Control Of Ornithine Decarboxylase Activity Research Articles

Control of ornithine decarboxylase activity by polyamines and absence of antizyme in Tetrahymena

1. In cells of Tetrahymena pyriformis and thermophila, ODC activity was significantly suppressed but ODC decay was not stimulated by putrescine. 2. Free antizyme and ODC-antizyme complex were both not detected in extracts of cells of T. pyriformis treated with putrescine. 3. It was concluded that in Tetrahymena, unlike vertebrate cells, ODC is not subject to polyamine-induced destabilization mediated by antizyme.

Polyamine Metabolism and Growth of Neurospora Strains Lacking Cis-Acting Control Sites in the Ornithine Decarboxylase Gene

Ornithine decarboxylase (ODC) initiates the synthesis of polyamines (putrescine, spermidine, and spermine) and is highly regulated. We wished to know the importance of the control of ODC synthesis to the rates of growth and polyamine synthesis in the fungus, Neurospora crassa. We identified two control sites of the spe-1 gene, encoding ODC. One was an upstream activation region (UAR) and the other was the DNA encoding the long ODC mRNA leader, which governs polyamine-mediated repression of enzyme synthesis. Transformants receiving copies of spe-1 sequences lacking the UAR compensated for the deficiency by derepression or enzyme stabilization; polyamine synthesis was almost normal. A transformant lacking the spe-1 mRNA leader DNA constitutively expressed ODC mRNA and ODC activity, and synthesized excessive putrescine, especially when provided exogenous ornithine. This transformant grew normally and had only mildly elevated pools of spermidine, the major polyamine of this organism. We conclude that ODC activity normally limits polyamine synthesis, and ornithine becomes limiting in the ODC-constitutive strain. In this strain, however, spermidine synthesis remains rigorously limited by another step of the pathway, as yet unidentified. Thus the control of ODC activity in Neurospora is not vital to growth in laboratory culture, and synthesis of toxic levels of spermidine is limited by other mechanisms.

Control of ornithine decarboxylase activity in jute seeds by antizyme

The control of ornithine decarboxylase activity by antizyme was studied during early germination of jute seeds(Corchorus olitorius). When 2 mM of putrescine and spermidine were applied to the germinating medium, the enzyme activity was markedly inhibited (1.7-fold) during 16 h imbibition. This inhibition could be attributed to the formation of an inhibitory protein termed antizyme. The antizyme was partially purified from jute and barley seedlings. The activity of jute ornithine decarboxylase antizyme was weaker than that of barley.

Control of ornithine decarboxylase activity in alpha-difluoromethylornithine-resistant L1210 cells by polyamines and synthetic analogues.

The regulation of ornithine decarboxylase (ODC) activity by the polyamine derivatives N1,N8-bis(ethyl)-spermidine and N1,N12-bis(ethyl)spermine was studied using a line of L1210 cells resistant to alpha-difluoromethylornithine (D-R cells), which contain very high levels of ODC, and a synthetic mRNA prepared from a plasmid containing an insert corresponding to ODC mRNA adjacent to an SP6 RNA polymerase promoter. Studies in which ODC protein was labeled in the D-R cells by exposure to [35S]methionine indicated that the polyamine derivatives and their physiological counterparts led to an increased rate of degradation of ODC and to a rapid reduction in ODC synthesis without affecting the content of ODC mRNA. Direct evidence that the polyamine derivatives act by inhibiting the translation of the ODC mRNA was obtained by studying their effects on the translation of ODC mRNA in reticulocyte lysates. This translation was strongly inhibited by the addition of N1,N8-bis(ethyl)spermidine, spermidine, N1,N12-bis(ethyl)spermine, or spermine but was not affected much by putrescine. The inhibition of the translation of ODC mRNA by either of the bis(ethyl) polyamine derivatives occurred at concentrations which stimulated total protein synthesis showing the selectivity of the reduction in ODC. The effects of polyamine derivatives and polyamines on translation of the plasmid-derived ODC mRNA were identical with those found with the D-R L1210 cell mRNA. This synthetic ODC mRNA lacks 261 bases of the 5'-leader sequences and 200 bases plus the poly(A) section from the 3'-nontranslated sequence. Therefore, these regions appear not to influence sensitivity of the ODC mRNA to inhibition of translation by polyamine derivatives.

Restoration of normal ornithine decarboxylase antizyme activity in rat liver after acute carcinogen treatment.

This study was undertaken to see whether or not the decrease in ornithine decarboxylase antizyme activity caused in rat liver by a hepatocarcinogen could be reversed. Thioacetamide was administered only once, in a single i.p. injection and at a non-carcinogenic, non-necrogenic dose. The activities of both hepatic ornithine decarboxylase and hepatic ornithine decarboxylase antizyme were measured at intervals of hours after the injection of thioacetamide. The hepatic ornithine decarboxylase antizyme in thioacetamide-treated rats was minimal at 40 and 80 h after carcinogen administration. The reversal process requires a very long time, namely 450 h for normal levels of hepatic ornithine decarboxylase antizyme activity to be restored in treated rats. This time is much longer than that required to restore normal ornithine decarboxylase activity in liver of thioacetamide-treated rats. The results of this study, combined with those of the preceding paper, demonstrate that hepatocarcinogens cause a relative inability of rat liver cells to make the ornithine decarboxylase antizyme and that the irreversibility of this defect in cellular control of ornithine decarboxylase activity may be a constant feature in the neoplastic transformation of the rat liver.

