Deficits in the Activation and Phosphorylation of Hippocampal Tyrosine Hydroxylase in the Aged Fischer 344 Rat Following Intraventricular Administration of 6-Hydroxydopamine

Abstract: Tyrosine hydroxylase activity was measured under optimal and suboptimal assay conditions in hippocampal extracts from young (2 month), mature (12 month), and old (24 month) Fischer 344 male rats 72 h after the infusion of 200 µg of the neurotoxin 6-hydroxydopamine or vehicle into the lateral ventricle. The lesion resulted in a 45–55% decrease of tyrosine hydroxylase activity measured under optimal conditions (pH 6.1, 3.0 m M 6-methyl-5,6,7,8-tetrahydropterin) and an ∼35% decrease in the relative concentration of immunoreactive tyrosine hydroxylase. When measured under suboptimal conditions (pH 6.6, 0.7 m M 6-methyl-5,6,7,8-tetrahydropterin), tyrosine hydroxylase activity in 2- and 12-month-old lesioned animals was twice that measured in vehicle-treated animals. However, in the old lesioned animals, tyrosine hydroxylase activity measured under suboptimal conditions was not different from that measured in age-matched vehicle-treated animals. Isoforms of tyrosine hydroxylase were identified on immunoblots after two-dimensional gel electrophoresis using enhanced chemiluminescence. The relative proportion of lower pl isoforms of tyrosine hydroxylase in the 2-month-old lesioned animals was greater than that observed in vehicle-treated controls. In contrast, no difference was seen in the relative proportion of tyrosine hydroxylase isoforms in the 24-month-old lesioned versus control animals. These data indicate that the ability of locus ceruleus neurons to rapidly respond to and compensate for insult is attenuated in 24-month-old Fischer 344 rats due to a deficit in stimulus-evoked enzyme phosphorylation.     

Polypeptide composition,assembly and phosphorylation patterns of the photosystem II antenna system of Chlamydomonas reinhardtii

In recent years major progress has been made in describing the gene families that encode the polypeptides of the light-harvesting antenna system of photosystem II (PSII). At the same time, advances in the biochemical characterization of these antennae have been hampered by the high degree of similarity between the apoproteins. To help interpret the molecular results, we have re-examined the composition, the assembly and the phosphorylation patterns of the light-harvesting antenna of PSII (LHCII) in the green alga Chlamydomonas reinhardtii Dang, using a non-Tris SDS-PAGE system capable of resolving polypeptides that differ by as little as 200 daltons. Research to date has suggested that in C. reinhardtii the LHCII comprises just four polypeptides (p11, p13, p16 and p17), and CP29 and CP26 just one polypeptide each (p9 and p10, respectively), i.e. a total of six polypeptides. We report here that these antenna systems contain at least 15 polypeptides, 10 associated with LHCII, 3 with CP29, and 2 with CP26. All of these polypeptides have been positively identified by means of appropriate antibodies. We also demonstrate substantial heterogeneity to the pattern of in-vitro phosphorylation, with major differences found among members of closely spaced and immunologically related polypeptides. Most intriguing is the fact that the polypeptides that cross-react with the anti-type 2 LHCII antibodies of higher plants (p16, and to a lesser extent p11) are not phosphorylated, whereas in higher plants these are the most highly phosphorylated polypeptides. Also, unlike in higher plants, CP29 is heavily phosphorylated. Phosphorylation does not appear to have any effect on the mobility of polypeptides on fully denaturing SDS-PAGE gels. To learn more about the accumulation and organization of the light-harvesting polypeptides, we have also investigated a chlorophyll b-less mutant, cbn1-48. The LHCII is almost completely lost in this mutant, along with at least some LHCI. But the accumulation of CP29 and CP26 and their binding to PSII core complexes, is relatively unaffected. As expected, the loss of antenna polypeptides is accompanied by a reduction of the size of large reaction-center complexes. Following in-vitro phosphorylation the number of phosphorylated proteins is greatly increased in the mutant thylakoids compared to wildtype thylakoids. We present a model of the PSII antenna system to account for the new polypeptide complexity we have demonstrated.This work was supported by National Institute of Health grant GM22912 to L.A.S. We would like to thank Anastasios Melis for helpful discussions.     

