CAPABILITY OF TUBULIN AND MICROTUBULES TO INCORPORATE AND TO RELEASE TYROSINE AND PHENYLALANINE AND THE EFFECT OF THE INCORPORATION OF THESE AMINO ACIDS ON TUBULIN ASSEMBLY

Abstract— Incorporation of [¹⁴C]tyrosine into the C-terminal position of α-tubulin of rat brain cytosol was 10-fold higher for non-assembled than for assembled tubulin. The incorporation into tubulin from disassembled microtubules was higher than into non-assembled tubulin; therefore, the low incorporation into microtubules was not due to a lower acceptor capacity of their tubulin constituent.
[¹⁴C]Tyrosine was released from assembled and non-assembled [¹⁴C]tyrosinated tubulin by the action of an endogenous carboxypeptidase. Release from non-assembled tubulin was shown by incubating a tubulinyl-[¹⁴C]tyrosine preparation in the presence of CaCl₂ at a concentration that abolished microtubule formation. Release from microtubules was inferred from the observation that the percentages of [¹⁴C]tyrosine released and the decrease of the specific radioactivity of the recovered microtubules were practically identical and did not change after a 10-fold dilution of the incubated microtubules.
[³H]Phenylalanine was released from a preparation of tubulinyl-[³H]phenylalanine also by an enzymatic activity.
The capacity of a tubulin preparation to incorporate tyrosine was increased 43% by pre-treatment with endogenous carboxypeptidase.
Tubulin tyrosinated in vitro was assembled to the same extent as native tubulin. After a mixture of tubulinyl-[¹⁴C]tyrosine and tubulinyl-[³H]phenylalanine was partially assembled, the ratio of ¹⁴C/³H found in the microtubules was the same as in the non-assembled tubulin fraction.     

Effects of cytoprotective antioxidants on lymphocytes from representative mitochondrial neurodegenerative diseases

Two new aza analogues of the neuroprotective agent idebenone have been synthesized and characterized. Their antioxidant activity, and ability to augment ATP levels have been evaluated in several different cell lines having suboptimal mitochondrial function. Both compounds were found to be good ROS scavengers, and to protect the cells from oxidative stress induced by glutathione depletion. The compounds were more effective than idebenone in neurodegenerative disease cells. These novel pyrimidinol derivatives were also shown to augment ATP levels in coenzyme Q₁₀-deficient human lymphocytes. The more lipophilic side chains attached to the pyrimidinol redox core in these compounds resulted in less inhibition of the electron transport chain and improved antioxidant activity.     

Growth, mortality and reproduction of the blue tilapia Oreochromis aureus (Perciformes: Cichlidae) in the Aguamilpa Reservoir, Mexico

Tilapia production has increased in Aguamilpa Reservoir, in Nayarit, Mexico, in the last few years and represents a good economic activity for rural communities and the country. We determined growth parameters, mortality and reproductive aspects for 2413 specimens of blue tilapia Oreochromis aureus in this reservoir. Samples were taken monthly from July 2000 through June 2001, of which 1 371 were males and 1 042 were females. Standard length (SL) and total weight (TW) were measured in each organism. The SL/TW relationships through power models for sexes were determined. The growth parameters L infinity k, and t0 of the von Bertalanffy equation were estimated using frequency distribution of length through ELEFAN-I computer program. Finally the reproductive cycle and size of first maturity were established using morph chromatic maturity scale. The results suggested that the males and females had negative allometric growth (b < 3). Significant differences were found between SL/TW model for the sexes, suggesting separate models for males and females. Results indicate that there are no differences in growth rates between sexes; the proposed parameters were L infinity = 43.33 cm standard length, k = 0.36/year and t0 = -0.43 years. Natural and fishing mortality coefficients were 0.83/year and 1.10/year, respectively. The estimated exploitation rate (0.57/year) suggested that during the study period the fishery showed signs of overfishing. Blue tilapia reproduces year-round; the highest activity occurs from January through May and size of first maturity was 23 cm SL. We conclude that it is necessary to establish a minimum catch size in this reservoir based on the reproductive behavior of this species.     

Composition of carrageenan blends inferred from C-NMR and infrared spectroscopic analysis

Carrageenan blends mainly composed of kappa, iota and lambda carrageenan were investigated by ¹³C-NMR and infrared (IR) spectroscopy. ¹³C-NMR was shown to be a powerful tool for quantifying the kappa-iota ratio in a carrageenan blend. The technique, however, was not helpful for identifying lambda carrageenan. Consequently, IR spectroscopy was used to achieve a semi-quantitative determination of lambda carrageenan. By combining both techniques, the carrageenan composition of several commercial samples was established.     

