Primordial germ cell proliferation in the salamander Pleurodeles waltl: genetic control before gonadal differentiation

PGC counts were carried out on larvae of Pleurodeles waltl (urodele amphibia) issued from standard, monosexual male and monosexual female offspring while the genital ridges were settling. During this period, which is characterized by a zero mitotic index (and is therefore called the Po period), and which lasts from stage 35 to stage 41, no PGC proliferation occurs. A statistical analysis indicated that PGC counts per larva are sex genotype independent and that offspring may be divided into three groups with average PGC counts of 96.9, 51.0 and 31.1, respectively. A fourth group with an average of 18.3 PGCs has been identified using experimental larvae reared at 30 degrees C from stage 30. The PGC count of 96.9 would result from at least three mitotic cycles. Before the Po period, germ cells are not identifiable. A hypothesis concerning genetic control of PGC proliferation before Po was deduced from this analysis.     

Life history patterns of parental and hybrid Daphnia differ between lakes

1. We investigated whether Daphnia galeata × hyalina hybrids of Lake Constance and Lake Greifensee show the same pattern of life history parameters as previously reported for D. galeata × cucullata hybrids and whether such a pattern is consistent between Daphnia populations from those two lakes. 2. Hybrids in Lake Constance were intermediate in size compared with the parental species. Hybrids in Lake Greifensee were smaller than D. galeata. The intrinsic growth rate (r) of hybrids from Lake Constance was not significantly different from the faster growing parental taxon D. galeata. However, r of hybrids from Lake Greifensee was significantly lower than that of D. galeata. 3. The observed juvenile body length differences between the taxa varied with the clutch number. The first clutch juvenile lengths of the three taxa did not differ for Lake Constance. First clutch juveniles of Lake Greifensee D. galeata were smaller than hybrid first clutch juveniles. The third clutch juvenile length did not differ between taxa from Lake Greifensee, but D. galeata juveniles from Lake Constance were bigger than those of D. hyalina. 4. The life history pattern found in Lake Constance corresponds to previous findings from other studies. The hybrids in this lake combine the faster population growth of one parental species with a relatively small size. In the case of Lake Greifensee hybrids, the relatively large size of first clutch juveniles and the small size of the adults could be interpreted as dual adaptations to invertebrate and fish predation. We speculate that the lower population growth rate of the hybrids is a trade‐off for this twofold protection.     

Tests for gene clustering.

Comparing chromosomal gene order in two or more related species is an important approach to studying the forces that guide genome organization and evolution. Linked clusters of similar genes found in related genomes are often used to support arguments of evolutionary relatedness or functional selection. However, as the gene order and the gene complement of sister genomes diverge progressively due to large scale rearrangements, horizontal gene transfer, gene duplication and gene loss, it becomes increasingly difficult to determine whether observed similarities in local genomic structure are indeed remnants of common ancestral gene order, or are merely coincidences. A rigorous comparative genomics requires principled methods for distinguishing chance commonalities, within or between genomes, from genuine historical or functional relationships. In this paper, we construct tests for significant groupings against null hypotheses of random gene order, taking incomplete clusters, multiple genomes, and gene families into account. We consider both the significance of individual clusters of prespecified genes and the overall degree of clustering in whole genomes.     

EPR and multi-field magnetisation of reduced forms of the binuclear iron centre in ribonucleotide reductase from mouse

Mouse ribonucleotide reductase is composed of a 1?:?1 complex of two homodimeric subunits and catalyses the first unique step on the biochemical pathway to DNA synthesis. The small subunit, protein R2, contains dinuclear iron-oxygen clusters and a tyrosyl free radical required for catalytic activity. We have studied the mixed valent and fully reduced forms of the diiron oxygen cluster from mouse R2 protein by low-temperature EPR. EPR signals of the mixed-valent states of proteins R2 reconstituted with ferrous iron and oxygen in normal and deuterated water, using the same buffers, show apparent g values of 1.92, 1.73, and 1.60 for the mixed-valent state in H₂O and 1.93, 1.73, and 1.62 in D₂O. These g values are typical for diiron-oxygen proteins, while the effect of D₂O is unprecedented for this class of proteins. We estimate the coupling constant J for the Heisenberg exchange (H?=?2J*S₁*S₂) to be J?=?–7.5±1?cm^–1 for the mixed-valent form. The diferrous R2 protein shows an integer spin EPR signal in the presence of azide or 20% glycerol. Variable temperature variable field saturation magnetisation measurements show that only in the azide-complexed R2 protein does a weak ferromagnetic coupling occur (J?=?0.26±0.05?cm^–1), while R2 protein in the absence or presence of 20% glycerol contains non-coupled mononuclear ferrous iron (S?=?2) sites.     

A second L-type isozyme of potato glucan phosphorylase: cloning,antisense inhibition and expression analysis

In potato tubers two starch phosphorylase isozymes, types L and H, have been described and are believed to be responsible for the complete starch breakdown in this tissue. Type L has been localized in amyloplasts, whereas type H is located within the cytosol. In order to investigate whether the same isozymes are also present in potato leaf tissue a cDNA expression library from potato leaves was screened using a monoclonal antibody recognizing both isozyme forms. Besides the already described tuber L-type isozyme a cDNA clone encoding a second L-type isozyme was isolated. The 3171 nucleotide long cDNA clone contains an uninterrupted open reading frame of 2922 nucleotides which encodes a polypeptide of 974 amino acids. Sequence comparison between both L-type isozymes on the amino acid level showed that the polypeptides are highly homologous to each other, reaching 81–84% identity over most parts of the polypeptide. However the regions containing the transit peptide (amino acids 1–81) and the insertion sequence (amino acids 463–570) are highly diverse, reaching identities of only 22.0% and 29.0% respectively.Northern analysis revealed that both forms are differentially expressed. The steady-state mRNA levels of the tuber L-type isozyme accumulates strongly in potato tubers and only weakly in leaf tissues, whereas the mRNA of the leaf L-type isozyme accumulates in both tissues to the same extent. Constitutive expression of an antisense RNA specific for the leaf L-type gene resulted in a strong reduction of starch phosphorylase L-type activity in leaf tissue, but had only sparse effects in potato tuber tissues. Determination of the leaf starch content revealed that antisense repression of the starch phosphorylase activity has no significant influence on starch accumulation in leaves of transgenic potato plants. This result indicated that different L-type genes are responsible for the starch phosphorylase activity in different tissues, but the function of the different enzymes remains unclear.     

