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51.
SiuShing Wong Zachary M Wilmott Saroj Saurya Ines AlvarezRodrigo Felix Y Zhou KwaiYin Chau Alain Goriely Jordan W Raff 《The EMBO journal》2022,41(11)
Mitotic centrosomes are formed when centrioles start to recruit large amounts of pericentriolar material (PCM) around themselves in preparation for mitosis. This centrosome “maturation” requires the centrioles and also Polo/PLK1 protein kinase. The PCM comprises several hundred proteins and, in Drosophila, Polo cooperates with the conserved centrosome proteins Spd‐2/CEP192 and Cnn/CDK5RAP2 to assemble a PCM scaffold around the mother centriole that then recruits other PCM client proteins. We show here that in Drosophila syncytial blastoderm embryos, centrosomal Polo levels rise and fall during the assembly process—peaking, and then starting to decline, even as levels of the PCM scaffold continue to rise and plateau. Experiments and mathematical modelling indicate that a centriolar pulse of Polo activity, potentially generated by the interaction between Polo and its centriole receptor Ana1 (CEP295 in humans), could explain these unexpected scaffold assembly dynamics. We propose that centrioles generate a local pulse of Polo activity prior to mitotic entry to initiate centrosome maturation, explaining why centrioles and Polo/PLK1 are normally essential for this process. 相似文献
52.
Albert K. Dearden Brian Callahan Patrick Van Roey Zhong Li Utsav Kumar Marlene Belfort Saroj K. Nayak 《Protein science : a publication of the Protein Society》2013,22(5):557-563
Protein splicing is an autocatalytic process where an “intein” self‐cleaves from a precursor and ligates the flanking N‐ and C‐“extein” polypeptides. Inteins occur in all domains of life and have myriad uses in biotechnology. Although the reaction steps of protein splicing are known, mechanistic details remain incomplete, particularly the initial peptide rearrangement at the N‐terminal extein/intein junction. Recently, we proposed that this transformation, an N‐S acyl shift, is accelerated by a localized conformational strain, between the intein's catalytic cysteine (Cys1) and the neighboring glycine (Gly‐1) in the N‐extein. That proposal was based on the crystal structure of a catalytically competent trapped precursor. Here, we define the structural origins and mechanistic relevance of the conformational strain using a combination of quantum mechanical simulations, mutational analysis, and X‐ray crystallography. Our results implicate a conserved, but largely unstudied, threonine residue of the Ssp DnaE intein (Thr69) as the mediator of conformational strain through hydrogen bonding. Further, the strain imposed by this residue is shown to position the splice junction in a manner that enhances the rate of the N‐S acyl shift substantially. Taken together, our results not only provide fundamental understanding of the control of the first step of protein splicing but also have important implications in various biotechnological applications that require precursor manipulation. 相似文献
53.
Modulating furin activity with designed mini-PDX peptides: synthesis and in vitro kinetic evaluation
A peptide was designed from reactive site loop structure of alpha1 Antitrypsin Portland known as alpha1 PDX as a novel mini-PDX inhibitor of furin. The sequence was derived from (367-394) that contains the crucial furin cleavage motif RIPR382. A P3 mutant replacing Ile380 by Leu was prepared as a first model peptide. A Cys residue was inserted at each terminal of the peptide for purpose of cyclisation which was accomplished by air or iodine-induced oxidation. This mini-PDX peptide both cyclic and acyclic form inhibited in vitro furin activity (IC50 in nM) when measured against either substrates Boc-RVRRdown double arrow MCA or QVEGF-C [Abz-QVHSIIRRdown double arrow SLP-Y(NO2)-A-CONH2, Abz=2-amino benzoic acid and Y(NO2)=3-nitro tyrosine], latter being derived from vascular endothelial growth factor-C (VEGF-C) processing site. The geometrically constrained structure mimicking PDX reactive loop is crucial for enzyme inhibition. Our study further revealed that both mini-PDX peptides inactivate furin in a slow tight binding manner, with disulfide-bridged cyclic form being slightly more potent. Unlike PDX, these peptides inhibit furin via a different mechanistic pathway. The study provides an alternate strategy for development of efficient peptide-based inhibitors of Proprotein Convertases including furin. 相似文献
54.
55.
