Effect of Temperature, Light, Nutrients and Dehardening on Abscisic Acid Induced Cold Hardiness in Bromus inermis Leyss Suspension Cultured Cells

The effects of culture conditions on abscisic acid (ABA)-inducedfreezing tolerance were determined in smooth bromegrass Bromusinermis Leyss cv. Manchar) cell suspension cultures. Bromegrasscultures initiated with 2 g fr wt of cells achieved maximumfreezing tolerances (greater than –32?C) at 25 to 30?Cin the presence of 75 to 100 µM ABA. High levels of freezingtolerance induced by ABA were correlated with high growth ratesat 25 and 30?C. In control cells, incubation at 10?C inducedoptimum levels of hardiness with minimal growth. Prolonged exposure(6 weeks) of cells to 3?C, with or without ABA, increased freezingtolerance only by several degrees. Exogenous ABA concentrationsgreater than 100 µM were not inhibitory to growth. Repeatedexposure to ABA, however, retarded growth and made the cellstolerant to temperatures below –40?C. Removal of ABA fromthe medium resulted in dehardening of the cells both at 25 and3?C. Nitrogen had a marginal effect on ABA-induced hardeningat 25?C, but inhibited age-dependent hardening of control cellcultures. Light had no effect on the freezing tolerance of culturedcells. Addition of 10% sucrose, 30 min prior to freezing, tobromegrass cells treated with ABA for 4 days increased freezingtolerance more than 15?C. These observations are discussed inrelation to the contrasting behaviour of the low temperatureand photoperiod dependent cold acclimation of plants (Received July 14, 1989; Accepted October 23, 1989)     

Coevolution and Hierarchical Interactions of Tomato mosaic virus and the Resistance Gene Tm-1

During antagonistic coevolution between viruses and their hosts, viruses have a major advantage by evolving more rapidly. Nevertheless, viruses and their hosts coexist and have coevolved, although the processes remain largely unknown. We previously identified Tm-1 that confers resistance to Tomato mosaic virus (ToMV), and revealed that it encodes a protein that binds ToMV replication proteins and inhibits RNA replication. Tm-1 was introgressed from a wild tomato species Solanum habrochaites into the cultivated tomato species Solanum lycopersicum. In this study, we analyzed Tm-1 alleles in S. habrochaites. Although most part of this gene was under purifying selection, a cluster of nonsynonymous substitutions in a small region important for inhibitory activity was identified, suggesting that the region is under positive selection. We then examined the resistance of S. habrochaites plants to ToMV. Approximately 60% of 149 individuals from 24 accessions were resistant to ToMV, while the others accumulated detectable levels of coat protein after inoculation. Unexpectedly, many S. habrochaites plants were observed in which even multiplication of the Tm-1-resistance-breaking ToMV mutant LT1 was inhibited. An amino acid change in the positively selected region of the Tm-1 protein was responsible for the inhibition of LT1 multiplication. This amino acid change allowed Tm-1 to bind LT1 replication proteins without losing the ability to bind replication proteins of wild-type ToMV. The antiviral spectra and biochemical properties suggest that Tm-1 has evolved by changing the strengths of its inhibitory activity rather than diversifying the recognition spectra. In the LT1-resistant S. habrochaites plants inoculated with LT1, mutant viruses emerged whose multiplication was not inhibited by the Tm-1 allele that confers resistance to LT1. However, the resistance-breaking mutants were less competitive than the parental strains in the absence of Tm-1. Based on these results, we discuss possible coevolutionary processes of ToMV and Tm-1.     

