Accumulation of Species-Specific Amino Acid Replacements That Cause Loss of Particular Protein Functions in Buchnera, an Endocellular Bacterial Symbiont

Endosymbiotic bacteria live in animal cells and are transmitted vertically at the time of the host's reproduction. In view of their small and asexual populations with infrequent chances of recombination, these endocellular bacteria are expected to accumulate mildly deleterious mutations. Previous studies showed that the DNA sequences of these bacteria evolved faster than those of free-living bacteria. In this study, we compared all the ORFs of Buchnera, an endocellular bacterial symbiont of aphids, with those of 34 other prokaryotic organisms and estimated the effect of the accelerated evolution of Buchnera on the functions of its proteins. It was revealed that Buchnera proteins contain many mutations at the sites where sequences are conserved in their orthologues in many other organisms. In addition, amino acid replacements at the conserved sites are mostly changes to physicochemically different amino acids. These results suggest that functions and conformations of Buchnera proteins have been seriously impaired or strongly modified. Indeed, extensive loss of functional motifs was observed in some Buchnera proteins. In many Buchnera proteins mutations were not detected evenly throughout each molecule but tended to accumulate in some functional units, possibly leading to loss of specific functions. As Buchnera has an unusual and limited gene repertory, it is conceivable that the manner of interactions among its proteins has been changed, and thus, functional constraints over their amino acid residues have also been changed during evolution. This may account for the loss of some functional units only in the Buchnera proteins. We obtained evidence that amino acid replacements in Buchnera were not always deleterious, but neutral or, in some cases, even positively selected. Received: 14 December 2000 / Accepted: 12 March 2001     

Prevention of adriamycin-induced interphase death by 3-aminobenzamide and nicotinamide in a human promyelocytic leukemia cell line

Adriamycin caused significant interphase death in HL-60 cells during six hours of incubation, which was abolished by the poly(ADP-ribose) polymerase inhibitors, 3-aminobenzamide or nicotinamide. Neither agent changed adriamycin uptake by HL-60 cells. Although 3-aminobenzamide did not alter the number of DNA strand breaks caused by adriamycin, it prevented adriamycin-induced depletion of intracellular NAD+ and ATP, and maintained energy charge. These findings suggest that the activation of poly(ADP-ribose) synthesis plays an important role in the adriamycin-induced interphase death of proliferating HL-60 cells.     

Physical Basis of Flower-opening in Pharbitis nil

Flower buds of Pharbitis nil cut from plants growing in thefield open rapidly when subjected to darkness (20–25°C)or low temperature (20°C) in light. Petals of the buds arethe sites of photo- and thermo-perception; flower-opening iscaused mainly by the epinasty of petal midribs. ¹Dedicated to Professor Dr. Erwin Bunning on the occasion ofhis 75th birthday. (Received October 23, 1980; Accepted December 15, 1980)     

Flower-Inducing Activity of Benzoic Acid Derivatives for Lemna minor

Lemna minor, strain M601, scarcely flowers in full or 1/10 strengthM medium supplemented with 1% sucrose either in long-day orshort-day conditions, but easily flowers when some benzoic acidderivatives are added to 1/10 strength M-sucrose medium. Thepresence of cytokinin in the medium improves the activity ofbenzoic acid but not significantly that of salicylic acid. Thestructure-activity relationships for the flower-inducing activityof substituted benzoic acid derivatives in the presence of cytokininwere analyzed quantitatively in terms of physicochemical substituentparameters. In the presence of cytokinin, the flower-inducingactivity is determined by electronic and steric effects of thesubstituent. The higher the electron withdrawing ability andthe lower the bulkiness of the substituent, the higher is theactivity. In the absence of cytokinin, the activity is enhancedby higher electron withdrawing ability of the substituent, butthe steric effect is insignificant. In both conditions, thehydroxyl group in the ortho position exhibits an additionaleffect to enhance activity, and the molecular form responsiblefor the activity seems to be undissociated neutral species.The structure-activity relationship in L. minor M601 is basicallysimilar to that for L. paucicostata 151. (Received February 2, 1983; Accepted May 2, 1983)     

Determination of the cross-points of rat liver peroxisomes,peroxisomal core and the core components by cross-partition

