Genomic structure,expression and evolution of the alfalfa aspartate aminotransferase genes

Genomic clones encoding two isozymes of aspartate aminotransferase (AAT) were isolated from an alfalfa genomic library and their DNA sequences were determined. The AAT1 gene contains 12 exons that encode a cytosolic protein expressed at similar levels in roots, stems and nodules. In nodules, the amount of AAT1 mRNA was similar at all stages of development, and was slightly reduced in nodules incapable of fixing nitrogen. The AAT1 mRNA is polyadenylated at multiple sites differing by more than 250 bp. The AAT2 gene contains 11 exons, with 5 introns located in positions identical to those found in animal AAT genes, and encodes a plastid-localized isozyme. The AAT2 mRNA is polyadenylated at a very limited range of sites. The transit peptide of AAT2 is encoded by the first two and part of the third exon. AAT2 mRNA is much more abundant in nodules than in other organs, and increases dramatically during the course of nodule development. Unlike AAT1, expression of AAT2 is significantly reduced in nodules incapable of fixing nitrogen. Phylogenetic analysis of deduced AAT proteins revealed 4 separate but related groups of AAT proteins; the animal cytosolic AATs, the plant cytosolic AATs, the plant plastid AATs, and the mitochondrial AATs.     

Nitrogen fixation by periphyton and plankton on the Amazon floodplain at Lake Calado

Nitrogen fixation by periphyton and plankton was measured on the Amazon flood-plain using the acetylene reduction method calibrated with¹⁵N-N₂. The average ratio (± SD) of moles C₂H₄ reduced per mole N₂-N fixed was 3.4 ± 0.7, similar to other studies. Periphyton and plankton had high rates of light-dependent nitrogen fixation, with dark nitrogen fixation averaging 26% of the average rates in the light. The average daily (24 h) rates for periphyton nitrogen fixation in 1989 and 1990 were 1.79 and 0.51 mmol N₂-N·m^–2·d^–1 respectively, which are comparable to summer rates in many temperate cyanobacterial assemblages. Nitrogen fixation was depressed at N0₃ ^– concentrations as low as 0.5 M, and was below detection limits at concentrations of 4 M, which occurred during periods of river flooding. Planktonic nitrogen fixation rates were high (0.5–0.8 mmol N₂-N·m^–2·d^–1) during the high-water and drainage phases of the annual hydrograph when the floodplain waters were draining towards the river (low NO₃ ^–), but rates were undetectable (< 0.05 mmol N₂-N·m^–2·d^–1) when there was river flooding (high NO₃ ^–). Nitrogen fixation by periphyton and plankton in 1989–1990 accounted for approximately 8% of previously reported total annual nitrogen inputs to the floodplain at Lake Calado.     

Cloning of Multiple Forms of Goldfish Vimentin: Differential Expression in CNS

Abstract: In efforts to determine the primary structure of intermediate filament proteins in the goldfish visual pathway, we isolated clones from a retinal λgt11 cDNA expression library that represent goldfish vimentin. We show that there are at least two forms of goldfish vimentin, designated as vimentin α and vimentin β. RNase protection assays indicate that vimentin α mRNA is expressed in low amounts in retina, optic nerve, and brain and in higher amounts in spinal cord. In contrast, vimentin β mRNA is expressed in low amounts in retina, optic nerve, brain, and spinal cord and in very high amounts in eye lens. Immunohistochemical studies show that in the optic nerve, vimentin α is mainly restricted to blood vessels, meninges, and septa. Light staining is observed with this antibody in an astrocytic glial pattern throughout the optic nerve. Two-dimensional gel analysis shows that all of these goldfish vimentins are low abundant components of optic nerve cytoskeletal preparations.     

Cloning and Expression of a 5-Hydroxytryptamine7 Receptor Positively Coupled to Adenylyl Cyclase

Abstract: In a number of different cell types, phosphorylation of a 63-kDa protein has been shown to increase rapidly in response to stimuli that lead to an increase in intracellular calcium. Here, a stimulus-sensitive protein at this molecular weight is identified in PC12 cells and rat cortical synaptosomes as phosphoglucomutase. In addition, the added phosphate is shown to be in an oligosaccharide terminating in phosphodiester-linked glucose. In synaptosomes, incorporated radioactivity, following incubation with [¹⁴C]glucose or the [β-³⁵S]phosphorothioate analogue of UDP-glucose, was found to increase within 5 s of stimulation and return to baseline within 25 s. Despite the many pathways utilizing glucose, this was the only detectable protein glycosylation observed in synaptosomes. These results indicate that cytoplasmic glycosylation is reversible and rapidly regulated, and suggest that phosphoglucomutase undergoes an alteration in function and/or topography in response to increases in intracellular calcium.     