Determinants of Slow Walking Speed in Ambulatory Patients Undergoing Maintenance Hemodialysis

Walking ability is significantly lower in hemodialysis patients compared to healthy people. Decreased walking ability characterized by slow walking speed is associated with adverse clinical events, but determinants of decreased walking speed in hemodialysis patients are unknown. The purpose of this study was to identify factors associated with slow walking speed in ambulatory hemodialysis patients. Subjects were 122 outpatients (64 men, 58 women; mean age, 68 years) undergoing hemodialysis. Clinical characteristics including comorbidities, motor function (strength, flexibility, and balance), and maximum walking speed (MWS) were measured and compared across sex-specific tertiles of MWS. Univariate and multivariate logistic regression analyses were performed to examine whether clinical characteristics and motor function could discriminate between the lowest, middle, and highest tertiles of MWS. Significant and common factors that discriminated the lowest and highest tertiles of MWS from other categories were presence of cardiac disease (lowest: odds ratio [OR] = 3.33, 95% confidence interval [CI] = 1.26–8.83, P<0.05; highest: OR = 2.84, 95% CI = 1.18–6.84, P<0.05), leg strength (OR = 0.62, 95% CI = 0.40–0.95, P<0.05; OR = 0.57, 95% CI = 0.39–0.82, P<0.01), and standing balance (OR = 0.76, 95% CI = 0.63–0.92, P<0.01; OR = 0.81, 95% CI = 0.68–0.97, P<0.05). History of fracture (OR = 3.35, 95% CI = 1.08–10.38; P<0.05) was a significant factor only in the lowest tertile. Cardiac disease, history of fracture, decreased leg strength, and poor standing balance were independently associated with slow walking speed in ambulatory hemodialysis patients. These findings provide useful data for planning effective therapeutic regimens to prevent decreases in walking ability in ambulatory hemodialysis patients.     

Analysis of water in Autonomous Biological Systems (ABS) samples.

Several soluble components, peptidase and amino acids, and carbon isotopic ratio in the water retrieved from flight experiments of Autonomous Biological Systems (ABS) as well as ground control samples are analyzed to interpret the condition, dynamics, material balance of the ABS ecosystems. Organic carbons in flight samples were found to be more abundant compared with the control ones, which suggested the uniform ecosystems in low gravity might easily dissolve more soluble components. The Mir-1997 flight sample showed higher C/N ratio probably because of the dissolution of carbon-rich plant materials.     

Turnover of the aggregates and cross-linked products of the D1 protein generated by acceptor-side photoinhibition of photosystem II.

It is known that the reaction-center binding protein D1 in photosystem (PS) II is degraded significantly during photoinhibition. The D1 protein also cross-links covalently or aggregates non-covalently with the nearby polypeptides in PS II complexes by illumination. In the present study, we detected the adducts between the D1 protein and the other reaction-center binding protein D2 (D1/D2), the alpha-subunit of cyt b(559) (D1/cyt b(559)), and the antenna chlorophyll-binding protein CP43 (D1/CP43) by SDS/urea-polyacrylamide gel electrophoresis and Western blotting with specific antibodies. The adducts were observed by weak and strong illumination (light intensity: 50-5000 microE m(-2) s(-1)) of PS II membranes, thylakoids and intact chloroplasts from spinach, under aerobic conditions. These results indicate that the cross-linking or aggregation of the D1 protein is a general phenomenon which occurs in vivo as well as in vitro with photodamaged D1 proteins. We found that the formation of the D1/D2, D1/cyt b(559) and D1/CP43 adducts is differently dependent on the light intensity; the D1/D2 heterodimers and D1/cyt b(559) were formed even by illumination with weak light, whereas generation of the D1/CP43 aggregates required strong illumination. We also detected that these D1 adducts were efficiently removed by the addition of stromal components, which may contain proteases, molecular chaperones and the associated proteins. By two-dimensional SDS/urea-polyacrylamide gel electrophoresis, we found that several stromal proteins, including a 15-kDa protein are effective in removing the D1/CP43 aggregates, and that their activity is resistant to SDS.     

