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This paper outlines a novel, non-invasive procedure to obtain DNA from Mexican tarantulas (Brachypelma spp.) using exuvia. These species are important in the pet trade and species identification is important for international wildlife law enforcement. Mitochondrial DNA sequence from the cytochrome c oxidase subunit I gene was used to investigate the relationship between various Brachypelma spp. This phylogeny was used as a framework to assign unknown specimens and spiderlings to species. The benefits to conservation, research, and international wildlife law enforcement that are gained by the ability to accurately identify species without the death of the specimen are explored. Our data also suggest that there is no support for the genus Brachypelmides as some authors have proposed and upholds the synonymy of Locht et al. (1999) J Arachnol 27:196-200. [ABSTRACT FROM AUTHOR]

This article is a critique of the meaning and importance of some of the recently published work in the field whose development as a special area began with papers by Zuckerkandl & Pauling and Pauling & Zuckerkandl entitled Chemical Paleogenetics, Molecular Restoration Studies of Extinct Forms of Life and the paper by Margoliash on the evolution of cytochrome c. The primary emphasis is on conclusions and inferences derivable from amino acid sequences, followed bya section on nucleotide sequences. These are preceded by a brief indication of some of the literature that antedates the origin of the genetic code as it now stands. This will not treat immunoglobulin evolution, reviewed by Gally & Edelma and Smith, Hood & Fitch or related material on the evolution of enzymes and proteins reviewed by Smith and Arnheim. The functional interpretation of these changes has made significant progress [Dickerson and Takano et al]. For the evolution of metabolic pathways, see brew, Vogel, and Lejohn. [ABSTRACT FROM AUTHOR]

Amino acid analyses of L-lactate dehydrogenase from baker's show that the minimum molecular weight (53 000 daltons) of the protein is much lower than found in the literature (80 000). This result, combined with those reported in the following papers, leads to a revision of the dimeric model generally accepted for cytochrome b2 [ABSTRACT FROM AUTHOR]

Amino acid analyses of L-lactate dehydrogenase from baker's show that the minimum molecular weight (53 000 daltons) of the protein is much lower than found in the literature (80 000). This result, combined with those reported in the following papers, leads to a revision of the dimeric model generally accepted for cytochrome b2 [ABSTRACT FROM AUTHOR]

Background: Cystic echinococcosis (CE) caused by the Echinococcus granulosus, is a major public health problem worldwide, including India. The different genotypes of E. granulosus responsible for human hydatidosis have been reported from endemic areas throughout the world. However, the genetic characterization of E. granulosus infecting the human population in India is lacking. The aim of study was to ascertain the genotype(s) of the parasite responsible for human hydatidosis in North India. Methodology/Principal Findings: To study the transmission patterns of E. granulosus, genotypic analysis was performed on hydatid cysts obtained from 32 cystic echinococcosis (CE) patients residing in 7 different states of North India. Mitochondrial cytochrome c oxidase subunit1 (cox1) sequencing was done for molecular identification of the isolates. Most of the CE patients (30/32) were found to be infected with hydatid cyst of either G3 (53.1%) or G1 (40.62%) genotype and one each of G5 (cattle strain) and G6 (camel strain) genotype. Conclusions/Significance: These findings demonstrate the zoonotic potential of G1 (sheep strain) and G3 (buffalo strain) genotypes of E. granulosus as these emerged as predominant genotypes infecting the humans in India. In addition to this, the present study reports the first human CE case infected with G5 genotype (cattle strain) in an Asian country and presence of G6 genotype (camel strain) in India. The results may have important implications in the planning of control strategies for human hydatidosis. [ABSTRACT FROM AUTHOR]

