Standing and flowing: the complex origins of adaptive variation

A population faced with a new selection pressure can only adapt if appropriate genetic variation is available. This genetic variation might come from new mutations or from gene exchange with other populations or species, or it might already segregate in the population as standing genetic variation (which might itself have arisen from either mutation or gene flow). Understanding the relative importance of these sources of adaptive variation is a fundamental issue in evolutionary genetics (Orr & Betancourt 2001 ; Barrett & Schluter 2008 ; Gladyshev et al. 2008 ) and has practical implications for conservation, plant and animal breeding, biological control and infectious disease prevention (e.g. Robertson 1960 ; Soulé & Wilcox 1980 ; Prentis et al. 2008 ; Pennings 2012 ). In this issue of Molecular Ecology, Roesti et al. ( 2014 ) make an important contribution to this longstanding debate.     

<Emphasis Type="Italic">Drosophila</Emphasis>-based <Emphasis Type="Italic">in vivo</Emphasis> assay for the validation of inhibitors of the epidermal growth factor receptor/Ras pathway

Overexpression of epidermal growth factor receptor (EGFR) is a common phenomenon observed in most cancers. Clinical treatment of such cancer involves the use of chemotherapeutic agents such as gefitinib and erlotinib which are inhibitors of tyrosine kinase (TK). These small molecules bind to the ATP-binding sites of the TK domain of EGFR. Our in silico analysis suggests that the TK domains of Drosophila and human EGFR are highly conserved. We therefore employed the Drosophila system to validate the in silico observations made with two important anticancer drugs. Since a large number of mutant flies are available, it was possible to investigate the various components of the EGFR/Ras/Raf/ MAPK pathways and the phosphorylation status of diphosphorylated extracellular signal-regulated kinase (dp-ERK1/2). These studies confirm the binding of the anilinoquinazolines to the Drosophila EGFR protein and modulation of its activity. Thus, Drosophila appears to be a robust and simple model system for screening newer anticancer drugs that act as TK inhibitors (TKIs). Electronic Supplementary Material  Supplementary material is available for this article at and is accessible for authorized users. Supplementary figures and tables pertaining to this article are available on the Journal of Biosciences Website at http://www.ias.ac.in/jbiosci/dec2008/pp731-742-suppl.pdf     

Inhibition of the yeast-mycelial transition and the phorogenesis of Mucorales by diamino butanone

Diamino butanone (DAB), a competitive inhibitor of ornithine decarboxylase (ODC) a key enzyme in polyamine biosynthesis, inhibited the yeast to hyphae transition in Mucor rouxii, induced by transfer from anaerobiosis to aerobiosis, but not the opposite phenomenon. Addition of DAB to anaerobic yeast cells brought about a decrease in ODC and polyamine levels. In these conditions, the aerobic shift produced only a weak increase in ODC activity and no change in polyamine levels. DAB also blocked phorogenesis in M. rouxii and in Phycomyces blakesleeanus. At the effective concentrations DAB did not affect cell growth of either fungus. It is suggested that low, constant levels of ODC and polyamines are necessary for cell growth, and that high transient levels are required during the differentiative steps. DAB, at the concentrations used, affects this last process, but does not interfere with the maintenance level of polyamines.Abbreviations ODC ornithine decarboxylase - DAB 1,4-diamino butanone     

Inferring the Pattern of Spontaneous Mutation from the Pattern of Substitution in Unitary Pseudogenes of Mycobacterium leprae and a Comparison of Mutation Patterns Among Distantly Related Organisms

