Accumulation of pyrophosphate correlates with the increased level of nucleoside triphosphates in Escherichia coli

We measured the concentration of nucleoside triphosphates and inorganic pyrophosphate in Escherichia coli in conditions where nucleotide synthesis or nucleic acid synthesis was inhibited. The inhibitors that brought about an accumulation of some of the four ribonucleoside triphosphates also increased the pyrophosphate level. In a pyrimidine auxotrophic strain uracil starvation led to simultaneous accumulation of ATP and pyrophosphate, and they both rapidly returned to normal level when starvation was relieved. These results indicate the possible involvement of pyrophosphate in the reactions leading to the accumulation of nucleoside triphosphates.     

The effect of different night conditions on the CO2 fixation in a lichen Xanthoria parietina

CO₂ fixation was studied in a lichen, Xanthoria parietina, kept in continuous light, and with cyclic changes in light intensity, dark period or temperature. The diurnal and seasonal courses of CO₂ exchange were followed. The rate of net photosynthesis was observed to fall from morning to evening, and this decline was more pronounced in winter than in summer. The maximal net photosynthetic rate, 223 ng CO₂g^-1dws^-1, occured in winter and the minimum, 94 ng CO₂g^-1dws^-1, late in spring. The light compensation point in summer was four times as high as in winter. In continuous light (180 or 90 mol photons m^-2s^-1, 15°C) net photosynthesis decreased noticeably during one week, falling below the level maintained in a 12 h light: 12 h dark cycle. Photosynthetic activity did not decrease, however, in lichens held in continuous light (90 mol photons m^-2s^-1) with cyclic changes of temperature (12 h 20 °C: 12 h 5 °C). Active photosynthesis was also maintained in light of cyclically changing intensity (12 h: 12 h, 15 °C) when night-time light was at least 75% lower than illumination by day. A dark period of 4 hours in a 24-h light:dark cycle was sufficient to keep CO₂ fixation at the control level. It seems that plants need an unproductive period during the day to survive and this can be induced by fluctuations in light and/or temperature.     

Tumourigenicity, cell-surface glycoprotein changes and ornithine decarboxylase gene pattern in Ehrlich ascites-carcinoma cells.

We selected a 2-difluoromethylornithine-resistant Ehrlich ascites-carcinoma cell line that grows in the presence of 20 mM-difluoromethylornithine. These cells contain 10-20 times the normal amount of hybridizable sequences for ornithine decarboxylase (EC 4.1.1.17) in their genomic DNA. We used these gene-amplified cells, their revertant counterparts (grown in the absence of the drug after an established gene amplification) and tumour cells grown in the presence of putrescine to investigate the changes of ornithine decarboxylase gene pattern and simultaneously occurring phenotypic changes, such as tumourigenicity and the expression of cell-surface glycoproteins. In the tumour cells reverted back to the normal gene frequency, not only did the amplified sequences disappear, but there were also signs of gene re-arrangements seen as a "gene jump', when a signal evidently moved to a heavier restriction fragment. Similar gene re-arrangement likewise occurred in cells exposed to putrescine. Although the wild-type tumour cells and the gene-amplified cells readily grew in the peritoneal cavity of mice, the revertant cells and the putrescine-treated cells had lost their tumourigenicity in mice. Gene-amplified tumour cells and the revertant cells showed distinct changes in their surface glycoprotein pattern in comparison with the parental cell line. These findings indicate that alterations of ornithine decarboxylase gene pattern/dosage may be associated with phenotypic changes possibly related to the tumourigenicity of these carcinoma cells.     

Bacterial oxidation of 2-tridecanone to 1-undecanol

A study of the microbial utilization of long-chain methyl ketones was under-taken. In general, enrichment culture experiments revealed that soil microorganisms capable of utilizing these compounds as growth substrates are ubiquitous. Gram-negative, rod-shaped bacteria were the prominent organisms exhibiting this capability. In particular, a strain of Pseudomonas isolated from soil degraded 2-tridecanone into several products that were recovered from cell-free culture fluid. These products were identified by gas-liquid chromatography as 2-tridecanol, 1-undecanol, 1-decanol, and undecanoic acid. A large amount of the substrate was converted to 1-undecanol. This compound was characterized further by classical methods of organic analysis. Unequivocal identification of 1-undecanol has established that some unique mechanism that involves subterminal oxidation must exist to degrade 2-tridecanone. No such mechanism has been reported for the biological degradation of long-chain, aliphatic, methyl ketones. A pathway for utilization of 2-tridecanone was proposed that is consistent with, but not confirmed by, the data presented.     

