bluuebunny/crossref_metadata_2025 huggingface dataset:
Population systematics of the snake genus Naja (Reptilia: Serpentes: Elapidae) in Indochina: Multivariate morphometrics and comparative mitochondrial DNA sequencing (cytochrome oxidase I)
Authors: Wüster W. , Thorpe R. S. , J. Cox M. , Jintakune P. , Nabhitabhata J.
Published: December 2004

We analyze the population systematics of Asiatic cobras in Indochina, China and the Andaman Islands by means of comparative sequencing of the cytochrome oxidase subunit I gene of the mitochondrial DNA molecule and multivariate analysis of morphological characters. Canonical variate analysis and mtDNA sequence information reveal that the cobras of this region comprise four distinct species: Naja atra from China and northern Vietnam, Naja kaouthia from Burma, central Thailand, Cambodia and southern Vietnam, Naja siamensis from Thailand, Cambodia and southern Vietnam, and Naja sagittifera from the Andaman Islands. The subspecies N. kaouthia suphanensis Nutaphand 1986 shows no mtDNA sequence difference from typical N. kaouthia from central Thailand, and multivariate analysis does not reveal differences in general phenotypic profile; the subspecies is therefore synonymised with Naja kaouthia. The cytochrome oxidase subunit I gene, little used in molecular taxonomy, is shown to be well suited for studies at the species level, as it shows taxonomically useful levels of interspecific divergence but low levels of intraspecific variation; this is particularly relevant for studies of rare species, where sample size is a problem. The combination of multivariate morphometrics and molecular systematics can be particularly powerful in resolving systematic problems in such cases.
Inbreeding depression in Lychnis flos‐cuculi (Caryophyllaceae): effects of different levels of inbreeding
Authors: Hauser Thure P. , Loeschcke Volker
Published: December 2004

In a controlled crossing experiment on Lychnis flos‐cuculi plants in the greenhouse, outbred and selfed maternal plants were each treated with pollen from unrelated plants, siblings and selves. The seeds thus obtained had expected inbreeding coefficients of 0, 0.25 and 0.5 for the outbred maternal plants, and 0, 0.5 and 0.75 for the selfed maternal plants. Seed abortion rate, seed weight and germination rate were estimated. Seedlings were transplanted to an outdoor garden, and monitored for survival, probability of flowering, number of capsules and area of capsules next spring. Inbred seeds germinated slower and in lower proportions than those less inbred, and seedlings had lower survival, flowering, fruit set and area of capsules if inbred.

Combined fitness values were estimated from the survival and fecundity components, and severe inbreeding depression was detected for these estimates (0.51 and 0.56 for one generation of selfing). The fitness function decreased linearly with the increase in inbreeding coefficient, which is as expected if the inbreeding depression is additive among loci.
Quantitative genetic analysis of cranial morphology in the cotton‐top (Saguinus oedipus) and saddle‐back (S. fuscicollis) tamarins
Authors: Cheverud J. M.
Published: December 2004

Patterns of morphological variation play an important role in evolutionary diversification and are critical to an informed interpretation of interspecific differences. When patterns of genetic variation have not diverged substantially, it is possible to reconstruct the differences in selection which gave rise to morphological differences among extant species. Morphological variation patterns are compared between two tamarin species, the cotton‐top tamarin (Saguinus oedipus) and the saddle‐back tamarin (S. fuscicollis illigeri).

Genetic, phenotypic, and environmental variance/covariance and correlation matrices were obtained for a series of 39 cranial characters in each species (cotton‐top tamarin, N = 328; saddle‐back tamarin, N = 209) and for the species combined using crania from individuals of known genealogical relationship. After accounting for the effects of estimation error on measures of matrix similarity, patterns of phenotypic, genetic, and environmental variation and correlation were found to be very similar across species and among the types of variance within species. Taking the saddle‐back tamarins as the standard, cotton‐top tamarins have been selected for an enlarged anterior temporalis attachment area and increased facial prognathism. In primates, an enlarged anterior temporalis muscle is associated with incisive food preparation, especially at wide gape.
Ecological and morphological diversification within single species and character displacement in Mandarina, endemic land snails of the Bonin Islands
Authors: Chiba S.
Published: December 2004

The endemic land snail species Mandarina hahajimana has undergone extensive habitat and morphological diversification within the Hahajima islands in the Bonin archipelago. This species has diversified into populations with ground, arboreal and semi‐arboreal life histories. In addition, arboreal populations and semi‐arboreal populations show diversification in preferences of species and positions of the tree on which they are found. Shell morphologies of M. hahajimana exhibit remarkable geographical variation, and they have a clear relationship with their life histories. The morphological variation of M. hahajimana results from adaptation to different lifestyles. The habitats of these populations influence the relationships with other species of Mandarina coexisting with M. hahajimana. This suggests that the morphological and ecological divergence within M. hahajimana has been induced by competitive interaction with other species of Mandarina. Character displacement may have played an important role in promoting adaptive radiation of Mandarina in the Bonin Islands.
The evolutionary consequences of niche construction: a theoretical investigation using two‐locus theory
Authors: Laland K. N. , Odling‐Smee F. J. , Feldman M. W.
Published: December 2004