The response of tomato roots (Lycopersicon esculentum Mill.) to iron deficiency stress: alterations in the pattern of protein synthesis

Dicotyledon plants adapt to iron (Fe) deficiency through a seriesof reactions that increase the ability of the plant to assimilateFe and to increase the efficiency of Fe utilization. In an attemptto gain an insight into these adaptive processes, the specificchanges in protein synthesis associated with the onset of theFe deficiency response in tomato roots (Lycopersicon esculentumMill cv. Rutgers) have been investigated. Roots were grown underFe—sufficient and —deficient conditions, and thepattern of protein synthesis was analysed by in vitro translationof root mRNA and by in vivo labelling of root proteins. Polypeptideswere resolved by two—dimensional polyacrylamide gel elec—trophoresis.Seven polypeptides were identified by in vitro translation,whose synthesis was significantly increased during Fe deficiency.The increase was probably specific to Fe deficiency in thatthe polypep—tide synthesis was not increased during phosphatedeficiency stress, was less prominent following prolonged Fedeficiency and was decreased following re—supply of Feto the hydroponic medium. The pattern of in vitro translation of mRNA isolated from Fe—deficientroots was compared to the results obtainedin vivo followingradiolabelling of proteins. In these analyses, eight polypeptideswere identified, tentatively including the seven polypeptidespreviously identified by in vitro translation. All polypeptideswere characterized with regard to molecular mass and pl andtheir localization in the cell, whether being membrane boundor soluble. It is suggested that members of this group of polypeptidesare involved in the response of the root to Fe deficiency: althoughtheir functions remain to be identified. Key words: In vitro protein synthesis, iron, iron deficiency, root, 2-dimensional PAGE     

Ribonuclease P from plant nuclei and photosynthetic organelles

RNase P consists of both protein and RNA subunits in all organisms and organelles investigated so far, with the exception of chloroplasts and plant nuclei where no enzyme-associated RNA has been detected to date. Studies on substrate specificity revealed that cleavage by plant nuclear RNase P is critically dependent on a complete and intact structure of the substrate. No clearcut answer is yet possible regarding the order of processing events at the 5 or 3 end of tRNAs in the case of nuclear or chloroplast processing enzymes. RNase P from a phylogenetically ancient photosynthetic organelle will be discussed in greater detail: The enzyme from theCyanophora paradoxa cyanelle is the first RNase P from a photosynthetic organelle which has been shown to contain an essential RNA subunit. This RNA is strikingly similar to its counterpart from cyanobacteria, yet it lacks catalytic activity. Properties of the holoenzyme suggest an intermediate position in RNA enzyme evolution, with an eukaryotic-type, inactive RNA and a prokaryotic-type small protein subunit. The possible presence of an RNA component in RNase P from plant nuclei and modern chloroplasts will be discussed, including a critical evaluation of some criteria that have been frequently applied to elucidate the subunit composition of RNase P from different organisms.Abbreviations RNase P Ribonuclease P - (pre-)tRNA transfer ribonucleic acid (precursor) - tRNA ^Ser (- ^Tyr , - ^Phe ) transfer ribonucleic acid specific for serine (tyrosine, phenylalanine) - CyRP RNA RNA component of cyanelle RNase P     

Location by fluorescence microscopy of glycosidases and a xylanase in the anaerobic gut fungi Caecomyces communis,Neocallimastix frontalis,and Piromyces rhizinflata

-d-Glucosidase, -d-fucosidase -d-xylosidase, and -cellobiopyranosidase activities in Caecomyces communis, Neocallimastix frontalis, and Piromyces rhizinflata, located with fluorescent conjugates, occur throughout the whole thallus as from zoospore germination and disappear before sporulation. -d-Galactosidase and -l-arabinopyranosidase activities are low or nonexistent. A xylanase, detected by indirect immunofluorescence, was observed at the surface of the vegetative cells, vesicles, or rhizoids. Cross-reactions prove the existence of analogies in structure among the enzymes of these anaerobic gut fungi.     

Growth of Nitrosomonas europaea on hydroxylamine

Abstract Hydroxylamine is an intermediate in the oxidation of ammonia to nitrite, but until now it has not been possible to grow Nitrosomonas europaea on hydroxylamine. This study demonstrates that cells of N. europaea are capable of growing mixotrophically on ammonia and hydroxylamine. The molar growth yield on hydroxylamine (4.74 g mol⁻¹ at a growth rate of 0.03 h⁻¹) was higher than expected. Aerobically growing cells of N. europaea oxidized ammonia to nitrite with little loss of inorganic nitrogen, while significant inorganic nitrogen losses occurred when cells were growing mixotrophically on ammonia and hydroxylamine. In the absence of oxygen, hydroxylamine was oxidized with nitrite as electron acceptor, while nitrous oxide was produced. Anaerobic growth of N. europaea on ammonium, hydroxylamine and nitrite could not be observed at growth rates of 0.03 h⁻¹ and 0.01 h⁻¹.