Demirci C Gargili A Kandil A Cetinkaya H Uyaner I Boynuegri B Gumustas MK 《The Chinese journal of physiology》2006,49(6):326-334
The roles of nitric oxide production and oxidative process were studied in mice infected with Toxocara canis and treated with aminoguanidine which is a specific inhibitor of inducible nitric oxide synthase (iNOS). Relations of nitric oxide synthase inhibition and tissue pathology were assessed by biochemical, histological and immunohistochemical methods. In experiments, Balb/c albino mice were inoculated with T. canis eggs either with or without aminoguanidine treatment or alone, at 24th, 48th hours and on 7th days. LPx and SOD values in liver tissue and plasma were measured. Liver and lung tissues were evaluated for the pathological lesions. The expression of eNOS and iNOS in both tissues were studied with immunohistochemistry in the same intervals. We observed significant differences between T. canis infected and aminoguanidine treated animals. Larval toxocarosis led to oxidative stress elevation in plasma. Microscopic examination of the liver histological sections revealed pathological lesions in the hepatic parenchyma in infected mice. In the mice received T. canis eggs plus aminoguanidine, the sinusoidal areas were enlarged. Histological lesions were more severe at 48 hours after infection. Numbers of eNOS and iNOS expressing epithelial cells were increased in the T. canis infected mice. The activities of eNOS and iNOS were also observed in the body of the larvae which have migrated to lung and liver. As a result, we have demonstrated that in vivo production of eNO and iNO during T. canis infection cause direct host damages and it is strongly related to the oxidative stress. We propose that larval NO can also be effective in larval migration, but it needs further investigation on distribution of NO in larvae. 相似文献
56.
Basak JM Verghese PB Yoon H Kim J Holtzman DM 《The Journal of biological chemistry》2012,287(17):13959-13971
Accumulation of the amyloid β (Aβ) peptide within the brain is hypothesized to be one of the main causes underlying the pathogenic events that occur in Alzheimer disease (AD). Consequently, identifying pathways by which Aβ is cleared from the brain is crucial for better understanding of the disease pathogenesis and developing novel therapeutics. Cellular uptake and degradation by glial cells is one means by which Aβ may be cleared from the brain. In the current study, we demonstrate that modulating levels of the low-density lipoprotein receptor (LDLR), a cell surface receptor that regulates the amount of apolipoprotein E (apoE) in the brain, altered both the uptake and degradation of Aβ by astrocytes. Deletion of LDLR caused a decrease in Aβ uptake, whereas increasing LDLR levels significantly enhanced both the uptake and clearance of Aβ. Increasing LDLR levels also enhanced the cellular degradation of Aβ and facilitated the vesicular transport of Aβ to lysosomes. Despite the fact that LDLR regulated the uptake of apoE by astrocytes, we found that the effect of LDLR on Aβ uptake and clearance occurred in the absence of apoE. Finally, we provide evidence that Aβ can directly bind to LDLR, suggesting that an interaction between LDLR and Aβ could be responsible for LDLR-mediated Aβ uptake. Therefore, these results identify LDLR as a receptor that mediates Aβ uptake and clearance by astrocytes, and provide evidence that increasing glial LDLR levels may promote Aβ degradation within the brain. 相似文献
57.
2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a persistent and ubiquitous environmental contaminant. The health impact of TCDD exposure is of great concern
to the general public. Recent reports have implied that eicosapentaenoic acid (EPA) might be a potential chemopreventive agent
and influence hepatotoxicity. The aim of the current study was to explore the effectiveness of EPA in alleviating the toxicity
of TCDD on primary cultured rat hepatocytes. EPA (5, 10 and 20 μM) was added to cultures alone or simultaneously with TCDD
(5 and 10 μM). Rat hepatocytes were treated with TCDD and EPA for 48 h, and then cytotoxicity was detected by [3-(4,5-dimethyl-thiazol-2-yl)
2,5-diphenyltetrazolium bromide] (MTT) assay and lactate dehydrogenase (LDH) release, while total antioxidant capacity (TAC)
and total oxidative stress (TOS) levels were determined to evaluate the oxidative injury. The DNA damage was also analyzed
by liver micronucleus assay (LMN) and 8-oxo-2-deoxyguanosine (8-OH-dG). The results of MTT and LDH assays showed that TCDD
but not EPA decreased cell viability. TCDD also increased TOS level and significantly decreased TAC level in rat hepatocytes
in a clear dose dependent manner. On the basis of increasing doses, the dioxin caused significant increases of micronucleated
hepatocytes (MNHEPs) and 8-OH-dG as compared to control culture. Whereas, in cultures treated with EPA alone, TOS level did
not change and the level of TAC significantly increased. The presence of EPA with TCDD minimized the toxic effects of the
dioxin on primary hepatocytes cultures. Noteworthy, EPA has a protective effect against TCDD-mediated DNA damages. 相似文献
58.