This paper addresses the joint evolution of environment‐altering (niche constructing) traits, and traits whose fitness depends on alterable sources of natural selection in environments. We explore the evolutionary consequences of this niche construction using a two‐locus population genetic model. The novel conclusions are that niche construction can (1) cause evolutionary inertia and momentum, (2) lead to the fixation of otherwise deleterious alleles, (3) support stable polymorphisms where none are expected, (4) eliminate what would otherwise be stable polymorphisms, and (5) influence disequilibrium. The results suggest that the changes that organisms bring about in their niche can themselves be an important source of natural selection pressures, and imply that evolution may proceed in cycles of selection and niche construction.
Laboratory evolution of longevity in the bean weevil (Acanthoscelides obtectus)
Authors: Tucić N. , Gliksman I. , Šešlija D. , Milanović D. , Mikuljanac S. , Stojković O.
Published: December 2004

In this study we examined the effects of long‐term selection on early and late reproduction in the bean weevil. The pure lines and the hybrids between the lines within a selection regime were compared for longevity, early and late female fecundity, male mating ability, pre‐adult developmental time and wet adult weight. Comparison of hybrid with pure lines provided some evidence for inbreeding despression in the lines from both selection regimes. We found that virgin and mated adults of both sexes from the “old” lines lived longer than “young” line beetles. Comparisons of the hybrid “young” with hybrid “old” lines revealed a trade‐off between early and late fecundity of females. For noncompetitive mating ability of males there was no difference between the lines with different rates of senescence when the males were young. But, when the males were older, beetles from the lines selected for delayed senescence expressed superior mating ability. In addition, the “old” line beetles take longer to develop and are heavier than those from “young” line beetles. Although these data suggest that shorter pre‐adult developmental time may imply more rapid senescence, there is the possibility of inadvertent selection for rapid development in the “young” lines and this complicates the interpretation of the observed trade‐off between the pre‐adult and adult performances.
Adaptive significance of environmental sex determination in an amphipod
Authors: McCabe J. , Dunn A. M.
Published: December 2004

Environmental sex determination (ESD) permits adaptive sex choice under patchy environmental conditions, where the environment affects sex‐specific fitness and where offspring can predict their likely adult status by monitoring an appropriate environmental cue. For Gammarus duebeni, an amphipod with ESD, it has been proposed that this flexible sex determination system is adaptive because males gain more from large size. Under ESD, young which are born earlier in the season become mostly males and, experiencing longer to grow, are therefore larger at breeding than females which are born later in the season. In order to test the hypothesis that ESD is adaptive for this species we investigated the relationship between size and fitness for both males and females, in a population of G. duebeni known to have ESD. We measured size related pairing success and fecundity, and used these two measures to calculate the relative fitness gains achieved through an increase in size for either sex. The fitness of both males and females increased with size, but males gained more from an increase in size than did females, throughout the breeding season. The data support the adaptive explanation for the evolution and maintenance of ESD in this species.
Reaction norms of Arabidopsis. V. Flowering time controls phenotypic architecture in response to nutrient stress
Authors: Pigliucci M. , Schlichting C. D.
Published: December 2004

The evolutionary and environmental stability of character correlations has increasingly been the focus of ecological and quantitative genetic studies. Although the genetic stability of character correlations is a central assumption of quantitative genetic models of phenotypic evolution, theoretical considerations suggest that both the genetic and the phenotypic architecture should change in response to selection and to environmental heterogeneity. We investigate genetic (population) differences and plasticity to nutrient availability of the phenotypic architecture describing the whole‐plant phenotype of Arabidopsis thaliana (Brassicaceae). We found significant genetic differences among early and late flowering ecotypes in the relationships between several traits, when a path‐analytical model was used to estimate character correlations. Furthermore, we found significant plasticity of several path coefficients when nutrient levels were altered. A whole‐plant analysis considering all paths in the model simultaneously confirmed that populations of A. thaliana are characterized by distinct phenotypic architectures, and that these are altered in different ways by environmental changes. We discuss the implications of these findings for our understanding of selective pressure on and response by multivariate phenotypes.
The genetics of phenotypic plasticity. VII. Evolution in a spatially‐structured environment
Authors: Scheiner Samuel M.
Published: December 2004

Previous models of the evolution of phenotypic plasticity have, for the most part, not considered the effects of genetic architecture and spatial structure. I examine those factors with an individual‐based simulation model. With regard to genetic architecture, I considered how the presence of different types of loci would affect medium‐term evolutionary outcomes. The types of loci differed in how the environment determined phenotypic expression and included loci that were insensitive to the environment (non‐plastic loci), sensitive in a linear fashion, and sensitive in a quadratic fashion (both plastic loci). With regard to spatial structure, I investigated the affects of migration patterns. These simulations demonstrated that two general conditions are necessary for phenotypic plasticity to be selected. (1) The environment must have a strong influence on genotypic expression. (2) The between‐generation changes in the environment must be large and predictable, in the current instance because of migration in a spatially‐structured (clinal) environment. Responses to selection were not simple, however. Rarely were pure strategies — genetic specialization or phenotypic plasticity — selected for. Instead, the existence of multiple types of loci led to mixed genetic outcomes. The result of this mixed outcome were individuals with reaction norms that were less steep than the optimal reaction norm (when non‐plastic and linear‐plastic loci were present) or individuals with curved reaction norms when the optimal reaction norms was linear (when all three types of loci were present). A pure plasticity strategy had the highest global fitness because plastic individuals would match the optimal phenotype everywhere. The reason that the metapopulation did not achieve this global fitness optimum is that local selection is stronger than global selection. Each deme is driven to a local fitness peak based on the combined, locally additive effects of the non‐plastic and plastic loci. Plasticity is only selected globally, so plasticity becomes more highly favored with high migration rates. This effect was greatest in parts of the cline where the plasticity loci were not being expressed and, thus, not locally selected upon. That is, in these demes local selection was weak or absent allowing global fitness effects to predominate.
Physiological and metabolic aspects of experimental heterochrony in colonial hydroids
Authors: Blackstone N. W.
Published: December 2004

