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51.
Eiichi Tachikawa A. William Tank David H. Weiner Werner F. Mosimann Nobuyuki Yanagihara Norman Weiner 《Journal of neurochemistry》1987,48(5):1366-1376
Abstract: Incubation of rat pheochromocytoma PC12 cells with 4β-phorbol-12β-myristate-13α-acetate (PMA), an activator of Ca2+/phospholipid-dependent protein kinase (protein kinase C), or forskolin, an activator of adenylate cyclase, is associated with increased activity and enhanced phosphorylation of tyrosine hydroxylase. Neither the activation nor increased phosphorylation of tyrosine hydroxylase produced by PMA is dependent on extracellular Ca2+. Both activation and phosphorylation of the enzyme by PMA are inhibited by pretreatment of the cells with trifluo-perazine (TFP). Treatment of PC 12 cells with l-oleoyl-2-acetylglycerol also leads to increases in the phosphorylation and enzymatic activity of tyrosine hydroxylase; 1, 2-diolein and 1, 3-diolein are ineffective. The effects of forskolin on the activation and phosphorylation of the enzyme are independent of Ca2+ and are not inhibited by TIT5. Forskolin elicits an increase in cyclic AMP levels in PC 12 cells. The increases in both cyclic AMP content and the enzymatic activity and phosphorylation of tyrosine hydroxylase following exposure of PC 12 cells to different concentrations of forskolin are closely correlated. In contrast, cyclic AMP levels do not increase in cells treated with PMA. Tryptic digestion of the phosphorylated enzyme isolated from untreated cells yields four phosphopeptides separable by HPLC. Incubation of the cells in the presence of the Ca2+ ionophore ionomycin increases the phosphorylation of three of these tryptic peptides. However, in cells treated with either PMA or forskolin, there is an increase in the phosphorylation of only one of these peptides derived from tyrosine hydroxylase. The peptide phosphorylated in PMA-treated cells is different from that phosphorylated in forskolin-treated cells. The latter peptide is identical to the peptide phosphorylated in dibutyryl cyclic AMP-treated cells. These results indicate that tyrosine hydroxylase is activated and phosphorylated on different sites in PC 12 cells exposed to PMA and forskolin and that phosphorylation of either of these sites is associated with activation of tyrosine hydroxylase. The results further suggest that cyclic AMP-dependent and Ca2+/ phospholipid-dependent protein kinases may play a role in the regulation of tyrosine hydroxylase in PC 12 cells. 相似文献
52.
E. Korpimäki 《Oecologia》1987,74(2):277-285
Summary Food samples of breeding Kestrels (Falco tinnunculus) and Long-eared Owls (Asio otus) were collected in the peak and low phase of their preferred prey (Microtus voles) in western Finland. Diets of pairs that bred as neighbours (1 km) with interspecifics were compared with those of non-neighbours. In both species, neighbouring pairs fed less on Microtus voles and more on alternative prey than did non-neighbours. Competition theory predicts that diet overlap should be lower during prey shortage and that diet similarity should be especially reduced in neighbouring pairs. Observations were consistent with expectations: diet similarity was lower in the low vole years and neighbouring pairs showed less diet overlap that non-neighbours. Differences in habitat composition and prey availability at the sample sites should not confuse the results. In addition to the high diet similarity, hunting habitats and nest sites of the species overlapped almost completely; they only showed clear temporal segregation in hunting. Probably because of food competition, the neighbouring pairs of both species produced significantly fewer young than the non-neighbours. These results contrast with the view that the diet composition and dietary shift of rodent-feeding predatory birds can be interpreted in terms of simple opportunistic foraging. In the breeding season, interspecific competition for food seems to be an important factor that affects the niches of these species, especially in northern areas, where the seasonal low phase of voles in spring and the number of alternative prey are lower than in more southern areas. 相似文献
53.
