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Xiao-Xiao Li Bun Tsoi Yi-Fang Li Hiroshi Kurihara Rong-Rong He 《The journal of histochemistry and cytochemistry》2015,63(5):301-311
Cardiolipin (CL) is a unique dimeric phospholipid that exists almost exclusively in the inner mitochondrial membrane (IMM) in eukaryotic cells. Two chiral carbons and four fatty acyl chains in CL result in a flexible body allowing interactions with respiratory chain complexes and mitochondrial substrate carriers. Due to its high content of unsaturated fatty acids, CL is particularly prone to reactive oxygen species (ROS)-induced oxidative attacks. Under mild mitochondrial damage, CL is redistributed to the outer mitochondrial membrane (OMM) and serves as a recognition signal for dysfunctional mitochondria, which are rapidly sequestered by autophagosomes. However, peroxidation of CL is far greater in response to severe stress than under normal or mild-damage conditions. The accumulation of oxidized CL on the OMM results in recruitment of Bax and formation of the mitochondrial permeability transition pore (MPTP), which releases Cytochrome c (Cyt c) from mitochondria. Over the past decade, the significance of CL in the function of mitochondrial bioenergy has been explored. Moreover, approaches to analyzing CL have become more effective and accurate. In this review, we discuss the unique structural features of CL as well as the current understanding of CL-based molecular mechanisms of mitophagy and apoptosis. 相似文献
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Nacrein-like proteins have carbonic anhydrase (CA)-like domains, but their coding regions are flanked by inserted repeat sequence, such as Gly-X-Asn. Reportedly, nacrein-like proteins show the highest similarity to human carbonic anhydrase 1(α-CA1), possess CA catalytic functions, and play a key role in shell biomineralization. In the present study, two novel nacrein-like proteins were firstly identified from the shell-forming mantle of the Pacific oyster Crassostrea gigas. With numerous analyses, it was identified and characterized that both the nacrein-like proteins F1 and F2 were secreted and most closely related to the nacrein-like protein of California mussel Mytilus californianus via phylogenetic analysis. RT-PCR analysis showed that the nacrein-like proteins F1 and F2 were expressed in multiple tissues and the expression levels remarkably rose after entering the spat stage, which were basically consistent with the increase of calcite fractions in the total shell volume. Surprisingly, the Gly-X-Asn repeat domain, which is distinctive in most nacrein-like proteins, was absent in the two newly identified nacrein-like proteins in C. gigas and replaced with a series of acidic amino acids (D/E). Regardless, nacrein-like proteins in mollusks seem to be vital to the deposition of calcium carbonate and likely perform diverse functions. 相似文献
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Gen-cheng Gong Wen-zhu Fan Di-zheng Li Xiong Tian Shao-jun Chen Yu-cai Fu Wen-can Xu Chi-ju Wei 《PloS one》2015,10(6)
Ectopically expressed Cre recombinase in extrapancreatic tissues in RIP-Cre mice has been well documented. The objective of this study was to find a simple solution that allows for improved beta-cell specific targeting. To this end, the RIP-Cre and reporter CMV-loxP-DsRed-loxP-EGFP expression cassettes were configurated into a one-plasmid and two-plasmid systems, which labeled approximately 80% insulin-positive INS-1 cells after 48 h transfection. However, off-target labeling was robustly found in more than 15% insulin-negative Ad293 cells. When an IRES element was inserted in front of Cre to reduce the translation efficiency, the ratio of recombination between INS-1 and Ad293 cells increased 3-4-fold. Further, a series of Cre mutants were generated by site-directed mutagenesis. When one of the mutants, Cre(H289P) in both configurations, was used in the experiment, the percentage of recombination dropped to background levels in a number of insulin-negative cell lines, but decreased only slightly in INS-1 cells. Consistently, DNA substrate digestion assay showed that the enzymatic activity of Cre(H289P) was reduced by 30-fold as compared to that of wild-type. In this study, we reported the generation of constructs containing RIP and Cre mutants, which enabled enhanced beta-cell specific labeling in vitro. These tools could be invaluable for beta-cell targeting and to the study of islet development. 相似文献
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Glioblastoma multiforme (GBM) is one of the utmost malignant tumors. Excessive angiogenesis and invasiveness are the major reasons for their uncontrolled growth and resistance toward conventional strategies resulting in poor prognosis. In this study, we found that low-dose JSI-124 reduced invasiveness and tumorigenicity of GBM cells. JSI-124 effectively inhibited VEGF expression in GBM cells. In a coculture study, JSI-124 completely prevented U87MG cell–mediated capillary formation of HUVECs and the migration of HUVECs when cultured alone or cocultured with U87MG cells. Furthermore, JSI-124 inhibited VEGF-induced cell proliferation, motility, invasion and the formation of capillary-like structures in HUVECs in a dose-dependent manner. JSI-124 suppressed VEGF-induced p-VEGFR2 activity through STAT3 signaling cascade in HUVECs. Immunohistochemistry analysis showed that the expression of CD34, Ki67, p-STAT3 and p-VEGFR2 protein in xenografts was remarkably decreased. Taken together, our findings provide the first evidence that JSI-124 effectively inhibits tumor angiogenesis and invasion, which might be a viable drug in anti-angiogenesis and anti-invasion therapies. 相似文献
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L Li K Lin I J Kurland J J Correia S J Pilkis 《The Journal of biological chemistry》1992,267(7):4386-4393
To identify those residues involved in fructose 6-phosphate binding to the kinase domain of rat liver 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase site-directed mutations were engineered at Lys194, Arg195, Arg230, and Arg238. The mutant enzymes were purified to homogeneity by anion exchange and Blue-Sepharose chromatography and/or substrate elution from phosphocellulose columns. Circular dichroism experiments demonstrated that all of the single amino acid mutations had no effect on the secondary structure of the protein. In addition, when fructose-2,6-bisphosphatase activity was measured, all mutants had Km values for fructose 2,6-bisphosphate, Ki values for fructose 6-phosphate, and maximal velocities similar to that of the wild-type enzyme. Mutation of Arg195----Ala, or His, had little or no effect on the maximal velocity of the kinase but increased the Km for fructose 6-phosphate greater than 3,000-fold. Furthermore, the Ka for phosphate for Arg195Ala was increased 100-fold compared with the wild-type enzyme. Mutation of Lys194----Ala had no effect on maximal velocity or the Km for fructose 6-phosphate. Mutation of either Arg230 or Arg238----Ala increased the maximal velocity and the Km for fructose-6 phosphate of the kinase by 2-3-fold but had no effect on fructose-2,6-bisphosphatase. However, the Km values for ATP of the Arg230Ala and Arg238Ala mutants were 30-40-fold higher than that for the wild-type enzyme. Mutation of Gly48----Ala resulted in a form with no kinase activity, but fructose-2,6-bisphosphatase activity was identical to that of the wild-type enzyme. The results indicate that: 1) Arg195 is a critical residue for the binding of fructose 6-phosphate to the 6-phospho-fructo-2-kinase domain, and that interaction of the sugar phosphate with Arg195 is highly specific since mutation of the adjacent Lys194----Ala had no effect on fructose 6-phosphate binding; 2) Arg195 also play an important role in the binding of inorganic phosphate; and 3) Gly48 is an important residue in the nucleotide binding fold of 6-phosphofructo-2-kinase and that both Arg230 and Arg238 are also involved in ATP binding; and 4) the bifunctional enzyme has two separate and independent fructose 6-phosphate binding sites. 相似文献