The pattern of spontaneous mutation can be inferred from the pattern of substitution in pseudogenes, which are known to be under very weak or no selective constraint. We modified an existing method (Gojobori T, et al., J Mol Evol 18:360, 1982) to infer the pattern of mutation in bacteria by using 569 pseudogenes from Mycobacterium leprae. In Gojobori et al.’s method, the pattern is inferred by using comparisons involving a pseudogene, a conspecific functional paralog, and an outgroup functional ortholog. Because pseudogenes in M. leprae are unitary, we replaced the missing paralogs by functional orthologs from M. tuberculosis. Functional orthologs from Streptomyces coelicolor served as outgroups. We compiled a database consisting of 69,378 inferred mutations. Transitional mutations were found to constitute more than 56% of all mutations. The transitional bias was mainly due to C→T and G→A, which were also the most frequent mutations on the leading strand and the only ones that were significantly more frequent than the random expectation. The least frequent mutations on the leading strand were A→T and T→A, each with a relative frequency of less than 3%. The mutation pattern was found to differ between the leading and the lagging strands. This asymmetry is thought to be the cause for the typical chirochoric structure of bacterial genomes. The physical distance of the pseudogene from the origin of replication (ori) was found to have almost no effect on the pattern of mutation. A surprising similarity was found between the mutation pattern in M. leprae and previously inferred patterns for such distant taxa as human and Drosophila. The mutation pattern on the leading strand of M. leprae was also found to share some common features with the pattern inferred for the heavy strand of the human mitochondrial genome. These findings indicate that taxon-specific factors may only play secondary roles in determining patterns of mutation. Electronic Supplementary Material Electronic Supplementary material is available for this article at and accessible for authorised users. [Reviewing Editor:Dr. Dmitri Petrov]     

Polyamines in growth and dimorphism ofParacoccidioides brasiliensis

Putrescine and spermidine were the only polyamines found inParacoccidioides brasiliensis, a dimorphic fungus pathogenic for humans. Free polyamines (putrescine>spermidine) increased during the first 24 h of yeast growth, with a second peak at 42 h, and also during the first 12 h of mycelium-to-yeast transition (spermidine>putrescine). Conjugated and bound polyamines were also quantified. 1,4-Diamino-2-butanone decreased free putrescine and spermidine accumulation by inhibiting the activity of ornithine decarboxylase. The increase in free polyamines corresponds to bud emergence in yeast growth and to the mycelium-to-yeast transition ofP. brasiliensis.Abbreviations DAB 1,4-Diamino-2-butanone - Y Yeasts - M Mycelia - ODC Ornithine decarboxylase     

Unmarked gene deletion and host–vector system for the hyperthermophilic crenarchaeon Sulfolobus islandicus

Sulfolobus islandicus is being used as a model for studying archaeal biology, geo-biology and evolution. However, no genetic system is available for this organism. To produce an S. islandicus mutant suitable for genetic analyses, we screened for colonies with a spontaneous pyrEF mutation. One mutant was obtained containing only 233 bp of the original pyrE sequence in the mutant allele and it was used as a host to delete the β-glycosidase (lacS) gene. Two unmarked gene deletion methods were employed, namely plasmid integration and segregation, and marker replacement and looping out, and unmarked lacS mutants were obtained by each method. A new alternative recombination mechanism, i.e., marker circularization and integration, was shown to operate in the latter method, which did not yield the designed deletion mutation. Subsequently, Sulfolobus–E. coli plasmid shuttle vectors were constructed, which genetically complemented ΔpyrEFΔlacS mutation after transformation. Thus, a complete set of genetic tools was established for S. islandicus with pyrEF and lacS as genetic markers.     

Sulfur oxidation in mutants of the photosynthetic green sulfur bacterium Chlorobium tepidum devoid of cytochrome c-554 and SoxB

A mutant devoid of cytochrome c-554 (CT0075) in Chlorobium tepidum (syn. Chlorobaculum tepidum) exhibited a decreased growth rate but normal growth yield when compared to the wild type. From quantitative determinations of sulfur compounds in media, the mutant was found to oxidize thiosulfate more slowly than the wild type but completely to sulfate as the wild type. This indicates that cytochrome c-554 would increase the rate of thiosulfate oxidation by serving as an efficient electron carrier but is not indispensable for thiosulfate oxidation itself. On the other hand, mutants in which a portion of the soxB gene (CT1021) was replaced with the aacC1 cassette did not grow at all in a medium containing only thiosulfate as an electron source. They exhibited partial growth yields in media containing only sulfide when compared to the wild type. This indicates that SoxB is not only essential for thiosulfate oxidation but also responsible for sulfide oxidation. An alternative electron carrier or electron transfer path would thus be operating between the Sox system and the reaction center in the mutant devoid of cytochrome c-554. Cytochrome c-554 might function in any other pathway(s) as well as the thiosulfate oxidation one, since even green sulfur bacteria that cannot oxidize thiosulfate contain a cycA gene encoding this electron carrier.     