Background: Methotrexate is a chemotherapeutic agent used to treat a variety of cancers. However, the occurrence of resistance limits its effectiveness. Cytochrome c in its reduced state is less capable of triggering the apoptotic cascade. Thus, we set up to study the relationship among redox state of cytochrome c, apoptosis and the development of resistance to methotrexate in MCF7 human breast cancer cells. Results: Cell incubation with cytochrome c-reducing agents, such as tetramethylphenylenediamine, ascorbate or reduced glutathione, decreased the mortality and apoptosis triggered by methotrexate. Conversely, depletion of glutathione increased the apoptotic action of methotrexate, showing an involvement of cytochrome c redox state in methotrexate-induced apoptosis. Methotrexate-resistant MCF7 cells showed increased levels of endogenous reduced glutathione and a higher capability to reduce exogenous cytochrome c. Using functional genomics we detected the overexpression of GSTM1 and GSTM4 in methotrexate-resistant MCF7 breast cancer cells, and determined that methotrexate was susceptible of glutathionylation by GSTs. The inhibition of these GSTM isoforms caused an increase in methotrexate cytotoxicity in sensitive and resistant cells. Conclusions: We conclude that overexpression of specific GSTMs, GSTM1 and GSTM4, together with increased endogenous reduced glutathione levels help to maintain a more reduced state of cytochrome c which, in turn, would decrease apoptosis, thus contributing to methotrexate resistance in human MCF7 breast cancer cells. [ABSTRACT FROM AUTHOR]

Productive HIV infection of CD4+ T cells leads to a caspase-independent cell death pathway associated with lysosomal membrane permeabilization (LMP) and cathepsin release, resulting in mitochondrial outer membrane permeabilization (MOMP). Herein, we demonstrate that HIV infection induces damage-regulated autophagy modulator (DRAM) expression in a p53-dependent manner. Knocking down the expression of DRAM and p53 genes with specific siRNAs inhibited autophagy and LMP. However, inhibition of Atg5 and Beclin genes that prevents autophagy had a minor effect on LMP and cell death. The knock down of DRAM gene inhibited cytochrome C release, MOMP and cell death. However, knocking down DRAM, we increased viral infection and production. Our study shows for the first time the involvement of DRAM in host-pathogen interactions, which may represent a mechanism of defense via the elimination of infected cells. [ABSTRACT FROM AUTHOR]

Nelfinavir (NLF), an antiretroviral agent, preserves mitochondrial membranes integrity and protects mature brain against ischemic injury in rodents. Our study demonstrates that in neonatal mice NLF significantly limits mitochondrial calcium influx, the event associated with protection of the brain against hypoxic-ischemic insult (HI). Compared to the vehicle-treated mice, cerebral mitochondria from NLF-treated mice exhibited a significantly greater tolerance to the Ca2+-induced membrane permeabilization, greater ADP-phosphorylating activity and reduced cytochrome C release during reperfusion. Pre-treatment with NLF or Ruthenium red (RuR) significantly improved viability of murine hippocampal HT-22 cells, reduced Ca2+ content and preserved membrane potential (Ψm) in mitochondria following oxygen-glucose deprivation (OGD). Following histamine-stimulated Ca2+ release from endoplasmic reticulum, in contrast to the vehicle-treated cells, the cells treated with NLF or RuR also demonstrated reduced Ca2+ content in their mitochondria, the event associated with preserved Ψm. Because RuR inhibits mitochondrial Ca2+ uniporter, we tested whether the NLF acts via the mechanism similar to the RuR. However, in contrast to the RuR, in the experiment with direct interaction of these agents with mitochondria isolated from naïve mice, the NLF did not alter mitochondrial Ca2+ influx, and did not prevent Ca2+ induced collapse of the Ψm. These data strongly argues against interaction of NLF and mitochondrial Ca2+ uniporter. Although the exact mechanism remains unclear, our study is the first to show that NLF inhibits intramitochondrial Ca2+ flux and protects developing brain against HI-reperfusion injury. This novel action of NLF has important clinical implication, because it targets a fundamental mechanism of post-ischemic cell death: intramitochondrial Ca2+ overload → mitochondrial membrane permeabilization → secondary energy failure. [ABSTRACT FROM AUTHOR]