Hydractiniid hydroids display a range of morphological variation from sheet‐like to runner‐like forms, thus exemplifying the patterns of heterochrony found in many colonial animals. Experimental studies of clonal replicates of a Podocoryne carnea colony demonstrate a relationship between these heterochronic traits and physiological and metabolic factors. While colonies exposed to dilute solutions of 2, 4‐dinitrophenol (an uncoupler of oxidative phosphorylation) for 4 hours per day show a strong dose‐response relationship between within‐colony gastrovascular flow rate and the timing of morphological development, colonies exposed for 20 hours per day fail to show this relationship. Spectrofluorometric measures of redox state suggest that the degree of “loose‐coupling” of oxidative phosphorylation is proportional to the exposure time. Thus colonies exposed for 20 hours have significantly diminished ATP production relative to colonies exposed for 4 hours. Two underlying factors may be responsible for the breakdown of the dose‐response relationship between flow rate and development: (1) colonies exposed to a high dose of uncoupler for 20 hours per day allocate a larger‐than‐normal share of their total energy budget into maintaining the minimal flow rate required physiologically, and (2) colonies' redox state has a direct effect on the timing of morphological development. The latter effect is particularly intriguing, given the extensive literature relating “metabolic activity” to development. In general, these results suggest a relationship between energy allocation, metabolism, and development in these hydroids, and these interrelationships may be central to the evolutionary process of heterochrony.
Partial local mate competition and the sex ratio: A study on non‐pollinating fig wasps
Authors: West S. A. , Herre E. A.
Published: December 2004

In many species, mating takes place in temporary patches where only a small number of females produce offspring. In this situation Local Mate Competition (LMC) theory predicts that the optimal sex ratio (defined as proportion males) should become increasingly female biased as the number of females contributing offspring to a patch decreases. However, in a large number of these species, some mating is also likely to occur away from the natal patch (termed partial LMC). In this case the degree of LMC is reduced, and theory predicts a relatively less female biased sex ratio. We tested these two predictions with field data from 17 species of New World non‐pollinating fig wasps representing three genera. We present a model which suggests that the average number of females ovipositing in a fruit (i.e. patch) is positively correlated with the proportion of fruit of a given tree species in which that species of wasp occurs. Across species, the overall sex ratio was positively correlated with the proportion of fruit in which that species occurs. Furthermore, the males of some species are wingless, and in these species all mating must take place before females disperse from their natal fruit. In contrast, the males of other species are winged, and in these species mating may also take place away from the natal fruit. Species with winged males had less female biased sex ratios than species with wingless males that occurred in a similar proportion of fruit. Finally, the correlation between sex ratio and the proportion of fruit in which a species occurs was also observed within species when comparing between the fruit crops of different trees. This suggests that individual females facultatively adjust the sex ratio of their offspring in response to variable LMC.
Enhanced expression of Mcm proteins in cancer cells derived from uterine cervix
Authors: Ishimi Yukio , Okayasu Isao , Kato Chieko , Kwon Hyun‐Ju , Kimura Hiroshi , Yamada Kouichi , Song Si‐Young
Published: December 2004

Minichromosome maintenance proteins (Mcm) 2–7 play essential roles in eukaryotic DNA replication. Several reports have indicated the usefulness of Mcm proteins as markers of cancer cells in histopathological diagnosis. However, their mode of expression and pathophysiological significance in cancer cells remain to be clarified. We compared the level of expression of Mcm proteins among human HeLa uterine cervical carcinoma cells, SV40‐transformed human fibroblast GM00637 cells and normal human fibroblast WI‐38 cells. All the proteins examined were detected in HeLa and GM cells at 6–10 times the level found in WI‐38 cells on average. This increase was observed both in total cellular proteins and in the chromatin‐bound fraction. Consistently, Mcm2 mRNA was enriched in HeLa cells to approximately four times the level in WI‐38 cells, and the synthesis of Mcm4, 6 and 7 proteins was accelerated in HeLa cells. Immunohistochemical studies of surgical materials from human uterine cervix showed that Mcm3 and 4 are ubiquitously expressed in cancer cells. Further, the positive rate and level of Mcm3 and 4 expression appeared to be higher in cancer cells than in normal proliferating cells of the uterine cervix and dysplastic cells, suggesting that they can be useful markers to distinguish these cells.
Regulation of pyruvate dehydrogenase complex activity in plant cells
Authors: Tovar‐Méndez Alejandro , Miernyk Jan A. , Randall Douglas D.
Published: December 2004