The SOS induction of umu'-'lacZ fusion gene by oxidative damage is influenced by polyamines in Escherichia coli

We report that polyamines have an effect on the SOS response of the umu operon in polyamine-deficient mutant and wild-type Escherichia coli strains carrying the umu'-'lacZ fusion. H₂O₂ effectively induces umu'-'lacZ in the wild type, but not significantly in the mutant strain. Exogenous polyamines did not restore the umu induction in the mutant to the wild-type level. In logarithmically growing cells, the basal expression of umu gene in the mutant is about five times higher than that of the wild type. The addition of polyamines to the growth medium markedly reduces the basal expression to the wild-type level. This reduction is due not to growth rate but to the polyamine itself. Our results suggest that polyamines are essentially involved in the SOS induction of the umu operon in E. coli.     

Isopentenyl-diphosphate isomerase is essential for viability of <Emphasis Type="Italic">Caenorhabditis elegans</Emphasis>

Homozygosity for a mutation in the idi-1 gene of Caenorhabditis elegans results in paralysis during the first larval stage, followed by an arrest of growth and development late in the first larval stage. Apoptotic corpses, which are apparently the result of normal programmed cell death, persist in the arrested larvae. In genetic mosaics, an additional defect becomes evident upon examination with Nomarski optics: cells that are genotypically mutant enlarge, and their cytoplasm becomes dimpled. Electron microscopy indicates that the dimpling reflects an accumulation of many enlarged lysosomes and autophagosomes. The mosaics demonstrate that the lethal mutation acts cell autonomously with respect to this vesicular abnormality and that there is a maternal effect with respect to the time of developmental arrest of mutant progeny. Cloning of the gene reveals that it is the only gene in C. elegans for isopentenyl-diphosphate isomerase, an enzyme that is important for the synthesis of lipophilic molecules, including farnesyl and geranyl diphosphates.     

Levels of endogenous polyamines and NaCl-inhibited growth of rice seedlings

The role of endogenous polyamines in the control of NaCl-inhibited growth of rice seedlings was investigated. Putrescine, spermidine and spermine were all present in shoots and roots of rice seedlings. NaCl treatment did not affect spermine levels in shoots and roots. Spermidine levels in shoots and roots were increased with increasing concentrations of applied NaCl. NaCl at a concentration of 50 mM, which caused only slight growth inhibition, drastically lowered the level of putrescine in shoots and roots. Addition of precursors of putrescine biosynthesis (L-arginine and L-ornithine) resulted in an increase in putrescine levels in NaCl-treated shoots and roots, but did not allow recovery of the growth inhibition of rice seedlings induced by NaCl. Pretreatment of rice seeds with putrescine caused an increase in putrescine level in shoots, but could not alleviate the inhibition effect of NaCl on seedling growth. The current results suggest that endogenous polyamines may not play a significant role in the control of NaCl-inhibited growth of rice seedlings.Abbreviations PUT putrescine - SPD spermidine - SPM spermine     

The novel 5bp deletion polymorphism in the promoter region of bovine <Emphasis Type="Italic">ACRP30</Emphasis> gene

ACRP30 gene was located nearby the QTL affecting the marbling, ribeye muscle area and fat thickness on the BAT1 in Angus. In this study, a 5bp deletion mutation within the bovine ACRP30 gene was firstly detected and confirmed in 991 cattle by PCR-SSCP, DNA sequencing and direct PCR amplification. The deletion mutation was appeared in Qinchuan, Nanyang, Jiaxian and Hasake, but was not found in Jinnan, Chinese Holsteins and Angus. The association of the deletion polymorphism with growth traits (including birth weight, body weight, average daily gain and body sizes in different growth periods (6/12/18/24 month-old)) was analyzed in 224 Nanyang cattle. No signification association of the deletion polymorphism with growth traits were observed (P > 0.05). The deletion was located in the promoter region and it resulted in a new putative CCAAT/enhancer binding protein-β response element (C/EBP-RE).     

