Naturally occurring variation in bristle number and DNA polymorphisms at the scabrous locus of Drosophila melanogaster

The association between quantitative genetic variation in bristle number and molecular variation at a candidate neurogenic locus, scabrous, was examined in Drosophila melanogaster. Approximately 32 percent of the genetic variation in abdominal bristle number (21 percent for sternopleural bristle number) among 47 second chromosomes from a natural population was correlated with DNA sequence polymorphisms at this locus. Several polymorphic sites associated with large phenotypic effects occurred at intermediate frequency. Quantitative genetic variation in natural populations caused by alleles that have large effects at a few loci and that segregate at intermediate frequencies conflicts with the classical infinitesimal model of the genetic basis of quantitative variation.     

Polygenic mutation in Drosophila melanogaster: genotype x environment interaction for spontaneous mutations affecting bristle number

A highly inbred line of Drosophila melanogaster was subdivided into replicate sublines that were subsequently maintained independently with 10 pairs of parents per generation. The parents were randomly sampled for 19 'unselected' sublines and artificially selected for high or low abdominal or sternopleural bristle number for 12 'selected' sublines (with 3 replicate selection lines/trait/direction of selection). Divergence in mean bristle number among the unselected sublines, and response of the selected sublines to selection, are attributable to the accumulation of new mutations affecting bristle number. The input of mutational variance per generation, VM, can be estimated from the magnitude of response or divergence, assuming neutrality of mutations affecting the bristle traits. We reared unselected lines at generations 222 and 224, and selected lines at generations 182-184 of mutation accumulation at each of three temperatures (18 degrees C, 25 degrees C, 28 degrees C), and estimated the mutational variance common to all environments and the mutational variance from genotype x environment interaction. For sternopleural bristle number, the mutational interaction variance was 26% of the mutational variance common to all temperatures, and the interaction variance was due to temperature x line interaction. For abdominal bristle number, the mutational interaction variance was 142% of the mutational variance common to all temperatures, and the interaction variance was due to interactions of temperature x line, sex x line, and temperature x sex x line. It is possible that segregating variation for bristle number is maintained partly by genotype x environment interaction, but information on the fitness profiles of mutations affecting bristle number in each environment will be necessary to evaluate this hypothesis quantitatively.     

The dynamics of neurogenic signalling underlying bristle development in Drosophila melanogaster

We have examined expression of the neurogenic gene, Delta (Dl), and the regulatory relationships between the Delta-Notch signalling pathway and the proneural gene, achaete, during microchaeta development in Drosophila. Delta is expressed in all microchaeta proneural cells and microchaeta sensory organ precursors (SOPs) and is expressed dynamically in SOP progeny. We find that Delta expression in microchaeta proneural cells is detected prior to the onset of achaete expression and arises normally in the absence of achaete/scute function, indicating that initial Delta expression in the notum is not dependent on proneural gene function. Activation of the Delta-Notch pathway results in loss of Delta protein accumulation, suggesting that Delta expression is regulated, in part, by Delta-Notch signalling activity. We find that Delta signalling is required for correct delineation of early proneural gene expression in developing nota. Within microchaeta proneural stripes, we demonstrate that Delta-Notch signalling prohibits adoption of the SOP fate by repressing expression of proneural genes.     

Evolutionary history of the sex-peptide (Acp70A) gene region in Drosophila melanogaster

A phosphorothioate oligonucleotide (PS-ODN) with sequence identical to the repeat sequence of the mammalian telomere, 5'-d(TTAGGG)-3', was incubated with a Burkitt's lymphoma-derived (OMA-BL1) cell line. This hexanucleotide inhibits telomerase activity in cell lysates, lengthens cell doubling time, and induces apoptosis. Concatenated repeats (12-, 18-, and 24-mers) of the 5'-d(TTAGGG)-3' motif induce similar cellular responses. Scrambled sequences do not efficiently inhibit telomerase activity or significantly alter cell growth and viability. The in vivo efficacy of this PS-ODN was evaluated in a xenograft human-nude mouse model. Once tumors were established these animals were administered the telomere mimic, 5'-d(TTAGGG)-3', a control scrambled sequence 5'-d(TGTGAG)-3', or saline for 14 days via a subcutaneous osmotic pumps in a blinded study monitoring tumor size with dose and time. A significant decrease in tumor size was observed in animals given 50 micrograms/mouse/day 5'-d(TTAGGG)-3', but not following 5'-d(TGTGAG)-3', relative to the saline-treated animals. The antitumor activity of the 6-mer telomere mimic demonstrated a dose dependency including a reduction in metastatic nodules in the spleen. No activity was observed with the scrambled controls. In addition to antitumor activity we observed an increase in the mouse hematopoietic progenitor cell populations, BFU-e and CFU-GM. These results demonstrated the effects of a short hexameric oligonucleotide telomere mimic in vitro and in vivo and the potential utility of short oligonucleotides as telomerase inhibitors.     