The pyruvate dehydrogenase complex (PDC) is subjected to multiple interacting levels of control in plant cells. The first level is subcellular compartmentation. Plant cells are unique in having two distinct, spatially separated forms of the PDC; mitochondrial (mtPDC) and plastidial (plPDC). The mtPDC is the site of carbon entry into the tricarboxylic acid cycle, while the plPDC provides acetyl‐CoA and NADH forde novofatty acid biosynthesis. The second level of regulation of PDC activity is the control of gene expression. The genes encoding the subunits of the mt‐ and plPDCs are expressed following developmental programs, and are additionally subject to physiological and environmental cues. Thirdly, both the mt‐ and plPDCs are sensitive to product inhibition, and, potentially, to metabolite effectors. Finally, the two different forms of the complex are regulated by distinct organelle‐specific mechanisms. Activity of the mtPDC is regulated by reversible phosphorylation catalyzed by intrinsic kinase and phosphatase components. An additional level of sensitivity is provided by metabolite control of the kinase activity. The plPDC is not regulated by reversible phosphorylation. Instead, activity is controlled to a large extent by the physical environment that exists in the plastid stroma.
Interaction of ostreolysin, a cytolytic protein from the edible mushroom Pleurotus ostreatus, with lipid membranes and modulation by lysophospholipids
Authors: Sepčić Kristina , Berne Sabina , Potrich Cristina , Turk Tom , Maček Peter , Menestrina Gianfranco
Published: December 2004

Ostreolysin is a 16‐kDa cytolytic protein specifically expressed in primordia and fruiting bodies of the edible mushroom Pleurotus ostreatus. To understand its interaction with lipid membranes, we compared its effects on mammalian cells, on vesicles prepared with either pure lipids or total lipid extracts, and on dispersions of lysophospholipids or fatty acids. At nanomolar concentrations, the protein lysed human, bovine and sheep erythrocytes by a colloid‐osmotic mechanism, compatible with the formation of pores of 4 nm diameter, and was cytotoxic to mammalian tumor cells. A search for lipid inhibitors of hemolysis revealed a strong effect of lysophospholipids and fatty acids, occurring below their critical micellar concentration. This effect was distinct from the capacity of ostreolysin to bind to and permeabilize lipid membranes. In fact, permeabilization of vesicles occurred only when they were prepared with lipids extracted from erythrocytes, and not with lipids extracted from P. ostreatus or pure lipid mixtures, even if lysophospholipids or fatty acids were included. Interaction with lipid vesicles, and their permeabilization, correlated with an increase in the intrinsic fluorescence and α‐helical content of the protein, and with aggregation, which were not detected with lysophospholipids. It appears that either an unknown lipid acceptor or a specific lipid complex is required for binding, aggregation and pore formation. The inhibitory effect of lysophospholipids may reflect a regulatory role for these components on the physiological action of ostreolysin and related proteins during fruiting.
Molecular characterization and allergenic activity of Lyc e 2 (β‐fructofuranosidase), a glycosylated allergen of tomato
Authors: Westphal Sandra , Kolarich Daniel , Foetisch Kay , Lauer Iris , Altmann Friedrich , Conti Amedeo , Crespo Jesus F. , Rodríguez Julia , Enrique Ernesto , Vieths Stefan , Scheurer Stephan
Published: December 2004

Until now, only a small amount of information is available about tomato allergens. In the present study, a glycosylated allergen of tomato (Lycopersicon esculentum), Lyc e 2, was purified from tomato extract by a two‐step FPLC method. The cDNA of two different isoforms of the protein, Lyc e 2.01 and Lyc e 2.02, was cloned into the bacterial expression vector pET100D. The recombinant proteins were purified by electroelution and refolded. The IgE reactivity of both the recombinant and the natural proteins was investigated with sera of patients with adverse reactions to tomato. IgE‐binding to natural Lyc e 2 was completely inhibited by the pineapple stem bromelain glycopeptide MUXF (Manα1–6(Xylβ1–2)Manβ1–4GlcNAcβ1–4(Fucα1–3)GlcNAc). Accordingly, the nonglycosylated recombinant protein isoforms did not bind IgE of tomato allergic patients. Hence, we concluded that the IgE reactivity of the natural protein mainly depends on the glycan structure. The amino acid sequences of both isoforms of the allergen contain four possible N‐glycosylation sites. By application of MALDI‐TOF mass spectrometry the predominant glycan structure of the natural allergen was identified as MMXF (Manα1–6(Manα1–3)(Xylβ1–2)Manβ1–4GlcNAcβ1–4(Fucα1–3) GlcNAc). Natural Lyc e 2, but not the recombinant protein was able to trigger histamine release from passively sensitized basophils of patients with IgE to carbohydrate determinants, demonstrating that glycan structures can be important for the biological activity of allergens.
In vitro analysis of the relationship between endonuclease and maturase activities in the bi‐functional group I intron‐encoded protein, I‐AniI
Authors: Geese William J. , Kwon Yong K. , Wen Xiaoping , Waring Richard B.
Published: December 2004