Cellular control of ornithine decarboxylase activity by its antizyme.

Conditions have been established under which the antizyme of ornithine decarboxylase (E.C. 4.1.1.17, L-ornithine carboxy-lyase, ODC) a non-competitive protein inhibitor of ODC, can be detected in cells in response to as little as 10(-7) M putrescine. The maintenance of intracellular antizyme activity depends upon the continued presence of putrescine in the medium. Removal of putrescine results in a rapid decline of antizyme activity. These phenomena are unaffected by the presence of cycloheximide and are comparable to the requirement of L-asparagine for the maintenance of ODC activity. The extent to which the antizyme level is increased is inversely related to the preexisting level of intracellular ODC at the time of addition of putrescine. The time of appearance of free antizyme is delayed in cells that have high levels of ODC; the amount of free antizyme that can be assayed for in these cells, at any particular time is correspondingly less. The converse is also true. In cells that have high levels of antizyme, the delay in appearance of ODC is greater and the amount of ODC that can be assayed for is correspondingly less than in cells with low levels of antizyme. These experiments, as well as others, indicate that the ODC antizyme and ODC interact in vivo with each other to modify their respective activities.

Control of ornithine decarboxylase activity by cyclic nucleotides in the phytohemagglutinin induced lymphocyte transformation

Summary N2,O2′-Dibutyryl guanosine 3′:5′-cyclic monophosphate enhanced the ornithine decarboxylase activity which was induced during lymphocyte transformation by phytohemagglutinin, while N6,O2′-dibutyryl adenosine 3′:5′-cyclic monophosphate tended to inhibit it. The increased ornithine decarboxylase activity was not observed when N2,O2′-dibutyryl guanosine 3′:5′-cyclic monophosphate alone was added to the non-stimulated lymphocytes. In contrast to the dibutyryl derivative, guanosine 3′:5′-cyclic monophosphate did not show any effect on enzyme induction following phytohemagglutinin stimulation.

Control of ornithine decarboxylase activity in stimulated human lymphocytes by putrescine and spermidine.

Ornithine decarboxylase activity in extracts of phytohaemagglutinin-stimulated human lymphocytes is rapidly and extensively inhibited by additions of micromolar concentrations of putrescine or spermidine to the culture medium. This inhibition is not due to feedback inhibition of the enzyme by putrescine, spermidine or their metabolites. Inhibition is dependent on the continuation of protein synthesis, but does not require RNA synthesis. The effect of putrescine is abolished when its conversion into spermidine by the cells is prevented.

Ornithine decarboxylase activity and polyamine synthesis during kidney hypertrophy

We measured ornithine decarboxylase activity in rat kidneys obtained at various times following unilateral nephrectomy. The enzyme activity shows a biphasic response during renal compensatory hypertrophy, the major peak of activity, which is approx. 4–5 times higher than the activity in the kidney of sham-operated rats, is reached approx. i8 h following nephrectomy. The concentration of putrescine, spermidine and spermine show elevations in the first two days after the operation. Hydrocortisone or growth hormone administration rapidly increase ornithine decarboxylase activity in rat kidney. Actinomycin given together or i h after hydrocortisone injection prevents the increase in ornithine decarboxylase activity detected 4 h after hormone administration. Such inhibition does not occur when actinomycin is given 2 h following hydrocortisone. A comparison between the stimulation of polyamine metabolism occurring in compensatory growth of the kidney and liver is presented and the results discussed in relation to a possible post-transcriptional mechanism involved in the control of ornithine decarboxylase activity. Our results provide one more example of stimulation of polyamine metabolism at the onset of growth processes in the adult organism.

The control of ornithine decarboxylase activity during liver regeneration

The effect of amino acids and growth hormone on rat liver ornithine decarboxylase activity was investigated in an attempt to elucidate the mechanisms that control the activity of this enzyme during the early stages of liver regeneration. A single administration of casein hydrolyzate to intact rats produces a marked increase in liver ornithine decarboxylase activity. The casein effect is abolished by puromycin or actinomycin, is not modified by adrenalectomy and is slowed down by hypophysectomy. Histidine also causes a rapid increase in the activity of the enzyme but the amino acid effect is blocked by puromycin, is not inhibited by actinomycin, and is entirely absent in hypophysectomized rats. Growth hormone injections increase ornithine decarboxylase activity in normal or regenerating liver. The action of the hormone is only slightly inhibited by doses of actinomycin that prevent any increase in the specific activity of nuclear or cytoplasmic rapidly labeled RNA. 8-Azaguanine injection or small doses of actinomycin do not block the initial increase in ornithine decarboxylase activity occurring shortly after partial hepatectomy. The possibility that a post-transcriptional mechanism is involved in the control of ornithine decarboxylase activity during liver regeneration is discussed in detail.