Mitochondrial phospholipid hydroperoxide glutathione peroxidase suppresses apoptosis mediated by a mitochondrial death pathway.

Phospholipid hydroperoxide glutathione peroxidase (PHGPx) is a key enzyme in the protection of biomembranes exposed to oxidative stress. We investigated the role of mitochondrial PHGPx in apoptosis using RBL2H3 cells that overexpressed mitochondrial PHGPx (M15 cells), cells that overexpressed non-mitochondrial PHGPx (L9 cells), and control cells (S1 cells). The morphological changes and fragmentation of DNA associated with apoptosis occurred within 15 h in S1 and L9 cells upon exposure of cells to 2-deoxyglucose (2DG). The release of cytochrome c from mitochondria was observed in S1 cells after 4 h and was followed by the activation of caspase-3 within 6 h. Overexpression of mitochondrial PHGPx prevented the release of cytochrome c, the activation of caspase-3, and apoptosis, but non-mitochondrial PHGPx lacked the ability to prevent the induction of apoptosis by 2DG. An ability to protect cells from 2DG-induced apoptosis was abolished when the PHGPx activity of M15 cells was inhibited by diethylmalate, indicating that the resistance of M15 cells to apoptosis was indeed due to the overexpression of PHGPx in the mitochondria. The expression of members of the Bcl-2 family of proteins, such as Bcl-2, Bcl-xL, Bax, and Bad, was unchanged by the overexpression of PHGPx in cells. The levels of hydroperoxides, including hydrogen and lipid peroxide, in mitochondria isolated from S1 and L9 cells were significantly increased after the exposure to 2DG for 2 h, while the level of hydroperoxide in mitochondria isolated from M15 cells was lower than that in S1 and L9 cells. M15 cells were also resistant to apoptosis induced by etoposide, staurosporine, UV irradiation, cycloheximide, and actinomycin D, but not to apoptosis induced by Fas-specific antibodies, which induces apoptosis via a pathway distinct from the pathway initiated by 2DG. Our results suggest that hydroperoxide, produced in mitochondria, is a major factor in apoptosis and that mitochondrial PHGPx might play a critical role as an anti-apoptotic agent in mitochondrial death pathways.     

RNA-Induced Interference with Animal Virus Infection

Some RNAs, including both single- and double-stranded RNAs, when incubated with chick embryo cell culture induce cellular resistance against viruses. Evidence was now obtained indicating that the induction of cellular resistance by RNA depends on the cellular metabolic activity, especially on the synthesis of cellular RNA and protein. Thus, inhibitors of RNA and protein synthesis, actinomycin D and cycloheximide, were found to inhibit the development of an antiviral state when added before, or during the relatively early period of, incubation of the cells with RNA. In the course of induction of cellular resistance, three stages may be distinguished, the priming stage, the developing stage, and the established resistant stage.     

Organic Acid Metabolism in Aluminum-Phosphate Utilizing Cells of Carrot (Daucus carota L.)

Carbohydrate metabolism in Al-phosphate utilizing cells of carrot[designated as IPG, Koyama et al. (1992) Plant Cell Physiol.33: 171], which grow normally in Al-phosphate medium accompaniedby citrate excretion, was investigated. The excretion of citratewas strongly related to the availability of sucrose in medium,indicating that citrate excretion was severely limited by sucrosein medium. The ratio of the amount of carbon in the excretedcitrate to the consumed sucrose, was significantly higher inIPG cells than in wild-type cells. When 50% of the sucrose inthe medium was consumed, the ratio was 0.6% for the IPG cellsand 0.2% the wild-type cells. Under these conditions, IPG cellsshowed altered citrate synthesis metabolism, which resultedin increased citrate production. Specific activity of mitochondrialcitrate synthase was higher in IPG cells than in wild-type cells,whereas the activity of cytosolic NADP-specific isocitrate dehydrogenasewas lower in IPG cells than in wild-type cells. (Received August 27, 1998; Accepted February 21, 1999)     