Fluorescent protein based signaling probes are emerging as valuable tools to study cell signaling because of their ability to provide spatio- temporal information in non invasive live cell mode. Previously, multiple fluorescent protein probes were employed to characterize key events of apoptosis in diverse experimental systems. We have employed a live cell image based approach to visualize the key events of apoptosis signaling induced by zerumbone, the active principle from ginger Zingiber zerumbet, in cancer cells that enabled us to analyze prominent apoptotic changes in a hierarchical manner with temporal resolution. Our studies substantiate that mitochondrial permeabilisation and cytochrome c dependent caspase activation dominate in zerumbone induced cell death. Bax activation, the essential and early event of cell death, is independently activated by reactive oxygen species as well as calpains. Zerumbone failed to induce apoptosis or mitochondrial permeabilisation in Bax knockout cells and over-expression of Bax enhanced cell death induced by zerumbone confirming the essential role of Bax for mitochondrial permeabilsation. Simultaneous inhibition of reactive oxygen species and calpain is required for preventing Bax activation and cell death. However, apoptosis induced by zerumbone was prevented in Bcl 2 and Bcl-XL over-expressing cells, whereas more protection was afforded by Bcl 2 specifically targeted to endoplasmic reticulum. Even though zerumbone treatment down-regulated survival proteins such as XIAP, Survivin and Akt, it failed to affect the pro-apoptotic proteins such as PUMA and BIM. Multiple normal diploid cell lines were employed to address cytotoxic activity of zerumbone and, in general, mammary epithelial cells, endothelial progenitor cells and smooth muscle cells were relatively resistant to zerumbone induced cell death with lesser ROS accumulation than cancer cells. [ABSTRACT FROM AUTHOR]

We explored possible links between vector activity and genetic diversity in introduced populations of Limnoperna fortunei by characterizing the genetic structure in native and introduced ranges in Asia and South America. We surveyed 24 populations: ten in Asia and 14 in South America using the mitochondrial cytochrome c oxidase subunit I (COI) gene, as well as eight polymorphic microsatellite markers. We performed population genetics and phylogenetic analyses to investigate population genetic structure across native and introduced regions. Introduced populations in Asia exhibit higher genetic diversity (HE = 0.667–0.746) than those in South America (HE = 0.519–0.575), suggesting higher introduction effort for the former populations. We observed pronounced geographical structuring in introduced regions, as indicated by both mitochondrial and nuclear markers based on multiple genetic analyses including pairwise ФST, FST, Bayesian clustering method, and three-dimensional factorial correspondence analyses. Pairwise FST values within both Asia (FST = 0.017–0.126, P = 0.000–0.009) and South America (FST = 0.004–0.107, P = 0.000–0.721) were lower than those between continents (FST = 0.180–0.319, P = 0.000). Fine-scale genetic structuring was also apparent among introduced populations in both Asia and South America, suggesting either multiple introductions of distinct propagules or strong post-introduction selection and demographic stochasticity. Higher genetic diversity in Asia as compared to South America is likely due to more frequent propagule transfers associated with higher shipping activities between source and donor regions within Asia. This study suggests that the intensity of human-mediated introduction vectors influences patterns of genetic diversity in non-indigenous species. [ABSTRACT FROM AUTHOR]

