PLD activation in Chinese hamster ovary (CHO) cells transfected with PGF2α receptor cDNA

Pertussis toxin-insensitive GTP-binding protein was observed to be involved in prostaglandin F2α(PGF2α)-induced phosphoinositide metabolism in Chinese hamster ovary (CHO) cells transfected with PGF2α receptor cDNA (CHO-PGF2α·R cells) (Ito, S. et al. Biochem. Biophys. Res. Commun. 200: 756, 1994). In the present study, we investigated PGF2α-induced PLD activation in CHO-PGF2α·R cells. PLD activation was examined by measuring the production of [³H]phosphatidylbutanol ([³H]PBut), a specific product of the PLD-catalyzed transphosphatidylation reaction. PGF2α-induced [³H]PBut formation was concentration-dependent with the maximal level obtained at 1 μM PGF2α. The maximal [³H]PBut formation was observed at 2 min after addition of PGF2α. Depletion of extracellular Ca²⁺ with EGTA suppressed PGF2α-induced PLD activation by 50%. PKC inhibitors Ro31–8425 and calphostin C inhibited PGF2α-induced [³H]PBut formation by 50%. PTK inhibitors genistein and herbimycin A failed to inhibit PGF2α-induced PLD activation. A combination of maximal effective concentrations of PGF2α (1 μM) and PMA (100 nM) enhanced PLD activation in an additive manner. Pretreatment of the cells with PMA for 2 h down-regulated PKCα and decreased PGF2α-induced PLD activation. These results suggest that PLD activation by PGF2α is mediated by both PKC-dependent and -independent pathways and that PKCα is involved in the former pathway.     

Photoactive Photosystem I Particles with a Molar Ratio of Chlorophyll a to P700 of 9

Lyophilized photosystem I particles from spinach were treatedwith diethyl ether that contained various organic solvents withdifferent dielectric constants. More pigments were extractedas the dielectric constant of the solvent added to ether increased.The reaction-center chlorophylldimer, P700, was more resistantto extraction than the rest of the chlorophyll. Particles thatcontained only 6 chlorophylls in addition to P700 and the primaryelectron acceptor (A₀), in a single reaction-center unit, wereprepared by extraction with a mixture of ether and acetaldehyde.A distinct shoulder at 695 nm due to P700 or at 686 nm due toP700⁺ was observed in the absorption spectra of the reducedor oxidized particles, respectively, even at room temperature.No secondary acceptor phylloquinone remained in the particles.Stable charge separation was restored upon the addition of 2-amino-anthraquinone,even though the particles had the lowest molar ratio of chlorophyllto P700 of any reported particles. (Received February 20, 1995; Accepted May 8, 1995)     

Fluctuation of body temperature and cuckoo brood parasitism

Body temperatures of 11 bird species, including cuckoos, were measured in an artificial meteorological room. Ratios of change in body temperature to that in air temperature were thereby obtained for each species. Cuckoos demonstrate a remarkably high value, indicating a particularly low ability to regulate body temperature. Viewed in this light, the cuckoo's parasitic behavior is very likely an adaptation to overcome a physiological disadvantage. This in turn might be expected to reinforce delay in evolution of temperature homeostasis.     

Fluorometric measurement of tin-protoporphyrin in biological samples

Sn-protoporphyrin is a potent competitive inhibitor of heme oxygenase, can suppress neonatal and other forms of hyperbilirubinemia in laboratory animals, and represents a potential new approach to the treatment of neonatal jaundice in humans. In order to study the disposition of Sn-protoporphyrin in vivo we have developed a sensitive fluorometric method for the quantitation of this metalloporphyrin in biological samples. The method is sensitive to concentrations as low as 0.01 nmol/ml, and is specific for Sn-protoporphyrin even in the presence of other porphyrins such as protoporphyrin.     

Purification and properties of aldehyde dehydrogenase from Saccharomyces cerevisiae.

A procedure for the purification of aldehyde dehydrogenase from bakers' yeast (Saccharomyces cerevisiae) is reported. Treatment with acid, heat and organic solvents was avoided and chromatographic and filtration techniques in the presence of phenylmethylsulfonylfluoride were mainly used. An affinity chromatography step using the reactive dye Cibacron blue F3G-A, which was covalently bound to Sepharose 4B, was found to be essential. The enzyme was bound to and then released from the dye. The purified enzyme was shown to be homogeneous by gel filtration, disc electrophoresis and SDS electrophoresis. The molecular weight of the purified enzyme determined by gel filtration was 170,000, which agreed with that of the enzyme in the crude extract. The enzyme was composed of subunits of a molecular weight of 57,000. The specific activity of the enzyme was 20 units per mg of protein under the standard assay conditions. The substrate specificity, the relative maximal velocity, the michaelis constants, the pH optimum, the stability and the activation energy of the enzyme are reported.     

Heme transport from rat liver mitochondria to the microsomes invitro

When rat liver mitochondria labeled with [⁵⁹Fe]heme were incubated with microsomes in the presence of cytosol, about 16 % of the heme in mitochondria was transported to microsomes during a 1 hr-incubation period. In the absence of cytosol, little heme was transported. DEAE-Sephadex column chromatography of the cytosol partially purified by pH 5.1 treatment and ammonium sulfate precipitation (45–65%) revealed that there were at least two proteins with a releasing activity from mitochondria via heme transport.     

Permutation of the DNA in small-headed virions of coliphage P1

Summary Use of restriction endonucleases BglII, EcoRI, BamHI, and HindIII, has established that in small-headed (PIS) virions of coliphage P1, as a population, the entire genome found in big-headed (P1B) virions is represented. In addition, the origin and direction of DNA packaging are identical in P1S and P1B virions.