Effect of gliadins and HMW and LMW subunits of glutenin on dough properties in the F6 recombinant inbred lines from a bread wheat cross

The storage proteins of 64 F₂-derived F₆ recombinant inbred lines (RILs) from the bread wheat cross Prinqual/Marengo were analyzed. Parents differed at four loci: Gli-B1 (coding for gliadins), Glu-B1 (coding for HMW glutenin subunits), Glu-A3/Gli-A1 (coding for LMW glutenin subunits/gliadins) and Glu-D3 (coding for LMW glutenin subunits). The effect of allelic variation at these loci on tenacity, extensibility and dough strength as measured by the Chopin alveograph was determined. Allelic differences at the Glu-B1 locus had a significant effect on only tenacity. None of the allelic differences at either the Glu-A3/Gli-A1 or Glu-D3 loci had a significant effect on quality criteria. Allelic variation at the Gli-B1 locus significantly affected all of the dough properties. Epistatic effects between some of the loci considered contributed significantly to the variation in dough quality. Additive and epistatic effects each accounted for 15% of the variation in tenacity. Epistasis accounted for 15% of the variation in extensibility, whereas additive effects accounted for 4%. Epistasis accounted for 14% of the variation in dough strength, and additivity for 9%. The relative importance of epistatic effects suggest that they should be included in predictive models when breeding for breadmaking quality.     

Implications of the variance effective population size on the genetic conservation of monoecious species

The concept of variance effective population size [Ne_(v)] and other expressions are reviewed and described for specific sampling steps in germplasm collection and regeneration of monoecious species. Special attention is given to procedures for computing the variance of the number of contributed gametes [V(k)] to the next generation. Drift, as it occurs between generations, was considered to contain a component due to the sampling of parents and a subsequent component due to the sampling of gametes. This demonstrates that drift, caused by reduction of seed viability, damages the genetic integrity of accessions stored in germplasm banks. The study shows how mating designs, such as plant-to-plant or chain crossings with additional female gametic control, can partially alleviate this problem. Optimal procedures for increasing Ne_(v) when collecting germplasm in the field are also discussed. The effect of different female and male gametic control strategies on Ne_(v) is considered under several situations. Practical examples illustrating the use of V(k) and Ne_(v) expressions are given.     

Inheritance of resistance to blackmold (Alternaria alternata (Fr.) Keissler) in two interspecific crosses of tomato (Lycopersicon esculentum ×L. cheesmanii f.typicum)

Inheritance of resistance to blackmold, a disease of ripe tomato fruit caused byAlternaria alternata, was studied in two interspecific crosses. The parents, F₁ and F₂ generations of a cross between the susceptibleLycopersicon esculentum Mill. cultivar Hunt 100 and the resistantL. Cheesmanii f.typicum Riley accession LA 422, and the parents, F₁, F₂, F₃, and BC₁ P₂ generations of a cross between the susceptibleL. Esculentum cv. VF 145B-7879 and LA 422 were evaluated. The following disease evaluation traits were used: symptom rating (a symptom severity rating based on visual evaluation of lesions), diseased fruit (the number of diseased fruits divided by the total number of fruit scored), and lesion size (a function derived from the actual lesion diameter). Generation means analysis was used to determine gene action. The data of the Hunt 100 × LA 422 cross fit an additive-dominance model for all three traits. The VF 145B-7879 × LA 422 cross data best fit a model that included the additive × additive and additive × dominance interaction components for the trait diseased fruit, whereas higher-order epistatic models would have to be invoked to fit the data for the traits symptom rating and lesion size. A minimum of one gene segregated for all three traits. Broad-sense heritability estimates ranged from 0.09 to 0.16 for all three traits, indicating that selection for improved resistance to blackmold will require selection on a family performance basis.     