Cardiolipin, a major component of mitochondria, is critical for mitochondrial functioning including the regulation of cytochrome c release during apoptosis and proper electron transport. Mitochondrial cardiolipin with its unique bulky amphipathic structure is a potential substrate for phospholipase A2 (PLA2) in vivo. We have developed mass spectrometric methodology for analyzing PLA2 activity toward various cardiolipin forms and demonstrate that cardiolipin is a substrate for sPLA2, cPLA2 and iPLA2, but not for Lp-PLA2. Our results also show that none of these PLA2s have significant PLA1 activities toward dilyso-cardiolipin. To understand the mechanism of cardiolipin hydrolysis by PLA2, we also quantified the release of monolyso-cardiolipin and dilyso-cardiolipin in the PLA2 assays. The sPLA2s caused an accumulation of dilyso-cardiolipin, in contrast to iPLA2 which caused an accumulation of monolyso-cardiolipin. Moreover, cardiolipin inhibits iPLA2 and cPLA2, and activates sPLA2 at low mol fractions in mixed micelles of Triton X-100 with the substrate 1-palmitoyl-2-arachidonyl-sn-phosphtidylcholine. Thus, cardiolipin functions as both a substrate and a regulator of PLA2 activity and the ability to assay the various forms of PLA2 is important in understanding its function. [ABSTRACT FROM AUTHOR]

Two sulfate-reducing bacteria, which also reduce arsenate, were isolated; both organisms oxidized lactate incompletely to acetate. When using lactate as the electron donor, one of these organisms, Desulfomicrobium strain Ben-RB, rapidly reduced (doubling time = 8 h) 5.1 mM arsenate at the same time it reduced sulfate (9.6 mM). Sulfate reduction was not inhibited by the presence of arsenate. Arsenate could act as the terminal electron acceptor in minimal medium (doubling time = 9 h) in the absence of sulfate. Arsenate was reduced by a membrane-bound enzyme that is either a c-type cytochrome or is associated with such a cytochrome; benzyl-viologen-dependent arsenate reductase activity was greater in cells grown with arsenate/sulfate than in cells grown with sulfate only. The second organism, Desulfovibrio strain Ben-RA, also grew (doubling time = 8 h) while reducing arsenate (3.1 mM) and sulfate (8.3 mM) concomitantly. No evidence was found, however, that this organism is able to grow using arsenate as the terminal electron acceptor. Instead, it appears that arsenate reduction by the Desulfovibrio strain Ben-RA is catalyzed by an arsenate reductase that is encoded by a chromosomally-borne gene shown to be homologous to the arsC gene of the Escherichia coli plasmid, R773 ars system. [ABSTRACT FROM AUTHOR]

Cytochrome c′ of Methylococcus capsulatus Bath is involved in electron flow from the enzyme responsible for hydroxylamine oxidation, cytochrome P460, to cytochrome c555. This cytochrome is spectrally similar to other cytochromes c′ but is larger (16,000 Da) and has a lower midpoint potential (–205 mV). By a combination of Edman degradation, mass spectroscopy, and gene sequencing, we have obtained the primary structure of cytochrome c′ from M. capsulatus Bath. The cytochrome shows low sequence similarity to other cytochromes c′, only residues R12, Y53, G56, and the C-terminal heme-binding region (GXXCXXCHXXXK) being conserved. In contrast, cytochrome c′ from M. capsulatus Bath shows considerable sequence similarity to cytochromes P460 from M. capsulatus Bath (31% identity) and from Nitrosomonas europaea (18% identity). This suggests that P460-type cytochromes may have originated from a c′-type cytochrome which developed a covalent cross-link between a lysine residue and the c′-heme. [ABSTRACT FROM AUTHOR]

Ion-exchange chromatography of the total cytochrome c extracted from a wild-type laboratory strain of Saccharomyces cerevisiae shows the presence of two minor cytochrome c peaks, besides the already known iso-1 and iso-2 cytochromes c. Their amino acid composition is very similar to that of iso-1, but one of them lacks the ε-N-trimethylated lysine residue characteristic of yeast cytochromes c, thus excluding, at least for this peak, an artifactual origin. [ABSTRACT FROM AUTHOR]

Cites key research findings indicating a new and earlier pro-apoptotic role for cytochrome c. Role of cytochrome c release from the mitochondria as a component of the apoptotic cell-death program; Effects of the cytochrome c-catalyzed peroxidation of cardiolipin on the binding of cytochrome c to the mitochondrial inner membrane and the permeabilization of the outer membrane; Implications for chemistry and biology.