Genetic quality and offspring performance in Chinook salmon: implications for supportive breeding

Each year salmon and other fishes are caught and used for supportive breeding programs that attempt to augment natural populations that are threatened with extinction. These programs typically mate individuals randomly and as such they overlook the importance of genetic quality to offspring fitness and ultimately to ensuring population health. Here, we use Chinook salmon (Oncorhynchus tshawytscha) and a fully crossed quantitative genetic breeding design to partition genetic variance in offspring performance (growth and survival) to additive and non-additive genetic effects as well as maternal effects. We show that these three effects contribute about equally to the variation in survival, but only non-additive genetic and maternal effects contribute to variation in growth. Some of the genetic effects could be assigned to variation at the class IIB locus of the major histocompatibility complex, but the maternal effects were not associated with egg size and we found no relationship between dam phenotypic measures and offspring survival or growth. We also found no relationship between sire sexually selected characters and offspring survival or growth, which is inconsistent with a “good genes” hypothesis. Finally, we show that incorporation of genetic quality into supportive breeding programs can increase offspring growth or survival by between 3% and 19% during the endogenous feeding stage alone, and projections to adulthood suggest that survivorship could be over four fold higher. Electronic Supplementary Material  Supplementary material is available in the online version of this article at and is accessible for authorised users.     

Somatic embryogenesis of Panax ginseng in liquid cultures: a role for polyamines and their metabolic pathways

A callus with embryogenic capacity was generated fromroot sections of Panax ginseng and used as aninoculum source for embryogenic liquid cultures in athree-step process: – a suspension culture of cellaggregates in the presence of an auxin/cytokininmixture, – an induction medium containing auxin only(for 5 to 30 days), – a regeneration medium containingcytokinin only (for one month). Up to 25 embryos wererecovered per 2.5 g of aggregates in these conditions.Incorporation of polyamines or their precursorsarginine and ornithine into either the induction orregeneration media increased the number of embryosproduced by up to 4 times. Inhibitors of bothbiosynthesis and biodegradation of polyamines reducedthe number of embryos. These results support earlierfindings of the role of polyamines in the process ofsomatic embryogenesis. The success of these liquidcultures opens up the possibility of producing somaticembryos of Panax ginseng in bioreactors.     

Erythrocyte-based Pig-a gene mutation assay: Demonstration of cross-species potential

Glycosylphosphatidylinositol (GPI) anchors attach specific proteins to the cell surface of hematopoietic cells. Of the genes required to form GPI anchors, only Pig-a is located on the X-chromosome. Prior work with rats suggests that the GPI anchor deficient phenotype is a reliable indicator of Pig-a mutation [Bryce et al., Environ. Mol. Mutagen., 49 (2008) 256–264]. The current report extends this line of investigation by describing simplified blood handling procedures, and by testing the assay principle in a second species, Mus musculus. With this method, erythrocytes are isolated, incubated with anti-CD24-PE, and stained with SYTO 13. Flow cytometric analyses quantify GPI anchor-deficient erythrocytes and reticulocytes. After reconstruction experiments with mutant-mimicking cells demonstrated that the analytical performance of the method is high, CD-1 mice were treated on three occasions with 7,12-dimethyl-1,2-benz[a]anthracene (DMBA, 75 mg/kg/day) or ethyl-N-nitrosourea (ENU, 40 mg/kg/day). Two weeks after the final treatment, DMBA-treated mice were found to exhibit markedly elevated frequencies of GPI anchor deficient erythrocytes and reticulocytes. For the ENU experiment, blood specimens were collected at weekly intervals over a 5-week period. Whereas the frequencies of mutant reticulocytes were significantly elevated 1 week after the last administration, the erythrocyte population was unchanged until the second week. Thereafter, both populations exhibited persistently elevated frequencies for the duration of the experiment (mean frequency at termination = 310 × 10⁻⁶ and 523 × 10⁻⁶ for erythrocyte and reticulocyte populations, respectively). These data provide evidence that Pig-a mutation does not convey an appreciable positive or negative cell survival advantage to affected erythroid progenitors, although they do suggest that affected erythrocytes have a reduced lifespan in circulation. Collectively, accumulated data support the hypothesis that flow cytometric enumeration of GPI anchor deficient erythrocytes and/or reticulocytes represents an effective in vivo mutation assay that is applicable across species of toxicological interest.     