The role of sexual steroids in the modulation of growth hormone (GH) secretion in humans

Sex steroids contribute to modulate GH secretion in man. However, both the exact locus and mechanism by which their actions are exerted still remain not clearly understood. We undertook a number of studies designed to ascertain: (1) whether or not sudden or chronic changes in circulating gonadal steroids may affect GH secretion in normal adults; and (2) the reason(s) for gender-related dimorphic pattern of GH release. The pituitary reserve of GH, as evaluated by means of a GHRH challenge, was similar in women with anorexia nervosa and in normally menstruating women. Estrogenic receptor blockade with tamoxifen (TMX) did not significantly change GHRH-induced GH response in these normal women. Therefore, acute or chronic hypoestrogenism apparently had no important effects at level of somatotrophs. In another group of normal women we tested the possibility that changes in circulating estrogens might induce changes in the hypothalamic-somatotroph rhythm (HSR). GHRH challenges were performed throughout a menstrual cycle, and again after having achieved functional ovarian blockade with a GnRH agonist treatment. Short-term ovarian blockade did not significantly affect the parameters of GH response to GHRH, although it was accompanied by an increase in the number of women in a refractory HSR phase at testing. This suggested a low potentiating effect on the basic pattern of somatostatin (SS) release occurring as a consequence of the decrease in circulating estrogens. In normal men, neither the GH response to GHRH nor the HSR were affected by functional testicular blockade (after GnRH agonist treatment). However, the administration of testosterone enanthate (250 mg) to another group of men increased both the GHRH-induced GH release and the number of subjects in a spontaneous secretory HSR phase at testing; these were reversed by estrogenic receptor blockade with TMS. In another group of normal men, the fraction of GH secreted in pulses (FGHP) during a nocturnal sampling period was significantly decreased by testicular blockade. Other parameters of GH secretion,such as the number of GH pulses and their mean amplitude (A), and the mean plasma GH concentration (MCGH), showed a slight, although not significant, decrease following the lack of androgens. The administration of testosterone enanthate (500 mg) reversed these parameters to values similar to those in the basal study. Interestingly, when tamoxifen was given after testosterone enanthate, A, MCGH and FGHP increased to values significantly higher than in any other experimental condition in that study.(ABSTRACT TRUNCATED AT 400 WORDS)     

Regulation of plasma membrane Ca2 +-ATPase activity by acetylated tubulin: Influence of the lipid environment

We demonstrated previously that acetylated tubulin inhibits plasma membrane Ca^2 +-ATPase (PMCA) activity in plasma membrane vesicles (PMVs) of rat brain through a reversible interaction. Dissociation of the PMCA/tubulin complex leads to restoration of ATPase activity. We now report that, when the enzyme is reconstituted in phosphatidylcholine vesicles containing acidic or neutral lipids, tubulin not only loses its inhibitory effect but is also capable of activating PMCA. This alteration of the PMCA-inhibitory effect of tubulin was dependent on concentrations of both lipids and tubulin. Tubulin (300 μg/ml) in combination with acidic lipids at concentrations > 10%, increased PMCA activity up to 27-fold. The neutral lipid diacylglycerol (DAG), in combination with 50 μg/ml tubulin, increased PMCA activity > 12-fold, whereas tubulin alone at high concentration (≥ 300 μg/ml) produced only 80% increase. When DAG was generated in situ by phospholipase C incubation of PMVs pre-treated with exogenous tubulin, the inhibitory effect of tubulin on PMCA activity (ATP hydrolysis, and Ca^2 + transport within vesicles) was reversed. These findings indicate that PMCA is activated independently of surrounding lipid composition at low tubulin concentrations (< 50 μg/ml), whereas PMCA is activated mainly by reconstitution in acidic lipids at high tubulin concentrations. Regulation of PMCA activity by tubulin is thus dependent on both membrane lipid composition and tubulin concentration.     

The effects of drying on sediment nitrogen content in a Mediterranean intermittent stream: a microcosms study

Mediterranean climates predispose aquatic systems to both flood and drought periods, therefore, stream sediments may be exposed to desiccation periods. Changes in oxygen concentrations and sediment water content influence the biotic processes implicated in nitrogen dynamics. The objectives of this study were to identify (1) the changes of inorganic nitrogen in stream sediments during the transition from wet to dry conditions, and (2) the underlying processes in N dynamics and its regulation. Extractable sediment NO₃ ⁻-N and NH₄ ⁺-N, organic matter and extractable organic carbon content were assessed during natural desiccation in microcosms with sediments from an intermittent Mediterranean stream. In agreement with our initial hypothesis, our results showed how the NO₃ ⁻-N content of the sediment was enhanced during the first 10 days of sediment drying, whereas NH₄ ⁺-N was lost by 14 days post-drying. During the first 10 days, sediment desiccation seemed to stimulate the net N-mineralization and net nitrification from sediments. Afterwards, the extractable NO₃ ⁻-N concentration sharply dropped, which may be attributed to lower ammonium-oxidation rates as ammonium and organic matter are depleted, and to an increase in NO₃ ⁻-N consumption by microbial populations. Denitrification was inhibited, with a significant decrease as % water-filled pore space lowered. We hypothesize that the sediment inorganic N content enhanced during sediment desiccation could be released as part of the N pulse observed after sediment rewetting. However, the stream N availability after rewetting dried sediments would differ depending on desiccation period duration.