The cross-points of rat liver peroxisomes, peroxisomal core and the core components were determined by means of cross-partition in two phase systems. The partitions were carried out in the systems containing 6% (w/w) Dextran T 500 and 6% (w/w) polyethyleneglycol 4000 in sodium salts. The same crosspoint, pH 5.6, was obtained in peroxisomal marker enzymes in light mitochondrial fraction of liver homogenate, such as catalase, d-amino acid oxidase and urate oxidase. The cross-point as determined by cross-partition of purified peroxisomal core was 6.7. The cross-points of urate oxidase and framework protein fractions obtained by alkali treatment on the purified core were 7.8 and 4.2, respectively, and the ratio of the proteins of urate oxidase to framework protein was 2:1. The theoretical value of cross-point of the core calculated from the relationship between the cross-point and protein ratio of each component of the core coincided with the experimental value obtained by this method.     

Analysis of photosystem II using particle II preparation III. Roles of cytochrome b559 with different redox potentials and plastocyanin in the photosynthetic electron transport system

1. Using the P-II preparation (photosystem II preparation),the functional sites for Cyt-b₅₅₉ with different redox potentialsand for plastocyanin in the non-cyclic electron transport chainof chloroplasts were investigated. 2. In the absence of plastocyanin, the P-II preparation showedno light-induced absorption changes in Cyt-b₅₅₉. However, uponthe addition of a sufficient amount of plastocyanin, remarkablephotooxidation of this cytochrome was observed at room temperature. 3. Parallel measurements of the light-induced absorption changesin both Cyt-b₅₅₉ and plastocyanin revealed a close relationshipbetween them, and indicated that Cyt-b₅₅₉ and plastocyanin arelocated in series on the main path of the electron transportchain involving two photoreactions in PS-II. 4. Difference spectra and action spectra for the light-inducedabsorption changes in Cyt-b₅₅₉ and plastocyanin offered additionalevidence in support of the above conclusion. 5. Analysis of the relationship between the activity of ferricyanideHill reaction and the contents of Cyt-b₅₅₉ of different redoxpotentials showed that both forms of Cyt-b₅₅₉ (i.e. high- andlow-potential forms) play important roles under physiologicalconditions, being on the main pathway of the electron transportchain connecting PS-II and PS-I. 6. To these findings it is possible to give reasonable explanationsbased on our scheme presented previously, which suggested thatPS-II contains not one but two different photoreactions. Thisscheme is much more elaborately supported by this study whichshows the functional sites for Cyt-b₅₅₉ with two different potentials. ¹This work has been supported by a Grant from the Ministry ofEducation (Grant No. 844004), which we gratefully acknowledge.Part of this report has been presented at the Gordon ResearchConference on Regulatory Mechanisms in Photosynthesis, at Tilton,N.H., U.S.A., Aug. 13–17, 1973. (Received June 20, 1974; )     

Extremely low frequency magnetic fields suppress the reduction of germination rate of Arabidopsis thaliana seeds kept in saturated humidity.

The effects of an extremely low frequency magnetic field (ELFMF) on the germination of plant seeds were examined. The decrease in the germination activity of the seeds of Arabidopsis thaliana WS kept in saturated humidity and high temperature (37 degrees C) was suppressed by the exposure to a 400 mT ELFMF.     

C6 glioma cells produce basic fibroblast growth factor that can stimulate their own proliferation

With purified preparations of basic fibroblast growth factor (bFGF), we studied the effect of its growth-promoting activity on C6 glioma cells. We also examined with its antibody whether the cultured glioma cells could produce it. It was shown that bFGF stimulated the DNA synthesis and proliferation of C6 glioma cells in serum-free medium, and that the activity was potentiated by heparin, the bFGF concentrations for half-maximal stimulation being 0.2 and 5 ng/ml in the presence and absence of heparin, respectively. This effect of heparin was dose-dependent and was half-maximal at 0.5 microgram/ml. Next, we raised the antiserum against bFGF and detected a single immunoreactive band from extracts of C6 glioma cells by immunoblot analysis. The immunoreactive substance was partially purified on a heparin-Sepharose column and was shown to stimulate the DNA synthesis of C6 glioma cells. On the basis of its immunoreactivity, molecular weight, affinity for heparin, and growth-promoting activity, this substance was identified as bFGF. The content of bFGF in the cells was elevated as the cell density increased, but no immunoreactivity was detected in the conditioned medium of the cells. These results suggest that C6 glioma cells produce and store bFGF which is potent in stimulating their own growth.