Jérémie Fournier-Dit-Chabert Victoria Vinader Ana Rita Santos Mariano Redondo-Horcajo Aurore Dreneau Ramkrishna Basak Laura Cosentino Gemma Marston Hamdy Abdel-Rahman Paul M. Loadman Steven D. Shnyder José Fernando Díaz Isabel Barasoain Robert A. Falconer Klaus Pors 《Bioorganic & medicinal chemistry letters》2012,22(24):7693-7696
Colchicine was modified at the 10-OCH3 position of the C-ring by reaction with heterocyclic amines or commercially available amines to afford a library of target colchicinoids in high yields (62–99%). Molecular modeling revealed that the incorporation of the linker groups led to a reduction in entropy and therefore binding affinity when compared with colchicine. Some colchicinoids were shown to be equicytotoxic with colchicine when evaluated in the DLD-1 colon cancer cells and retained activity in resistant A2780AD or HeLa cells with mutant Class III β-tubulin. Importantly, unlike colchicine, the analogues in this study are amenable for prodrug derivatisation and with potential for tumor-selective delivery. 相似文献
59.
Plant defense responses are mediated by elementary regulatory proteins that affect expression of thousands of genes. Over
the last decade, microarray technology has played a key role in deciphering the underlying networks of gene regulation in
plants that lead to a wide variety of defence responses. Microarray is an important tool to quantify and profile the expression
of thousands of genes simultaneously, with two main aims: (1) gene discovery and (2) global expression profiling. Several
microarray technologies are currently in use; most include a glass slide platform with spotted cDNA or oligonucleotides. Till
date, microarray technology has been used in the identification of regulatory genes, end-point defence genes, to understand
the signal transduction processes underlying disease resistance and its intimate links to other physiological pathways. Microarray
technology can be used for in-depth, simultaneous profiling of host/pathogen genes as the disease progresses from infection
to resistance/susceptibility at different developmental stages of the host, which can be done in different environments, for
clearer understanding of the processes involved. A thorough knowledge of plant disease resistance using successful combination
of microarray and other high throughput techniques, as well as biochemical, genetic, and cell biological experiments is needed
for practical application to secure and stabilize yield of many crop plants. This review starts with a brief introduction
to microarray technology, followed by the basics of plant–pathogen interaction, the use of DNA microarrays over the last decade
to unravel the mysteries of plant–pathogen interaction, and ends with the future prospects of this technology. 相似文献
60.
ROR1 is expressed in human breast cancer and associated with enhanced tumor-cell growth 总被引:1,自引:0,他引:1
Zhang S Chen L Cui B Chuang HY Yu J Wang-Rodriguez J Tang L Chen G Basak GW Kipps TJ 《PloS one》2012,7(3):e31127
Receptor-tyrosine-kinase-like orphan receptor 1 (ROR1) is expressed during embryogenesis and by certain leukemias, but not by normal adult tissues. Here we show that the neoplastic cells of many human breast cancers express the ROR1 protein and high-level expression of ROR1 in breast adenocarcinoma was associated with aggressive disease. Silencing expression of ROR1 in human breast cancer cell lines found to express this protein impaired their growth in vitro and also in immune-deficient mice. We found that ROR1 could interact with casein kinase 1 epsilon (CK1ε) to activate phosphoinositide 3-kinase-mediated AKT phosphorylation and cAMP-response-element-binding protein (CREB), which was associated with enhanced tumor-cell growth. Wnt5a, a ligand of ROR1, could induce ROR1-dependent signaling and enhance cell growth. This study demonstrates that ROR1 is expressed in human breast cancers and has biological and clinical significance, indicating that it may be a potential target for breast cancer therapy. 相似文献