Localization of sequences regulating ancestral and acquired sites of esterase6 activity in Drosophila melanogaster

We have broadly defined the DNA regions regulating esterase6 activity in several life stages and tissue types of D. melanogaster using P- element-mediated transformation of constructs that contain the esterase6 coding region and deletions or substitutions in 5' or 3' flanking DNA. Hemolymph is a conserved ancestral site of EST6 activity in Drosophila and the primary sequences regulating its activity lie between -171 and -25 bp relative to the translation initiation site: deletion of these sequences decrease activity approximately 20-fold. Hemolymph activity is also modulated by four other DNA regions, three of which lie 5' and one of which lies 3' of the coding region. Of these, two have positive and two have negative effects, each of approximately twofold. Esterase6 activity is present also in two male reproductive tract tissues; the ejaculatory bulb, which is another ancestral activity site, and the ejaculatory duct, which is a recently acquired site within the melanogaster species subgroup. Activities in these tissues are at least in part independently regulated: activity in the ejaculatory bulb is conferred by sequences between -273 and -172 bp (threefold decrease when deleted), while activity in the ejaculatory duct is conferred by more distal sequences between -844 and -614 bp (fourfold decrease when deleted). The reproductive tract activity is further modulated by two additional DNA regions, one in 5' DNA (-613 to -284 bp; threefold decrease when deleted) and the other in 3' DNA (+1860 to +2731 bp; threefold decrease when deleted) that probably overlaps the adjacent esteraseP gene. Collating these data with previous studies suggests that expression of EST6 in the ancestral sites is mainly regulated by conserved proximal sequences while more variable distal sequences regulate expression in the acquired ejaculatory duct site.      

Phylogenetic diversification of immunoglobulin genes and the antibody repertoire

Immunoglobulins are encoded by a large multigene system that undergoes somatic rearrangement and additional genetic change during the development of immunoglobulin-producing cells. Inducible antibody and antibody-like responses are found in all vertebrates. However, immunoglobulin possessing disulfide-bonded heavy and light chains and domain-type organization has been described only in representatives of the jawed vertebrates. High degrees of nucleotide and predicted amino acid sequence identity are evident when the segmental elements that constitute the immunoglobulin gene loci in phylogenetically divergent vertebrates are compared. However, the organization of gene loci and the manner in which the independent elements recombine (and diversify) vary markedly among different taxa. One striking pattern of gene organization is the "cluster type" that appears to be restricted to the chondrichthyes (cartilaginous fishes) and limits segmental rearrangement to closely linked elements. This type of gene organization is associated with both heavy- and light-chain gene loci. In some cases, the clusters are "joined" or "partially joined" in the germ line, in effect predetermining or partially predetermining, respectively, the encoded specificities (the assumption being that these are expressed) of the individual loci. By relating the sequences of transcribed gene products to their respective germ-line genes, it is evident that, in some cases, joined-type genes are expressed. This raises a question about the existence and/or nature of allelic exclusion in these species. The extensive variation in gene organization found throughout the vertebrate species may relate directly to the role of intersegmental (V<==>D<==>J) distances in the commitment of the individual antibody-producing cell to a particular genetic specificity. Thus, the evolution of this locus, perhaps more so than that of others, may reflect the interrelationships between genetic organization and function.      

Differential effects of 2-difluoromethylornithine and methylglyoxal bis(guanylhydrazone) on the testosterone-induced growth of ventral prostate and seminal vesicles of castrated rats

2-Difluoromethylornithine totally prevented any increases in putrescine and spermidine concentrations in the ventral prostate of castrated rats during a 6-day testosterone treatment. Prostatic ornithine decarboxylase activity was inhibited by 80%, whereas S-adenosylmethionine decarboxylase was stimulated by more than 9-fold. In seminal vesicle, the inhibition of putrescine and spermidine accumulation, as well as of ornithine decarboxylase activity, was only minimal, and no stimulation of S-adenosylmethionine decarboxylase was observed. Administration of methylglyoxal bis(guanylhydrazone) to castrated androgen-treated rats resulted in a marked increase in concentrations of all prostatic polyamines. Prostatic ornithine decarboxylase activity was nearly 2 times and adenosylmethionine decarboxylase activity 9 times higher than that of the testosterone-treated animals. In contrast with ventral prostate, methylglyoxal bis(guanylhydrazone) treatment inhibited moderately the accumulation of spermidine and spermine in seminal vesicle, although both ornithine decarboxylase and S-adenosylmethionine decarboxylase activities were stimulated. Difluoromethylornithine inhibited significantly the weight gain of ventral prostate, but methylglyoxal bis(guanylhydrazone) produced a substantial increase in prostatic weight. These changes were largely due to the fact that the volume of prostatic secretion was greatly decreased by difluoromethylornithine, whereas methylglyoxal bis(guanylhydrazone) increased the amount of secretion. Treatment with difluoromethylornithine strikingly increased the methylglyoxal bis(guanylhydrazone) content of both ventral prostate and seminal vesicle, but even under these conditions the drug concentration remained low in comparison with other tissues. The results indicate that a combined use of these two polyamine anti-metabolites does not necessarily result in a synergistic growth inhibition of the androgen-induced growth of male accessory sexual glands.     

Transfer of intestine-derived diamines into tumour cells during treatment of Ehrlich-ascites--carcinoma-bearing mice with polyamine anti-metabolites.

Treatment of Ehrlich-ascites-carcinoma-bearing mice with methylglyoxal bis(guanylhydrazone) alone or in combination with 2-difluoromethylornithine greatly enhanced the transfer of intragastrically administered radioactive putrescine and cadaverine into the carcinoma cells. Difluoromethylornithine alone did not have any effect on the accumulation of intestine-derived diamines in the tumour cells. The frequently reported restoration of difluoromethylornithine-induced polyamine depletion on administration of methylglyoxal bis(guanylhydrazone) is in all likelihood attributable to a profound inhibition of intestinal diamine oxidase (EC 1.4.3.6), resulting in an enhanced entry of intestinal (bacterial) diamines into general circulation and finally into tumour cells.