Effect of Cadmium on Cellular Viability in Two Species of Microalgae (Scenedesmus sp. and Dunaliella viridis)

We determined the effect of several concentrations of cadmium (0, 5, 10, and 20 μg/l) on cellular viability in the microalgae Scenedesmus sp. and Dunaliella viridis, by measuring growth at 0, 24, 48, 72, and 96 h and pigment production at 10 days. Algae were obtained from the Nonvascular Plant Laboratory collection, in the Facultad Experimental de Ciencias, Universidad del Zulia, Venezuela. Growth was measured by cellular counting, while pigment content was evaluated using conventional spectrophotometric techniques. Growth of both species decreased in the exposed cultures comparing with the control, but its behavior was similar, because in both control and exposed cultures, its was observed an adaptive phase in the first hours, as well as a growth phase after 72 h. Cadmium concentrations above 10 μg/l produced an adverse effect on pigment production, depending on the concentration and/or exhibition time. However, even though cadmium inhibited growth and pigment production, levels of both parameters indicated cellular viability, demonstrating the adaptability of the algae cultures when they were exposed to the metal.     

Bottom-up controls on a mixed-species HAB assemblage: A comparison of sympatric Chattonella subsalsa and Heterosigma akashiwo (Raphidophyceae) isolates from the Delaware Inland Bays, USA

Recent novel mixed blooms of several species of toxic raphidophytes have caused fish kills and raised health concerns in the highly eutrophic Inland Bays of Delaware, USA. The factors that control their growth and dominance are not clear, including how these multi-species HAB events can persist without competitive exclusion occurring. We compared and contrasted the relative environmental niches of sympatric Chattonella subsalsa and Heterosigma akashiwo isolates from the bays using classic Monod-type experiments. C. subsalsa grew over a temperature range from 10 to 30 °C and a salinity range of 5–30 psu, with optimal growth occurring from 20 to 30 °C and 15 to 25 psu. H. akashiwo had similar upper temperature and salinity tolerances but also lower limits, with growth occurring from 4 to 30 °C and 5 to 30 psu and optimal growth between 16 and 30 °C and 10 and 30 psu. These culture results were confirmed by field observations of bloom occurrences in the Inland Bays. Maximum nutrient-saturated growth rates (μ_max) for C. subsalsa were 0.6 d⁻¹ and half-saturation concentrations for growth (K_s) were 9 μM for nitrate, 1.5 μM for ammonium, and 0.8 μM for phosphate. μ_max of H. akashiwo (0.7 d⁻¹) was slightly higher than C. subsalsa, but K_s values were nearly an order of magnitude lower at 0.3 μM for nitrate, 0.3 μM for ammonium, and 0.2 μM for phosphate. H. akashiwo is able to grow on urea but C. subsalsa cannot, while both can use glutamic acid. Cell yield experiments at environmentally relevant levels suggested an apparent preference by C. subsalsa for ammonium as a nitrogen source, while H. akashiwo produced more biomass on nitrate. Light intensity affected both species similarly, with the same growth responses for each over a range from 100 to 600 μmol photons m⁻² s⁻¹. Factors not examined here may allow C. subsalsa to persist during multi-species blooms in the bays, despite being competitively inferior to H. akashiwo under most conditions of nutrient availability, temperature, and salinity.