The AnCOB group I intron from Aspergillus nidulans encodes a homing DNA endonuclease called I‐AniI which also functions as a maturase, assisting in AnCOB intron RNA splicing. In this investigation we biochemically characterized the endonuclease activity of I‐AniI in vitro and utilized competition assays to probe the relationship between the RNA‐ and DNA‐binding sites. Despite functioning as an RNA maturase, I‐AniI still retains several characteristic properties of homing endonucleases including relaxed substrate specificity, DNA cleavage product retention and instability in the reaction buffer, which suggest that the protein has not undergone dramatic structural adaptations to function as an RNA‐binding protein. Nitrocellulose filter binding and kinetic burst assays showed that both nucleic acids bind I‐AniI with the same 1 : 1 stoichiometry. Furthermore, in vitro competition activity assays revealed that the RNA substrate, when prebound to I‐AniI, stoichiometrically inhibits DNA cleavage activity, yet in reciprocal experiments, saturating amounts of prebound DNA substrate fails to inhibit RNA splicing activity. The data suggest therefore that both nucleic acids do not bind the same single binding site, rather that I‐AniI appears to contain two binding sites.
Functional expression of the quinoline 2‐oxidoreductase genes (qorMSL) in Pseudomonas putida KT2440 pUF1 and in P. putida 86‐1 Δqor pUF1 and analysis of the Qor proteins
Authors: Frerichs‐Deeken Ursula , Goldenstedt Birgit , Gahl‐Janßen Renate , Kappl Reinhard , Hüttermann Jürgen , Fetzner Susanne
Published: December 2004

The availability of a system for the functional expression of genes coding for molybdenum hydroxylases is a prerequisite for the construction of enzyme variants by mutagenesis. For the expression cloning of quinoline 2‐oxidoreductase (Qor) from Pseudomonas putida 86 – that contains the molybdopterin cytosine dinucleotide molybdenum cofactor (Mo‐MCD), two distinct [2Fe−2S] clusters and FAD – the qorMSL genes were inserted into the broad host range vector, pJB653, generating pUF1. P. putida KT2440 and P. putida 86‐1 Δqor were used as recipients for pUF1. Whereas Qor from the wild‐type strain showed a specific activity of 19–23 U·mg−1, the specific activity of Qor purified from P. putida KT2440 pUF1 was only 0.8–2.5 U·mg−1, and its apparent kcat (quinoline) was about ninefold lower than that of wild‐type Qor. The apparent Km values for quinoline were similar for both proteins. UV/visible and EPR spectroscopy indicated the presence of the full set of [2Fe−2S] clusters and FAD in Qor from P. putida KT2440 pUF1, however, the very low intensity of the Mo(V)‐rapid signal, that occurs in the presence of quinoline, as well as metal analysis indicated a deficiency of the molybdenum center. In contrast, the metal content, and the spectroscopic and catalytic properties of Qor produced by P. putida 86‐1 Δqor pUF1 were essentially like those of wild‐type Qor. Release of CMP upon acidic hydrolysis of the Qor proteins suggested the presence of the MCD form of the pyranopterin cofactor; the CMP contents of the three enzymes were similar.
Alpha‐oxidation of 3‐methyl‐substituted fatty acids and its thiamine dependence
Authors: Casteels Minne , Foulon Veerle , Mannaerts Guy P. , Van Veldhoven Paul P.
Published: December 2004

3‐Methyl‐branched fatty acids, as phytanic acid, undergo peroxisomal α‐oxidation in which they are shortened by 1 carbon atom. This process includes four steps: activation, 2‐hydroxylation, thiamine pyrophosphate dependent cleavage and aldehyde dehydrogenation. The thiamine pyrophosphate dependence of the third step is unique in peroxisomal mammalian enzymology. Human pathology due to a deficient alpha‐oxidation is mostly linked to mutations in the gene coding for the second enzyme of the sequence, phytanoyl‐CoA hydroxylase.
Isocitrate dehydrogenase of Plasmodium falciparum
Authors: Wrenger Carsten , Müller Sylke
Published: December 2004

Erythrocytic stages of the malaria parasite Plasmodium falciparum rely on glycolysis for their energy supply and it is unclear whether they obtain energy via mitochondrial respiration albeit enzymes of the tricarboxylic acid (TCA) cycle appear to be expressed in these parasite stages. Isocitrate dehydrogenase (ICDH) is either an integral part of the mitochondrial TCA cycle or is involved in providing NADPH for reductive reactions in the cell. The gene encoding P. falciparum ICDH was cloned and analysis of the deduced amino‐acid sequence revealed that it possesses a putative mitochondrial targeting sequence. The protein is very similar to NADP+‐dependent mitochondrial counterparts of higher eukaryotes but not Escherichia coli. Expression of full‐length ICDH generated recombinant protein exclusively expressed in inclusion bodies but the removal of 27 N‐terminal amino acids yielded appreciable amounts of soluble ICDH consistent with the prediction that these residues confer targeting of the native protein to the parasites' mitochondrion. Recombinant ICDH forms homodimers of 90 kDa and its activity is dependent on the bivalent metal ions Mg2+ or Mn2+ with apparent Km values of 13 µm and 22 µm, respectively. Plasmodium ICDH requires NADP+ as cofactor and no activity with NAD+ was detectable; the for NADP+ was found to be 90 µm and that of d‐isocitrate was determined to be 40 µm. Incubation of P. falciparum under exogenous oxidative stress resulted in an up‐regulation of ICDH mRNA and protein levels indicating that the enzyme is involved in mitochondrial redox control rather than energy metabolism of the parasites.
Purple membrane lipid control of bacteriorhodopsin conformational flexibility and photocycle activity
Authors: Hendler Richard W. , Barnett Steven M. , Dracheva Swetlana , Bose Salil , Levin Ira W.
Published: December 2004