Meal feeding schedule and ventromedial hypothalamic lesions do not affect rhythm of pineal serotonin N-acetyltransferase activity in rats

Effects of meal feeding schedule and bilateral lesions of the ventromedial hypothalamus (VMH) on the circadian rhythm of pineal serotonin N-acetyltransferase (SNAT) activity were examined in rats, under LD (12:12) condition. Neither meal feeding nor VMH lesions affected the phase of the circadian rhythm of pineal SNAT activity, but the VMH lesions reduced the level. Meal feeding caused a shift of the phases of the daily rhythms of phosphoenolpyruvate carboxykinase and tyrosine aminotransferase activities in the liver. These findings suggest that the circadian rhythm of pineal SNAT activity is not entrained by the food intake, and that the VMH does not function as a master oscillator of the rhythm.     

Photoreduction of autooxidized albumin-heme hybrid in saline solution: revival of its O(2)-binding ability.

Recombinant human serum albumin (rHSA) incorporating 2-[8-[N-(2-methylimidazolyl)]octanoyloxymethyl]-5,10,15,20-tetrakis(alpha,alpha,alpha,alpha-o-pivalamido)phenylporphinatoiron(II)s (Fe(II)Ps) [rHSA-Fe(II)P] is a synthetic hemoprotein which can bind and release O(2) reversibly under physiological conditions (saline solution [NaCl]: 150 mM, pH 7.3) as do hemoglobin and myoglobin. However, the central ferrous ions of Fe(II)Ps are slowly oxidized to O(2)-inactive ferric forms. Based on the UV-vis. absorption spectroscopy, the majority of the autooxidized Fe(III)Ps in albumin are determined to be six-coordinate high-spin complexes with a proximal imidazole and a chloride anion, which show ligand-to-metal charge transfer (LMCT) absorption at 330 nm. Interestingly, photoirradiation of this LMCT band under an argon atmosphere led to reduction of the central ferric iron of Fe(III)P, allowing the revival of the O(2)-binding ability. The ratio of the photoreduction reached a maximum of 83%, which is probably due to the partial dissociation of the axial imidazole. The same photoirradiation under a CO atmosphere provides the corresponding carbonyl rHSA-Fe(II)P. Laser flash photolysis experiments revealed that the reduction was completed within 100 ns. The quantum yields (Phi) of these photoreductions were approximately 0.01.     

Distribution and evolution of C4 syndrome inRhynchospora (Rhynchosporeae-Cyperaceae)

In order to elucidate the evolution of C₄ syndrome, the taxonomic relationships, leaf anatomy, and ecological and global distribution of C₃ and C₄ species in the genusRhynchospora were investigated. The anatomical observation for 181 species revealed that 26 C₄ species occurred within theCapitatae group of the subgenusHaplostyleae, a natural group showing highly advanced morphological characteristics, together with several C₃ species. In spite of there being rather few C₄ species, they possessed two kinds of Kranz anatomical structure differing from each other in the location of Kranz cells. Some C₃ species ofCapitatae showed radial arrangement in mesophyll cells surrounding vascular bundles, which is distinguished from typical non-Kranz anatomy. The C₄ species extended their ecological ranges from wet habitats to dry savanna grasslands, while the C₃ species showed the best development in wet habitats. The C₃ species were widespread from tropical to temperate regions with partial range extension into subarctic regions of both hemispheres, showing conspicuously high concentration of species in the New World, but being absent from arid climatic regions. The C₄ species were distributed mostly in tropics and subtropics, showing two separate distributional centers in South and Central America and in Tropical Asia and Australia. The range of C₄ species was nearly completely included in the C₃ range. In conclusion, it seems that inRhynchospora the C₄ syndrome evolved relatively recently, and arose in at least two separate phylogenetic trends in the tropics and the subtropics, more probably in the Neotropics.