The intracellular localisation of phytochrome and ubiquitin in irradiated oat coleoptiles was analysed by electron microscopy. We applied indirect immunolabeling with polyclonal antibodies against phytochrome from etiolated oat seedlings or polyclonal antibodies against ubiquitin from rabbit reticulocytes, together with a goldcoupled second antibody, on serial ultrathin sections of resin-embedded material. Immediately after a 5-min pulse of red light-converting phytochrome from the red-absorbing (Pr) to the far-redabsorbing (Pfr) form-the label for phytochrome was found to be sequestered in electron-dense areas. For up to 2 h after irradiation, the size of these areas increased with increasing dark periods. The ubiquitin label was found in the same electrondense areas only after a dark period of 30 min. A 5 min pulse of far-red light, which reverts Pfr to Pr, given immediately after the red light did not cause the electron-dense structures to disappear; moreover, they contained the phytochrome label immediately after the far-red pulse. In contrast, after the reverting far-red light pulse, ubiquitin could only be visualised in the electron-dense areas after prolonged dark periods (i.e. 60 min). The relevance of these data to light-induced phytochrome pelletability and to the destruction of both Pr and Pfr is discussed.Abbreviations FR
far-red light; Pfr
- Pr
far-red-absorbing and red-absorbing forms of phytochrome, respectively
- R
red light 相似文献
54.
Seedlings of barley were grown either in continuous darkness or under a diurnal 12 h light/12 h dark cycle and the effects on NADPH-protochlorophyllide oxidoreductase were followed at two different levels. Firstly, the relative content of the mRNA encoding the NADPH-protochlorophyllide oxidoreductase was measured by dot-blot hybridization. Secondly, changes in the enzyme polypeptide were monitored either by the method of immunoblotting or by immunogold labelling of ultrathin sections of Lowicryl-embedded leaf tissue. Our results demonstrate that drastic diurnal changes in the level of mRNA sequences and the enzyme protein are unlikely to occur in plants which have been grown under natural light/dark conditions. In the dark, protein and mRNA accumulation occurs at an early developmental stage. These results are difficult to reconcile with the suggestion that the massive accumulation of mRNA and enzyme protein in dark-grown seedlings is primarily the consequence of an artificially extended darkperiod. In addition to the plastid-specific NADPH-protochlorophyllide oxidoreductase a closely related polypeptide has been detected outside the plastid in the surrounding cytoplasm (Dehseh et al. 1986b, Planta 169, 172–183). During the diurnal light/dark treatment of seedlings the concentrations of the two protein populations did not show any variation indicative of an exchange between the two protein populations across the plastid envelope.Abbreviation poly(A)+RNA
polyadenylated RNA 相似文献
55.
A procedure was developed which allows the large-scale isolation of root hairs from seedlings of Pisum sativum . L. cvs. Kleine Rheinländerin and Rosa Krone. The method may yield up to 50 g fresh weight of root hairs per 3.104 seedlings. In a modified form considerable amounts of root hair material may be harvested, even after incubation of the roots in aqueous solutions. Thus, detailed biochemical studies on the root hair system have become feasible.
The occurrence of specific proteins in membrane fractions of P. sativum root hairs was demonstrated as follows: Incubation of root hairs in situ with 3-azidonaphthalene-2,7-disulfonate – a strongly anionic, photoactivated fluorescent marker – followed by gel electrophoresis of membrane fractions showed the presence of root-hair specific proteins which, since the system was intact, suggests that they are on the outer surface of the cells. 相似文献
The occurrence of specific proteins in membrane fractions of P. sativum root hairs was demonstrated as follows: Incubation of root hairs in situ with 3-azidonaphthalene-2,7-disulfonate – a strongly anionic, photoactivated fluorescent marker – followed by gel electrophoresis of membrane fractions showed the presence of root-hair specific proteins which, since the system was intact, suggests that they are on the outer surface of the cells. 相似文献
56.