Specific lipids of the purple membrane of Halobacteria are required for normal bacteriorhodopsin structure, function, and photocycle kinetics [Hendler, R.W. & Dracheva, S. (2001) Biochemistry (Moscow)66, 1623–1627]. The decay of the M‐fast intermediate through a path including the O intermediate requires the presence of a hydrophobic environment near four charged aspartic acid residues within the cytoplasmic loop region of the protein (R. W. Hendler & S. Bose, unpublished results). On the basis of the unique ability of squalene, the most hydrophobic purple membrane lipid, to induce recovery of M‐fast activity in Triton‐treated purple membrane, we proposed that this uncharged lipid modulates an electrostatic repulsion between the membrane surface of the inner trimer space and the nearby charged aspartic acids of the cytoplasmic loop region to promote transmembrane α‐helical mobility with a concomitant increase in the speed of the photocycle. We examined Triton‐treated purple membranes in various stages of reconstitution with native lipid suspensions using infrared spectroscopic techniques. We demonstrate a correlation between the vibrational half‐width parameter of the protein α‐helical amide I mode at 1660 cm−1, reflecting the motional characteristics of the transmembrane helices, and the lipid‐induced recovery of native bacteriorhodopsin properties in terms of the visible absorbance maxima of ground state bacteriorhodopsin and the mean decay times of the photocycle M‐state intermediates.
No evidence for a role in signal‐transduction of Na+/K+‐ATPase interaction with putative endogenous ouabain
Authors: Hansen Otto
Published: December 2004

A cascade of events (signal‐transduction), mainly seen in rat cardiac myocytes and renal cells, is thought to occur after ouabain interaction with a minor fraction of Na+/K+‐ATPase. A higher intracellular Na+ concentration followed sodium pump inhibition by ouabain with a subsequent gradual increase or oscillations in intracellular Ca2+ concentration. Whether this increase in intracellular Ca2+ concentration is part of the cascade, a result of the cascade or a totally independent phenomenon are conflicting interpretations that are discussed. At best, however, the cascade is initiated by ouabain concentrations several orders of magnitude higher than the measured plasma concentrations of putative endogenous ouabain. The experimentally high ouabain concentration may be critical for another reason. Most tissues contain various isoforms of the catalytic α‐peptide of Na+/K+‐ATPase with an individual sublocalization and, in rats, with different ouabain‐sensitivity. The almost ouabain‐insensitive α1‐isoform of Na+/K+‐ATPase is essentially unaffected by the high ouabain concentration, whereas ouabain‐sensitive α‐isoforms, possibly confined to membrane structures near cytosolic microdomains and Na+/Ca2+ exchangers, may be totally blocked. Classifying endogenous ouabain as a physiological inducer of the signaling system on this background seems hazardous.
Human anionic trypsinogen
Authors: Kukor Zoltán , Tóth Miklós , Sahin‐Tóth Miklós
Published: December 2004

Human pancreatic secretions contain two major trypsinogen isoforms, cationic and anionic trypsinogen, normally at a ratio of 2 : 1. Pancreatitis, pancreatic cancer and chronic alcoholism lead to a characteristic reversal of the isoform ratio, and anionic trypsinogen becomes the predominant zymogen secreted. To understand the biochemical consequences of these alterations, we recombinantly expressed and purified both human trypsinogens and documented characteristics of autoactivation, autocatalytic degradation and Ca2+‐dependence. Even though the two trypsinogens are ≈ 90% identical in their primary structure, we found that human anionic trypsinogen and trypsin exhibited a significantly increased (10–20‐fold) propensity for autocatalytic degradation, relative to cationic trypsinogen and trypsin. Furthermore, in contrast to the characteristic stimulation of the cationic proenzyme, acidic pH inhibited autoactivation of anionic trypsinogen. In mixtures of cationic and anionic trypsinogen, an increase in the proportion of the anionic proenzyme had no significant effect on the levels of trypsin generated by autoactivation or by enterokinase at pH 8.0 in 1 mm Ca2+– conditions that were characteristic of the pancreatic juice. In contrast, rates of trypsinogen activation were markedly reduced with increasing ratios of anionic trypsinogen under conditions that were typical of potential sites of pathological intra‐acinar trypsinogen activation. Thus, at low Ca2+ concentrations at pH 8.0, selective degradation of anionic trypsinogen and trypsin caused diminished trypsin production; while at pH 5.0, inhibition of anionic trypsinogen activation resulted in lower trypsin yields. Taken together, the observations indicate that up‐regulation of anionic trypsinogen in pancreatic diseases does not affect physiological trypsinogen activation, but significantly limits trypsin generation under potential pathological conditions.
The expression of retinoblastoma and Sp1 is increased by low concentrations of cyclin‐dependent kinase inhibitors
Authors: Peñuelas Silvia , Alemany Cristina , Noé Véronique , Ciudad Carlos J.
Published: December 2004

