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Melvin J. Oliver David L. Ferguson John J. Burke Jeff Velten 《Molecular & general genetics : MGG》1993,239(3):425-434
Tobacco plants were genetically transformed to generate antisense RNA from a gene construct comprised of a full-length cucumber NADH-dependent hydroxypyruvate reductase (HPR) cDNA placed in reverse orientation between the cauliflower mosaic virus 35S promoter and a nopaline synthase termination/polyadenylation signal sequence. In vivo accumulation of antisense HPR RNA within eight independent transgenic tobacco plants resulted in reductions of up to 50% in both native HPR activity and protein accumulation relative to untransformed tobacco plants (mean transgenote HPR activity=67% wild type, mean transgenote HPR protein=63% wild type). However, in contrast to previous reports describing antisense RNA effects in plants, production of the heterologous HPR antisense RNA did not systematically reduce levels of native tobacco HPR mRNA (mean transgenote HPR mRNA level=135% wild type). Simple regression comparison of the steady-state levels of tobacco HPR mRNA to those of HPR antisense RNA showed a weak positive correlation (r value of 0.548, n=9 ; n is wild type control plus eight independent transformants; significant at 85% confidence level), supporting the conclusion that native mRNA levels were not reduced within antisense plants. Although all transgenic antisense plants examined displayed an apparent reduction in both tobacco HPR protein and enzyme activity, there is no clear correlation between HPR activity and the amount of either sense (r=0.267, n=9) or antisense RNA (r=0.175, n=9). This compares to a weak positive correlation between HPR mRNA levels and the amount of HPR activity observed in wild-type SRI tobacco plants (r=0.603, n=5). The results suggest that in vivo production of this heterologous HPR antisense RNA is inhibitory at the level of HPR-specific translation and produces its effect in a manner not dependent upon, nor resulting in, a reduction in steady-state native HPR mRNA levels. In this context, the observed antisense effect appears to differ mechanistically from most antisense systems described to date. 相似文献
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Cuperus R van Kuilenburg AB Leen R Bras J Caron HN Tytgat GA 《Free radical biology & medicine》2011,51(6):1213-1220
To enhance the efficacy of fenretinide (4HPR)-induced reactive oxygen species (ROS) in neuroblastoma, 4HPR was combined with buthionine sulfoximine (BSO), an inhibitor of glutathione (GSH) synthesis, in neuroblastoma cell lines and spheroids, the latter being a three-dimensional tumor model. 4HPR exposure (2.5-10 μM, 24 h) resulted in ROS induction (114-633%) and increased GSH levels (68-120%). A GSH depletion of 80% of basal levels was observed in the presence of BSO (25-100 μM, 24 h). The 4HPR-BSO combination resulted in slightly increased ROS levels (1.1- to 1.3-fold) accompanied by an increase in cytotoxicity (110-150%) compared to 4HPR treatment alone. A correlation was observed between the ROS-inducing capacity of each cell line and the increase in cytotoxicity induced by 4HPR-BSO compared to 4HPR. No significant correlation between baseline antioxidant levels and sensitivity to 4HPR or BSO was observed. In spheroids, 4HPR-BSO induced a strong synergistic growth retardation and induction of apoptosis. Our data show that BSO increased the cytotoxic effects of 4HPR in neuroblastoma monolayers and spheroids in ROS-producing cell lines. This indicates that the 4HPR-BSO combination might be a promising new strategy in the treatment of neuroblastoma. 相似文献
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All-trans-retinoic acid (ATRA) has been shown to inhibit the growth of a number of ovarian tumor cell lines while others have been found to be resistant to retinoid suppression of growth. Interestingly, two synthetic retinoids, CD437 and 4-HPR, inhibit the growth of both ATRA-sensitive (CA-OV-3) and ATRA-resistant (SK-OV-3) ovarian tumor cells. However, in contrast to ATRA, both induce apoptosis. Our goal was to elucidate the mechanism by which these two synthetic retinoids induce apoptosis in ovarian tumor cells. Since it has been documented that apoptosis induction is often mediated by the activation of a cascade of proteases known as caspases, we initially studied the role of caspases in induction of apoptosis by CD437 and 4-HPR. We found that both retinoids induced caspase-3 and caspase-9 enzyme activity. Furthermore, using caspase specific inhibitors we determined that caspase-3 and caspase-9 activity was essential for the induction of apoptosis by these synthetic retinoids since these inhibitors completely blocked CD437 and 4-HPR induced apoptosis. Interestingly, we found that treatment with bongkriekic acid (BA), a mitochondrial membrane depolarization inhibitor, blocked apoptosis, caspase-9 activation and caspase-3 activation induced by both retinoids. Finally, we were able to determine that CD437 treatment induced the translocation of TR3, a nuclear orphan receptor, whereas, 4-HPR did not. Our results suggest that CD437 and 4-HPR initially activate separate pathways to induce mitochondrial depolarization but both utilize mitochondrial depolarization, caspase-9 activation, and caspase-3 activation in the later stages of apoptosis induction. 相似文献
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Increase in p1 values was found between 11 and 30 days of postnatal life in papain-solubilized brush-border enzymes of rat small intestine by means of thin-layer analytical isoelectric focusing in agarose gel. Treatment with neuraminidase converted the acidic forms of enzymes from 11- and 20-day-old rats into the zymogram patterns identical with the adult, more basic forms. The zymograms of the respective enzymes are the same in 30-, 60- and 74-day-old animals and do not change on treatment with neuraminidase. 相似文献
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Mueller C Hutter P Sahli R Bretz A Toutoungi LN Péter O 《FEMS microbiology letters》2000,191(2):199-203
Borrelia garinii is one of the three major Borreliae responsible for Lyme borreliosis in Europe. We have characterized a protein of B. garinii (VS102) and a genomic fragment from the gene encoding this protein was cloned. The DNA sequence of the fragment showed high homology with a known gene of B. burgdorferi sensu stricto. The protein encoded by this gene in B. burgdorferi sensu stricto is a phosphocarrier protein (histidine-containing protein). A mutation T to G polymorphism at codon 57 was found to be specific to B. garinii. A PCR-based approach that allows the rapid detection of this mutation made it possible to specifically discriminate B. garinii from other B. burgdorferi genospecies with high sensitivity and specificity. 相似文献
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Transglutaminase 2 in the balance of cell death and survival 总被引:7,自引:0,他引:7
Transglutaminase 2 (TG2), a multifunctional enzyme with Ca(2+)-dependent protein crosslinking activity and GTP-dependent G protein functions, is often upregulated in cells undergoing apoptosis. In cultured cells TG2 may exert both pro- and anti-apoptotic effects depending upon the type of cell, the kind of death stimuli, the intracellular localization of the enzyme and the type of its activities switched on. The majority of data support the notion that transamidation by TG2 can both facilitate and inhibit apoptosis, while the GTP-bound form of the enzyme generally protects cells against death. In vivo studies confirm the Janus face of TG2 in the initiation of the apoptotic program. In addition, they reveal a further role: the prevention of inflammation, tissue injury and autoimmunity once the apoptosis has already been initiated. This function of TG2 is partially achieved by being expressed and activated also in macrophages digesting apoptotic cells and mediating a crosstalk between dying and phagocytic cells. 相似文献
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Francisco J. Corpas Marina Leterrier Juan C. Begara-Morales Raquel Valderrama Mounira Chaki Javier López-Jaramillo Francisco Luque José M. Palma María N. Padilla Beatriz Sánchez-Calvo Capilla Mata-Pérez Juan B. Barroso 《Biochimica et Biophysica Acta (BBA)/General Subjects》2013