On the origin of Spanish two-rowed barleys

To investigate the phylogenetic origin of Spanish two-rowed barleys, we studied 44 accessions of old land-races both morphologically and biochemically to ascertain their similarity with 51 entries of old cultivars and land-races of widespread origin across Europe. They were also compared with 20 accessions of Hordeum spontaneum from the Mediterranean basin and other regions of its distribution range, 14 accessions of Moroccan cultivated six-rowed barley land-races, and different six-rowed Spanish and two-and six-rowed European cultivars. CM-(trypsin inhibitors and subunits of the barley tetrameric -amylase inhibitor) proteins and hordeins, all of which are endosperm proteins, were used as biochemical markers. The appearance of separate clusters of the Spanish barleys in the numerical classifications for both protein systems as a result of the existence of characteristic gene combinations that do not exist in entries from other origins permitted us to postulate the existence of local ancestors for most of the Spanish two-rowed barleys studied, and, therefore, a possible in situ domestication.     

Dynamic approaches to the mechanism of photosynthesis

An account of the author's life and scientific research is presented. Two main lines of research have been pursued: (1) Studies on the physiological aspect of photosynthesis started from experiments with crops under field conditions and then extended to the study of photosynthesis in nature; and (2) studies on the mechanism of photophosphorylation and related problems which began with the measurement of quantum requirement of photophosphorylation. This work led to the discovery of the high energy state of phosphorylation and many other interesting findings. In recent years, efforts have been made to study the operation and regulation of photosynthetic apparatus with a view to link the above-mentioned lines of research together.Written at the invitation of Govindjee.     

Vanadium levels in urine and cystine levels in fingernails and hair of exposed and normal persons

Vanadium was determined by radiochemical neutron activation analysis (RNAA) with proven accuracy in urine of workers occupationally exposed to vanadium-rich dust in a vanadium pentoxide production plant, and values in the range of 3.02–762 ng/mL (median 33.0 ng/mL) were found. In a control group consisting of administrative workers of the plant, urinary vanadium levels were found in the range of 1.05–53.4 ng/mL (median 2.53 ng/mL), whereas in an another control group of occupationally nonexposed persons, these values amounted to 0.066–0.489 ng/mL (median 0.212 ng/mL). Accuracy of the results was tested by analysis of reference material IAEA A-13 Animal Blood and NIST SRM-1515 Apple Leaves, and very good agreement was found with literature and the NIST certified values, respectively. Unlike urine, no significant differences were found for cystine levels in fingernails and hair of exposed and control persons.     

Role of Liv-52 in protection against beryllium intoxication

An ayurvedic medicine, Liv-52, was studied as a prophylactic agent against beryllium-induced toxicity in rats. Administration of berylliumper se caused severe degenerative and necrotic changes in kidneys, liver, and uterus. Beryllium exposure also reduced glycogen content, activities of alkaline phosphatase, succinate-dehydrogenase, and adenosine-triphosphatase in these organs. On the contrary, activities of acid phosphatase and glucose-6-phosphatase showed marginal increase. Liv-52-primed rats exhibited comparatively less marked toxic effects.     

Metabolic shift analysis at high cell densities

Abstract: In high cell density cultures it is virtually inevitable that the environment to which the cells are exposed is heterogeneous. Thus, with suspended cultures, individual cells are subject to temporal changes in their environment whereas with aggregated or immobilized cells, the culture can be considered as being formed by a number of subpopulations, each with its own environmental characteristics. In addition, in a high cell density environment, high concentrations of end products may negatively influence the growth rate. This may result in the selection of organisms with an altered metabolic behaviour or with a decreased sensitivity to the adverse effects of the product. We discuss the consequences of this heterogeneity with regard to carbon source metabolism in view of the ability of many bacterial species to adapt to environmental conditions. Selection of variant organisms was found to occur with Clostridium butyricum when grown for a prolonged time in a medium containing approx. I-50 mM glucose. In contrast to the original strain, these variants could sustain a high maximal growth rate in the presence of butyric acid. In addition, they had acquired the capacity to spontaneously form aggregates and were able to carry out a completely solventogenic fermentation. Heterogeneous metabolic activity in aggregated cells is demonstrated with cultures of Lactobacillus laevolacticus , an aggregateforming lactic acid bacterium that converts glucose completely to o-lactate. By using microelectrodes, we show that the fraction of metabolically active cells decreases with increasing aggregate size: in larger aggregates steep pH gradients occur with the effect that only the outer layer of the aggregate is metabolically active, i.e. contributes to lactic acid formation, whereas with smaller aggregates all cells remain active. As a result, the net specific lactic acid production rate of the population as a whole is not invariably increased with increased aggregate size.