Chemical, physicochemical and spectrophotometric properties of crystalline chlorophyll-protein complexes from Lepidium virginicum L

Two kinds of water-soluble chlorophyll-protein complexes were prepared from leaves of Lepidium virginicum L., one (CP661) from the plant cultivated in a green house from seeds collected near Mono Lake, CA, and the other (CP-663) from a plant collected at Narashino, Chiba, Japan, by ammonium sulfate fractionation followed by column chromatography on DEAE-cellulose and Sephacryl S-200. The chlorophyll . proteins were further purified by crystallization. CP661 has absorption peaks at 661, 468, 439, 419, 380, 339 and 272 nm. CP663 had absorption peaks at 663, 469, 438, 419, 379, 338 and 272 nm. Estimated molecular weights were 78 000 for CP661 and 80 000 for CP663 by gel filtration chromatography and 83 000 for CP661 and 107 000 for CP663 by an equilibrium sedimentation method. 1 mol chlorophyll . protein contained 4 mol chlorophyll a and b with ratios of 1.0 in CP661 and 1.6 to 1.9 in CP663, but no carotenoids. These characters are different from those of chlorophyll-protein complexes which are prepared from the thylakoid membranes of chloroplasts with detergents.     

Seasonal Changes in Plasma Membranes and Mitochondria Isolated From Jerusalem Artichoke Tubers. Possible Relationship to Cold Hardiness

Plasma membranes and mitochondria were isolated from Jerusalemartichoke tubers during cold acclimation from September to December.The protein and lipid contents of the membranes were analyzedwith reference to physiological properties of the tubers, especiallycold hardiness. As cold hardiness increased from autumn to winter,the content of phospholipids and sterols on a mg protein basisincreased by 20–30% in plasma membranes, but little changewas observed in mitochondria. Minor changes were observed inthe fatty acid composition of phospholipids either in plasmamembranes or mitochondria. Membrane fluidity, assessed by fluorescentpolarization of 1,6-diphenyl-1,3,5-hexatriene, was found tobe relatively constant in both membranes during the season.One dimensional SDS-polyacrylamide gel electrophoresis revealedseasonal changes in proteins and glycoproteins in plasma membranes,but not in mitochondrial membranes. Plasma membrane ATPase increasedin specific activity from September to December, which was morenoticeable at higher assay temperatures. However, irrespectiveof the season, the plasma membrane ATPase had an inflectionon the slope of the Arrhenius plot around 15C. These resultssuggest that plasma membranes, in contrast to mitochondria,undergo several molecular changes from autumn to winter, whichmay be related to cold acclimation of the tubers. ¹ Contribution No. 2668 from the Institute of Low TemperatureScience. ² Present address: Crop Development Centre, University of Saskatchewan,Saskatoon, Canada S7N 0W0.     

Effects of epinephrine and oxytocin on the release of prostaglandin F from the rat uterus in vitro

Isolated uteri from rats with regular 4-day cycles were incubated in Krebs-Ringer bicarbonate buffer and the release of PGF into the medium was measured by radioimmunoassay after extraction of the incubation medium with ethyl acetate at pH 3.0-3.5. PGF was produced from endogenous precursors and accumulated in equal amounts in the medium during two successive 60 min periods on each day of cycle, but the magnitude of the production varied significantly during the cycle, being greatest in estrus. Oxytocin in doses up to 500 mU/ml had no effect on PGF accumulation in the incubation period at any stage of the cycle, while epinephrine (10(-3)) greatly stimulated PGF release from the estrous uterus but had no effect on PGF release from the diestrous uterus. Phentolamine, an alpha-blocking agent, had no effect on the epinephrine-induced release of PGF, while propranolol, a beta-blocking agent, not only prevented in increase in PGF production induced by epinephrine but also reduced the basal release of PGF by the estrous uterus. Since oxytocin contracts and epinephrine relaxes the nonpregnant rat uterus both in vivo and in vitro, it is unlikely that the effects of these two compounds on uterine contractility are mediated by the release of PGF2alpha.