Novel Technique for Measuring Tissue Firmness within Tomato (Lycopersicon esculentum Mill.) Fruit

Developmental changes of tomato (Lycopersicon esculentum) fruit tissues during maturation were analyzed by a physically defined method (stress-relaxation analysis). The tip of a conical probe connected to a load sensor was positioned on the cut surface of a sliced tomato fruit, and the decay of the imposed stress was monitored. Stress-relaxation data thus obtained were used for the calculation of three stress-relaxation parameters. Different zones within tomato fruit harvested at six different ripening stages were analyzed. One of the stress-relaxation parameters, minimum stress-relaxation time (T₀), decreased as the fruits matured. The decrease in T₀ was first found in the core of the carpel junction within the endopericarp at the blossom end during the breaker stage. The decrease in T₀ progressed from the blossom end, through the equatorial region and finally throughout the shoulder, as the fruit matured. In mature green fruit, T₀ values within the placenta and the proximal carpel junction were lower than those by other parts of the fruit. For all measurements the maximum stress-relaxation time was not substantially changed during maturation, nor were their changes observed in different regions of the fruit. The observed relaxation rate was therefore correlated with softening. The results indicate that fruit softening may be physically associated with the stress-relaxation parameter, T₀, and the extent of softening is a function of position within the fruit. Decreases in T₀ value appear to be correlated with the reported regional variation in the appearance of polygalacturonase.     

Zeaxanthin Formation and Energy-Dependent Fluorescence Quenching in Pea Chloroplasts under Artificially Mediated Linear and Cyclic Electron Transport

Artificially mediated linear (methylviologen) and cyclic (phenazine methosulfate) electron transport induced zeaxanthin-dependent and independent (constitutive) nonphotochemical quenching in osmotically shocked chloroplasts of pea (Pisum sativum L. cv Oregon). Nonphotochemical quenching was quantitated as Stern-Volmer quenching (SV_N) calculated as (F_m/F′_m)-1 where F_m is the fluorescence intensity with all PSII reaction centers closed in a nonenergized, dark-adapted state and F′_m is the fluorescence intensity with all PSII reaction centers closed in an energized state. Reversal of quenching by nigericin and electron-transport inhibitors showed that both quenching types were energy-dependent SV_N. Under light-induced saturating ΔpH, constitutive-SV_N reached steady-state in about 1 minute whereas zeaxanthin-SV_N continued to develop for several minutes in parallel with the slow kinetics of violaxanthin deepoxidation. SV_N above the constitutive level and relative zeaxanthin concentration showed high linear correlations at steady-state and during induction. Furthermore, F_o quenching, also treated as Stern-Volmer quenching (SV_O) and calculated as (F_o/F′_o)-1, showed high correlation with zeaxanthin and consequently with SV_N (F_o and F′_o are fluorescence intensities with all PSII reaction centers in nonenergized and energized states, respectively). These results support the view that zeaxanthin increases SV_N above the constitutive level in a concentration-dependent manner and that zeaxanthin-dependent SV_N occurs in the pigment bed. Preforming zeaxanthin increased the rate and extent of SV_N, indicating that slow events other than the amount of zeaxanthin also affect final zeaxanthin-SV_N expression. The redox state of the primary electron acceptor of photosystem II did not appear to determine SV_N. Antimycin, when added while chloroplasts were in a dark-adapted or nonenergized state, inhibited both zeaxanthin-SV_N and constitutive-SV_N induced by linear and cyclic electron transport. These similarities, including possible constitutive F_o quenching, suggest that zeaxanthin-dependent and constitutive SV_N are mechanistically related.     

Changes of fluorescence lifetime and rotational correlation time of bovine serum albumin labeled with 1-dimethylaminonaphthalene-5-sulfonyl chloride in guanidine and thermal denaturations

The nanosecond fluorescence depolarization method was applied to measure the fluorescence lifetime () and the rotational correlation time () of bovine serum albumin (BSA) labeled with 1-dimethylaminonaphthalene-5-sulfonyl chloride (dansyl-Cl). Changes of and of dansyl BSA in the guanidine denaturation and in the thermal denaturation were examined. In parallel, the secondary structural change of dansyl BSA was followed by circular dichroism measurements. The magnitude of was almost unchanged between 1 and 2 M guanidine, where the secondary structure of the protein was predominantly disrupted; whereas that of began to increase before the disruption of secondary structure in the guanidine denaturation. In the thermal denaturation, in contrast, changes of both and occurred in a temperature range where the secondary structure was predominantly disrupted. The volume of equivalent sphere (V _e ) and the axial ratio () for the BSA were 3.6–3.8×10^–19 cm³ and 3.6 at 2M guanidine as against 2.1×10^–19 cm³ and 2.2 in the absence of guanidine (25°C), respectively. The magnitudes ofV _e and were 4.9×10^–19 cm³ and 4.5 at 65°C, respectively. Although the secondary structural change of dansyl BSA was irreversible in the thermal denaturation,V _e and were reversible.     

