nondenturing polyacrylamide gel electrophoresis of CO-DH inactivated by inhibitor revealed that the mobilities of native enzyme and subunits were identical to those of active CO-DH. The inhibitor-treated CO-DH retained its original antigenic sites and exhibited enzyme activity upon activity staining. The CO-DH inhibitor of P. carboxydovorans was also active on CO-DHs from Pseudomonas carboxydohydrogena, Acinetobacter sp. strain JC1, and Pseudomonas carboxydoflava.Carbon monoxide dehydrogenase (CO-DH) is the key enzyme in the utilization of CO as the sole energy and carbon source in several aerobic carboxydobacteria (1,5,6,(15)(16)(17)22). The enzyme has generally been considered as an inducible enzyme, with the exception of the CO-DH in Pseudomonas carboxydoflava (1,2,5,15,17). Biochemical and immunological studies revealed that the enzymes from several carboxydobacteria share similar properties, with few exceptions (3,5,(15)(16)(17).We have recently found that Pseudomonas carboxydovorans growing under heterotrophic condition contains an inhibitor of CO-DH from this bacterium. Since this was the first observation of the presence of CO-DH inhibitor in carboxydobacteria and since nothing is known about the molecule, it was of interest to study the inhibitor in cell extracts. The present work was undertaken to study the physiological and biochemical properties of CO-DH inhibitor in cell extracts of P. carboxydovorans.
Background Although glioblastoma (GBM) is the most common primary brain tumor, the best available treatment options are still associated with poor prognosis. Recently, the Hippo/YAP signaling pathway has been emerged as an important driver of GBM. Nevertheless, extensive studies have not yet been focused on the importance of phosphorylation event in regulating endogenous YAP activity in GBM. Here, we sought to elucidate that the modulation and stabilization of YAP/TAZ in Hippo pathway promotes GBM progression. Material and Methods Expression patterns of YAP1, TAZ, CTGF, TEAD4 and LATS1 mRNA were confirmed in human normal and GBM tissue, and patient survival according to YAP1 expression was confirmed in TCGA and Severance cohort. The YAP1 expression patterns in several TSs were classified into SOH and AH according to transcriptome analysis and western blot. For each classified TSs, the characteristics of SOH and AH were comparatively analyzed through immunoprecipitation, nucleus fraction, TEAD4 luciferase assay, and confocal analysis. The effect of YAP1 knockdown/out and overexpression in SOH and AH TSs were measured by WST/ATP analysis, 3D invasion and neurosphere formation, and western blot. In addition, the drug reactivity of SOH and AH TSs was examined by treatment with the YAP1 inhibitors Peptide17 and verteporfin, and the effects of Peptide17, verteporfin and YAP1-shRNA were examined in in-vivo xenograft model. Results YAP1 mRNA levels including TAZ, CTGF, TEAD4 and LATS1 were higher in GBM than in normal tissues. Through transcriptome analysis and western blot, TS can be divided into SOH and AH groups according to the expression level of CTGF, but not YAP1 level. In the case of SOH TS, the binding of TEAD4 was strong in immunoprecipitation, the ratio of YAP1 was higher in the nucleus in the nucleus fractionation and confocal analysis, and the luciferase activity of TEAD4 was high in the TEAD4 luciferase assay. On the other hand, in the case of TS of AH, the opposite trend was observed. When YAP1 was knocked down or out, the proliferation, invasion, and stemness of TS tended to be decrease more in SOH TS than in AH TS, and SOH TS showed more sensitive drug response even in Peptide17 and Verteporfin treatment. In the case of YAP1 overexpression, the opposite trend was observed in SOH TS. In the in-vivo xenograft model, injection of Peptide17, verteporfin, and YAP1-shRNA showed a tendency to increase the survival rate of mice. Conclusion The association of YAP has been described in several cancers, but there has been no study comparing GBM TS by YAP expression and classifying it at the functional level. Now, for the first time, we show that the characteristics of GBM TS can be predicted according to the expression value of YAP, respectively, and YAP is expected to be applied as an important prognostic factor for GBM treatment in the future.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.