We examined the effect of suboptimal concentrations of cyclin‐dependent kinase inhibitors, which do not interfere with cell proliferation, on retinoblastoma expression in hamster (Chinese hamster ovary K1) and human (K562 and HeLa) cells. To achieve this, we used the chemical inhibitors roscovitine and olomoucine (which inhibit CDK2 preferentially), UCN‐01 (which also inhibits CDK4/6) and p21 (as an intrinsic inhibitor). All chemical inhibitors and overexpression of p21 strongly induced retinoblastoma protein expression. UCN‐01‐mediated retinoblastoma expression was caused by an increase in both the levels of retinoblastoma mRNA and the stability of the protein. The expression of the transcription factor Sp1, a retinoblastoma‐interacting protein, was also enhanced by all the cyclin‐dependent kinase inhibitors tested. However, Sp1 expression was caused by an increase in the levels of Sp1 mRNA without modification in the stability of the protein. By using luciferase experiments, the transcriptional activation of both retinoblastoma and Sp1 promoters by UCN‐01 was confirmed. Bisindolylmaleimide I, at concentrations causing a similar or higher inhibition of protein kinase C than UCN‐01, provoked a lower activation of retinoblastoma and Sp1 expression. Finally, the effects of cyclin‐dependent kinase inhibitors on dihydrofolate reductase gene expression were evaluated. Treatment with UCN‐01 increased cellular dihydrofolate reductase mRNA levels, and dihydrofolate reductase enzymatic activity was enhanced by UCN‐01, roscovitine, olomoucine and p21, in transient transfection experiments. These results support a mechanism for the self‐regulation of retinoblastoma expression, and point to the need to establish the appropriate dose of cyclin‐dependent kinase inhibitors as antiproliferative agents in anticancer treatments.
Salicylic acid and the hypersensitive response initiate distinct signal transduction pathways in tobacco that converge on the as‐1‐like element of the PR‐1a promoter
Authors: Grüner Rose , Strompen Georg , Pfitzner Artur J. P. , Pfitzner Ursula M.
Published: December 2004

Tobacco pathogenesis‐related protein 1a (PR‐1a) is induced in plants during the hypersensitive response (HR) after exposure of plants to salicylic acid (SA) and by developmental cues. Gene activation by these diverse stimuli is mediated via an as‐1‐like element in the PR‐1a upstream region. To further analyze the significance of this cis‐acting sequence, an authentic as‐1 element from the cauliflower mosaic virus 35S RNA promoter was inserted into the PR‐1a promoter in place of the as‐1‐like motif. Reporter gene analysis in transgenic tobacco plants demonstrated that as‐1 can functionally replace the as‐1‐like element in the PR‐1a promoter in response to all stimuli. However, reporter gene induction from the as‐1 carrying promoter was enhanced in response to SA compared to the wild‐type promoter, and the ratio of reporter gene activities in SA treated leaf tissue to tissue exhibiting the HR increased with the as‐1 promoter construct. Our findings support a model where PR‐1a gene expression relies on at least two distinct signal transduction pathways initiated by SA and by a yet unknown signal produced during the HR, that promote different, albeit related, transcription complexes on the PR‐1a as‐1‐like element. Analysis of PR‐1 proteins in plants expressing salicylate hydroxylase yielded additional evidence that an HR dependent pathway leads to high level PR‐1 gene induction in tobacco.
Acyl‐CoA dehydrogenases
Authors: Ghisla Sandro , Thorpe Colin
Published: December 2004

Acyl‐CoA dehydrogenases constitute a family of flavoproteins that catalyze the α,β‐dehydrogenation of fatty acid acyl‐CoA conjugates. While they differ widely in their specificity, they share the same basic chemical mechanism of α,β‐dehydrogenation. Medium chain acyl‐CoA dehydrogenase is probably the best‐studied member of the class and serves as a model for the study of catalytic mechanisms. Based on medium chain acyl‐CoA dehydrogenase we discuss the main factors that bring about catalysis, promote specificity and determine the selective transfer of electrons to electron transferring flavoprotein. The mechanism of α,β‐dehydrogenation is viewed as a process in which the substrate αC‐H and βC‐H bonds are ruptured concertedly, the first hydrogen being removed by the active center base Glu376‐COO– as an H+, the second being transferred as a hydride to the flavin N(5) position. Hereby the pKa of the substrate αC‐H is lowered from > 20 to ≈ 8 by the effect of specific hydrogen bonds. Concomitantly, the pKa of Glu376‐COO– is also raised to 8–9 due to the decrease in polarity brought about by substrate binding. The kinetic sequence of medium chain acyl‐CoA dehydrogenase is rather complex and involves several intermediates. A prominent one is the molecular complex of reduced enzyme with the enoyl‐CoA product that is characterized by an intense charge transfer absorption and serves as the point of transfer of electrons to the electron transferring flavoprotein. These views are also discussed in the context of the accompanying paper on the three‐dimensional properties of acyl‐CoA dehydrogenases.
Genetic defects in fatty acid β‐oxidation and acyl‐CoA dehydrogenases
Authors: Gregersen Niels , Bross Peter , Andresen Brage S.
Published: December 2004