Linkage analysis of sex determination in the honey bee (Apis mellifera)

A colony-level phenotype was used to map the major sex determination locus (designatedX) in the honey bee (Apis mellifera). Individual queen bees (reproductive females) were mated to single drones (fertile males) by instrumental insemination. Haploid drone progeny of an F1 queen were each backcrossed to daughter queens from one of the parental lines. Ninety-eight of the resulting colonies containing backcross progeny were evaluated for the trait low brood-viability resulting from the production of diploid drones that were homozygous atX. DNA samples from the haploid drone fathers of these colonies were used individually in polymerase chain reactions (PCR) with 10-base primers. These reactions generated random amplified polymorphic DNA (RAPD) markers that were analyzed for cosegregation with the colony-level phenotype. One RAPD marker allele was shared by 22 of 25 drones that fathered low brood-viability colonies. The RAPD marker fragment was cloned and partially sequenced. Two primers were designed that define a sequence-tagged site (STS) for this locus. The primers amplified DNA marker fragments that cosegregated with the original RAPD marker. In order to more precisely estimate the linkage betweenX and the STS locus, another group of bees consisting of progeny from one of the low-brood viability colonies was used in segregation analysis. Four diploid drones and 181 of their diploid sisters (workers, nonfertile females) were tested for segregation of the RAPD and STS markers. The cosegregating RAPD and STS markers were codominant due to the occurrence of fragment-length alleles. The four diploid drones were homozygous for these markers but only three of the 181 workers were homozygotes (recombinants). Therefore the distance betweenX and the STS locus was estimated at 1.6 cM. An additional linked marker was found that was 6.6 cM from the STS locus.     

The ecological significance of nickel hyperaccumulation: a plant chemical defense

Nickel hyperaccumulating plants have more than 1000 mg Ni kg^–1 dry weight when grown on nickel-bearing soils. We hypothesized that Ni hyperaccumulation could serve as a chemical defense against herbivores In feeding experiments with potential insect herbivores and Ni hyperaccumulating plants, only those inseets fed leaves from plants grown on non-nickel-bearing soil survived or showed a weight gain. Among chemical parameters measured, only Ni content of plants was sufficient to explain this result. When subjected to herbivory by lepidopteran larvae, plants grown on Ni-amended soil showed greater survival and yield than plants on unamended soil. Ni hyperaccumulation may be an effective plant chemical defense against herbivores because of its high lethality, apparent low cost, and broad spectrum of toxicity.     

Woody-tissue respiration for Simarouba amara and Minquartia guianensis,two tropical wet forest trees with different growth habits

We measured CO₂ efflux from stems of two tropical wet forest trees, both found in the canopy, but with very different growth habits. The species were Simarouba amara, a fast-growing species associated with gaps in old-growth forest and abundant in secondary forest, and Minquartia guianensis, a slow-growing species tolerant of low-light conditions in old-growth forest. Per unit of bole surface, CO₂ efflux averaged 1.24 mol m^–2 s^–1 for Simarouba and 0.83 mol m^–2s^–1 for Minquartia. CO₂ efflux was highly correlated with annual wood production (r ²=0.65), but only weakly correlated with stem diameter (r ²=0.22). We also partitioned the CO₂ efflux into the functional components of construction and maintenance respiration. Construction respiration was estimated from annual stem dry matter production and maintenance respiration by subtracting construction respiration from the instantaneous CO₂ flux. Estimated maintenance respiration was linearly related to sapwood volume (39.6 mol m^–3s^–1 at 24.6° C, r ²=0.58), with no difference in the rate for the two species. Maintenance respiration per unit of sapwood volume for these tropical wet forest trees was roughly twice that of temperate conifers. A model combining construction and maintenance respiration estimated CO₂ very well for these species (r ²=0.85). For our sample, maintenance respiration was 54% of the total CO₂ efflux for Simarouba and 82% for Minquartia. For our sample, sapwood volume averaged 23% of stem volume when weighted by tree size, or 40% with no size weighting. Using these fractions, and a published estimate of aboveground dry-matter production, we estimate the annual cost of woody tissue respiration for primary forest at La Selva to be 220 or 350 g C m^–2 year^–1, depending on the assumed sapwood volume. These costs are estimated to be less than 13% of the gross production for the forest.     

The evolutionary origin of the 35 kb circular DNA of Plasmodium falciparum: new evidence supports a possible rhodophyte ancestry

In common with other Apicomplexan parasites, Plasmodium falciparum carries two extrachromosomal DNAs, one of which, the 6 kb element, is undoubtedly mitochondrial. The second, generally referred to as the 35 kb circle, is of unknown provenance, but the nature and organization of its genetic content makes a mitochondrial association unlikely and the molecule has features reminiscent of plastid genomes. We now report the occurrence on the circle of an open reading frame specifying a predicted 470 amino acid protein that shares more than 50% identity with a gene currently known only on the plastome of red algae. This high degree of conservation confirms the 35 kb circle's plastid ancestry, and we speculate that it may have originated from the rhodoplast of an ancient red algal endosymbiont in the progenitor of the Apicomplexa.     

Partial Purification and Immunocytocharacterization of the Plasma Membrane ATPase of Jerusalem artichoke (Helianthus tuberosus L.) Tubers in Relation to Dormancy

Plasmalemma ATPase from Jerusalem artichoke tubers was studiedin relation to the dormancy of tubers. After partial purification,one peptide of 110 kDa appeared on SDS PAGE electrophoresisfrom dormant and non-dormant materials. ATPase specific activitywas twice higher on dormant material in the crude and solubilizedfractions, but was the same in both materials after partialpurification. Immunolabeling of this enzyme was made using aspecific antibody raised against the C terminal portion of theH⁺-ATPase from Arabidopsis thaliana. Immunolabeling was morepronounced in dormant material, in vitro and in situ. Severalworks had shown that the C terminal part of the enzyme couldbe involved in its regulation. The results presented are discussedin relation to the hypothesis according to which an internaleffector could modulated the plasmalemma ATPase activity, duringdormancy breaking. (Received October 25, 1993; Accepted September 6, 1994)