Effect of guanidinoethanesulfonic acid on brain monoamines in the mouse

Guanidinoethanesulfonic acid (GES) is known to induce convulsive seizures when administered intracisternally to rabbits and cats. The effects of GES on behavior, electroencephalographic recording and brain monoamine levels were examined in mice. When GES (900 nmol) was intraventricularly injected into mice, focal clonic movements of the face, vibrissae and ears together with twitching of the limbs were observed 0.5–1 min after the injection. Hypersensitivity was observed up to 7 min after the injection, after which the mice behaved normally. GES also induced sporadic spike discharges on electrocorticogram. The levels of 5-hydroxytryptamine (5-HT) of the GES-injected mice were lower than those of the saline-injected mice in the hippocampus, diencephalon, pons-medulla oblongata and cerebellum 5 min after the injection. No changes in the norepinephrine or dopamine levels were found after the GES injection. The level of 5-hydroxyindoleacetic acid increased in the striatum and cerebellum 5 min after the GES injection. These results suggest that GES-induced convulsive activities enhance the serotonergic neuroactivity in order to suppress the convulsions.     

Incorporation of fatty acids by Streptococcus mutans

In a series of investigations into the cariogenicity of Streptococcus mutans, we studied the incorporation of exogenous fatty acids with reference to glucosyltransferase secretion and membrane fatty acid changes. When cells were grown with different fatty acids, both saturated and unsaturated fatty acids were readily incorporated into the membrane lipids and were biotransformed and elongated preferentially to the longer 16- and 18-carbon-chain fatty acids. This incorporation and chain-elongation led to significant changes in fatty acids composition. By adding fatty acids to the medium, it was possible to appropriately modify the degree of unsaturation and the relative ratio between specific fatty acids in the membrane lipids of S. mutans.     

Homologous Recombination Involving a Large Heterology in Escherichia Coli

Recombination between two different deletion alleles of a gene (neo) for neomycin and kanamycin resistance was studied in an Escherichia coli sbcA- recB-C- strain. The two homologous regions were in an inverted orientation on the same plasmid molecule. Kanamycin-resistant plasmids were selected and analyzed. The rate of recombination to form kanamycin-resistant plasmids was decreased by mutations in the recE, recF and recJ genes, but was not decreased by a mutation in the recA gene. It was found that these plasmids often possessed one wild-type kanamycin-resistant allele (neo+) while the other neo allele was still in its original (deletion) form. Among kanamycin-resistant plasmids with one wild-type and one parental allele it was often found that the region between the inverted repeats had been flipped (turned around) with respect to sites outside the inverted repeats. These results were interpreted as follows. Gene conversion, analogous to gene conversion in eukaryotic meiosis, is responsible for a unidirectional transfer of information from one neo deletion allele to the other. The flipping of the region between the inverted repeats is interpreted as analogous to the crossing over associated with gene conversion in eukaryotic meiosis. In contrast with a rec+ strain, these products cannot be explained by two rounds of reciprocal crossing over involving a dimeric form as an intermediate. In the accompanying paper we present evidence that gene conversion by double-strand gap repair takes place in the same E. coli strain.     

The effects of ovarian hormones on glucose and fatty acid oxidation during exercise in female ovariectomized rats

The effects of ovarian hormones on glucose and fatty acid oxidation during exercise were investigated in adult female ovariectomized rats. Rats subdivided into 3 groups received intraperitoneal injections of hormones or sesame oil for 8 days. Estrogen (E) treated rats received 17-beta estradiol in daily doses of 2 micrograms. Estrogen and progesterone treated rats (EP) received 17-beta estradiol in daily doses of 2 micrograms and 2 mg, respectively. Control rats (S) received sesame oil alone. After an overnight fast, rats ran at the speed of 25 m.min-1 for 60 min. [U-14C]glucose or [1-14C]palmitate was injected into rats at 5 min of exercise and before 10 min of exercise, respectively. Expired 14CO2 was collected using bottomless chamber on a treadmill belt. No significant differences were found in mean blood glucose, lactate and plasma free fatty acid concentrations after the exercise. Until the end of the exercise 34.7 +/- 2.6 (E, n = 5), 40.8 +/- 2.9 (EP, n = 5) and 43.7 +/- 3.5% (S, n = 6) (mean +/- SE) of 14C which was injected as 14C-glucose was recovered as 14CO2. During 60 min of the exercise 27.5 +/- 1.0 (E, n = 7), 19.8 +/- 2.7 (EP, n = 6) and 25.0 +/- 1.9% (S, n = 6) of 14C which was injected as 14C-palmitate was recovered as 14CO2. A significant difference was found in this rate between E and EP (P less than 0.05). It was concluded that estrogen treatment stimulated fatty acid oxidation compared with the estrogen plus progesterone treatment and tended to inhibit glucose oxidation during prolonged exercise.     

Gene structure of human thrombomodulin, a cofactor for thrombin-catalyzed activation of protein C

The gene coding for human thrombomodulin, a thrombin receptor on endothelial cells and a cofactor for the activation of anticoagulant protein C zymogen, was isolated from a human genomic library by employing human thrombomodulin cDNA as a probe. The nucleotide sequences of the gene and the adjacent 5' and 3' flanking regions were then determined. The nucleotide sequence of this gene with approximately 3.7 kilobase pairs was identical to that of the cDNA, indicating that the gene for human thrombomodulin is free of introns. Hybridization data showed that there is only a single thrombomodulin gene in the human genome.