Mitochondrial fatty acid oxidation deficiencies are due to genetic defects in enzymes of fatty acid β‐oxidation and transport proteins. Genetic defects have been identified in most of the genes where nearly all types of sequence variations (mutation types) have been associated with disease. In this paper, we will discuss the effects of the various types of sequence variations encountered and review current knowledge regarding the genotype–phenotype relationship, especially in patients with acyl‐CoA dehydrogenase deficiencies where sufficient material exists for a meaningful discussion. Because mis‐sense sequence variations are prevalent in these diseases, we will discuss the implications of these types of sequence variations on the processing and folding of mis‐sense variant proteins. As the prevalent mis‐sense variant K304E MCAD protein has been studied intensively, the investigations on biogenesis, stability and kinetic properties for this variant enzyme will be discussed in detail and used as a paradigm for the study of other mis‐sense variant proteins. We conclude that the total effect of mis‐sense sequence variations may comprise an invariable – sequence variation specific – effect on the catalytic parameters and a conditional effect, which is dependent on cellular, physiological and genetic factors other than the sequence variation itself.
How Consistently are Good Parents Good Parents? Repeatability of Parental Care in the House Sparrow, Passer domesticus
Authors: Schwagmeyer P. L. , Mock Douglas W.
Published: December 2004

In many avian species, substantial individual variation occurs in parental food‐provisioning levels, which often is assumed to reflect variation in parental quality. Parental quality also has often been invoked as a key element in mate choice among biparental species, and many sexually‐selected traits have been investigated as potential predictors of parental quality. In recent studies of house sparrow (Passer domesticus) parents, we found that individuals behaved remarkably consistently across time, regardless of temporary manipulations of the nestling provisioning of their partners. This suggests that variation in parental competence may be attributable to quality differences among individuals. One prediction of the ‘parental quality differences’ hypothesis is that individuals also should show consistency in their provisioning behavior across broods. To test this, we compared the parental delivery rates of individual house sparrows across broods. Parents of both sexes reduced their per‐chick delivery rates as the season progressed; parents of both sexes were also responsive to changes in their brood sizes. Despite these sources of environmental variation in provisioning rates, the parental care of individual males was highly repeatable across broods. By contrast, female parental care showed extremely low repeatability, and standardized measures of among‐individual variation in parental behavior revealed females to be much less variable than males. These results indicate that females in this multi‐brooded species have much to gain from mate‐choice decisions predicated on male parental quality or accurate indicators of such, whereas males are less likely to profit from being highly selective about the ‘parental quality’ of their partners.
Flexible Helping Behaviour in the Azure‐Winged Magpie
Authors: Valencia Juliana , de la Cruz Carlos , González Beatriz
Published: December 2004

Helping to rear the offspring of others may be a way for younger birds to gain access to future reproduction especially when turnover of breeding opportunities is low. However, this explanation is not applicable to cases where adults also help, or when roles shift between helpers and breeders. Over a period of 6‐yr, we studied a marked population of azure‐winged Magpie (Cyanopica cyanus) breeding in a non‐territorial, colonial system. Magpies bred in a highly flexible cooperative system, in which individuals helped at different stages of the breeding cycle, including nest building, feeding the incubating female and feeding the young and removing the faecal sacs. On average, 50% of hatched nests were assisted by helpers‐at‐the‐nest, and nest success appeared to be positively related to the presence of helpers. Helpers were predominantly males. Although juveniles were more likely to help, both juvenile and adult birds helped. Individual birds behaved as helpers either as a first‐option or after having attempted their own breeding (second‐option helpers). An individual helper may assist more than one nest during the same breeding season and in different breeding seasons. Reversals between breeder and helper roles were common in both directions, within a breeding season and between years. Helping behaviour is an option for almost any member of the colony. Therefore, hypotheses related to the enhancement of future breeding opportunities for juveniles can be discarded as general explanation of helping in this species. Although the decision to help appeared to be influenced by proximal environmental conditions hindering successful breeding, the associated benefits of helping as opposed to simply recovering for future reproduction, especially for former breeders, deserve further study.
Population and Species Divergence of Chemical Cues that Influence Male Recognition of Females in Desmognathine Salamanders
Authors: Verrell Paul
Published: December 2004

Growing evidence indicates that males may be more discriminating of mating partners than often has been assumed. In the North American Ocoee dusky salamander, Desmognathus ocoee (Plethodontidae: Desmognathinae), sexual incompatibility among conspecific populations is high in encounters staged in the laboratory, at least in part because males fail to recognize ‘other’ females as appropriate targets for courtship. I used Y‐mazes to test the hypothesis that males of D. ocoee discriminate between substrate‐borne chemical cues produced by ‘own’ (homotypic) and ‘other’ (heterotypic) females. Males of four populations discriminated in favor of substrates soiled by homotypic females over clean (control) substrates (expt 1), suggesting that females produce chemical cues of sociosexual significance to males. Furthermore, males from these populations discriminated in favor of substrates soiled by homotypic females vs. substrates soiled by heterotypic females (expt 2), both conspecific and heterospecific (D. carolinensis and D. orestes). Thus, differences among populations and species in female chemical cues appear to affect the chemotactic responses of males. I suggest that, together with differences in behavioral signals and responses exhibited during courtship, differences in female chemical cues likely contribute to sexual incompatibility among populations and taxa of desmognathine salamanders.