Influx of na, k, and ca into roots of salt-stressed cotton seedlings : effects of supplemental ca
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High Na+ concentrations may disrupt K+ and Ca2+ transport and interfere with growth of many plant species, cotton (Gossypium hirsutum L.) included. Elevated Ca2+ levels often counteract these consequences of salinity. The effect of supplemental Ca2+ on influx of Ca2+, K+, and Na+ in roots of intact, salt-stressed cotton seedlings was therefore investigated. Eight-day-old seedlings were exposed to treatments ranging from 0 to 250 millimolar NaCl in the presence of nutrient solutions containing 0.4 or 10 millimolar Ca2+. Sodium influx increased proportionally to increasing salinity. At high external Ca2+, Na+ influx was less than at low Ca2+. Calcium influx was complex and exhibited two different responses to salinity. At low salt concentrations, influx decreased curvilinearly with increasing salt concentration. At 150 to 250 millimolar NaCl, 45Ca2+ influx increased in proportion to salt concentrations, especially with high Ca2+. Potassium influx declined significantly with increasing salinity, but was unaffected by external Ca2+. The rate of K+ uptake was dependent upon root weight, although influx was normalized for root weight. We conclude that the protection of root growth from salt stress by supplemental Ca2+ is related to improved Ca-status and maintenance of K+/Na+ selectivity. 相似文献
57.
Vacuoles of tobacco mesophyll and of suspension-cultured cells were isolated in order to study the localization of peroxidase isoenzymes. Only basic peroxidases were detectable by electrophoretic separation of the vacuolar sap. Some of the basic peroxidases have formerly been described as an ionically bound cell-wall fraction. This fraction, however, was found to be an artifact produced by incomplete cell breakage. Reinvestigation of isolated cell walls confirmed that mainly acidic peroxidases are localized in the cell walls where they move freely or are bound. As a consequence of former and present results we think it probable that all of the peroxidase isoenzymes are secretory proteins because they have to be transported from the sites of synthesis in the cytoplasm to the sites of function, the extracytoplasmic spaces, cell wall (acidic peroxidases), and vacuole (basic peroxidases).Abbreviation ER
endoplasmic reticulum
- PAGE
polyacrylamide gel electrophoresis 相似文献
58.
59.
It has been suggested that Locusta migratoria amplifies its ribosomal RNA genes in the growing oocytes (Kunz (1967) Chromosoma20, 332–370). Cloned ribosomal DNA of L. migratoria was used to analyze rDNA structure and number. The rDNA is localized on three chromosome pairs in six nucleolus organizers. It was found that all structural variants of the rRNA genes which have been described previously are represented in the same relative amounts in DNA from isolated oocytes as in somatic cells. Furthermore, the rRNA gene number is not increased in oocyte DNA, i.e., amplification does not occur. Therefore, the large number of multiple nucleoli seen in the growing oocytes has to be interpreted as the fully extended and fully active set of chromosomal rRNA genes. The total rRNA gene number was determined by dot blot hybridization to be about 3300 genes/haploid genome. 相似文献
60.
Summary Nonstationary pump currents which have been observed in K+-free Na+ media after activation of the Na,K-ATPase by an ATP-concentration jump (see the preceding paper) are analyzed on the basis of microscopic reaction models. It is shown that the behavior of the current signal at short times is governed by electrically silent reactions preceding phosphorylation of the protein; accordingly, the main information on charge-translocating processes is contained in the declining phase of the pump current. The experimental results support the Albers-Post reaction scheme of the Na,K-pump, in which the translocation of Na+ precedes translocation of K+. The transient pump current is represented as the sum of contributions of the individual transitions in the reaction cycle. Each term in the sum is the product of a net transition rate times a dielectric coefficient describing the amount of charge translocated in a given reaction step. Charge translocation may result from the motion of ion-binding sites in the course of conformational changes, as well as from movement of ions in access channels connecting the binding sites to the aqueous media. A likely interpretation of the observed nonstationary currents consists in the assumption that the principal electrogenic step is the E1-P/P-E2 conformational transition of the protein, followed by a release of Na+ to the extracellular side. This conclusion is supported by kinetic data from the literature, as well as on the finding that chymotrypsin treatment which is known to block the E1-P/P-E2 transition abolishes the current transient. By numerical simulation of the Albers-Post reaction cycle, the proposed mechanism of charge translocation has been shown to reproduce the experimentally observed time behavior of pump currents. 相似文献