Nucleotide variation in the triosephosphate isomerase (Tpi) locus of Drosophila melanogaster and Drosophila simulans

DNA sequence variation in a 1.1-kb region including the coding portion of the Tpi locus was examined in 25 homozygous third-chromosome lines of Drosophila melanogaster, nine lines of Drosophila simulans, and one line of Drosophila yakuba. Our data show that the widespread allozyme polymorphism observed in cosmopolitan D. melanogaster is due to a glutamic acid substitution occurring in a phylogenetically conserved lysine that has been identified as part of the "hinged-lid" active site of the enzyme. This observation suggests that the replacement polymorphism may have important functional consequences. One replacement polymorphism was also observed in D. simulans, although its functional relevance is more difficult to assess, since it affects a site that is not strongly conserved. This amino acid change in D. simulans is associated with a single lineage possessing seven unique silent substitutions, which may be indicative of balancing selection or population subdivision. The absence of fixed amino acid differences between D. melanogaster and D. simulans and only a single difference with D. yakuba suggests that triose phosphate isomerase is under strong functional constraint. Silent variation is slightly higher for D. melanogaster than for D. simulans. Finally, we outline the general lack of evidence for old balanced polymorphisms at allozyme loci in D. melanogaster.     

Microsatellite variation in Drosophila melanogaster and Drosophila simulans: a reciprocal test of the ascertainment bias hypothesis

Interspecific comparisons of microsatellite loci have repeatedly shown that the loci are longer and more variable in the species from which they are derived (the focal species) than are homologous loci in other (nonfocal) species. There is debate as to whether this is due to directional evolution or to an ascertainment bias during the cloning and locus selection processes. This study tests these hypotheses by performing a reciprocal study. Eighteen perfect dinucleotide microsatellite loci identified from a Drosophila simulans library screen and 18 previously identified in an identical Drosophila melanogaster library screen were used to survey natural populations of each species. No difference between focal and nonfocal species was observed for mean PCR fragment length. However, heterozygosity and number of alleles were significantly higher in the focal species than in the nonfocal species. The most common allele in the Zimbabwe population of both species was sequenced for 31 of the 36 loci. The length of the longest stretch of perfect repeat units is, on average, longer in the focal species than in the non-focal species. There is a positive correlation between the length of the longest stretch of perfect repeats and heterozygosity. The difference in heterozygosity can thus be explained by a reduction in the length of the longest stretch of perfect repeats in the nonfocal species. Furthermore, flanking-sequence length difference was noted between the two species at 58% of the loci sequenced. These data do not support the predictions of the directional-evolution hypothesis; however, consistent with the ascertainment bias hypothesis, the lower variability in nonfocal species is an artifact of the microsatellite cloning and isolation process. Our results also suggest that the magnitude of ascertainment bias for repeat unit length is a function of the microsatellite size distribution in the genomes of different species.     

Quantitative genetic variation of odor-guided behavior in a natural population of Drosophila melanogaster

Quantitative genetic variation in behavioral response to the odorant, benzaldehyde, was assessed among a sample of 43 X and 35 third chromosomes extracted from a natural population and substituted into a common inbred background. Significant genetic variation among chromosome lines was detected. Heritability estimates for olfactory response, however, were low, as is typical for traits under natural selection. Furthermore, the loci affecting naturally occurring variation in olfactory response to benzaldehyde were not the same in males and females, since the genetic correlation between the sexes was low and not significantly different from zero for the chromosome 3 lines. Competitive fitness, viability and fertility of the chromosome 3 lines were estimated using the balancer equilibrium technique. Genetic correlations between fitness and odor-guided behavior were not significantly different from zero, suggesting the number of loci causing variation in olfactory response is small relative to the number of loci causing variation in fitness. Since different genes affect variation in olfactory response in males and females, genetic variation for olfactory response could be maintained by genotype x sex environment interaction. This unusual genetic architecture implies that divergent evolutionary trajectories for olfactory behavior may occur in males and females.     

Changes in gene expression during post-eclosional development in the olfactory system of Drosophila melanogaster

We have found that the expression of some genes in Drosophila melanogaster changes during the life of the adult fly. These changes can be illustrated by the use of enhancer trap lines which mark the expression of particular genes in the adult fly. Although the fly is considered able to perform most necessary adult functions within the first 72 h after eclosion from the pupal case, we find changes in expression over the first 10 days of life in the antennae of several of the genes we have examined. Some genes change by increasing from an initially low level of expression of the marked gene, while other lines, which we have termed 'late-onset' genes, show no expression of the marked gene until 4-5 days following eclosion. In contrast, some genes decrease their expression during the first 10 days of life. The changes in gene expression seen over the first 10 days of the fly's adult life provides molecular evidence of the many maturational changes occurring during the early life of the adult fly.     

Regulation of the expression of the sn-glycerol-3-phosphate dehydrogenase gene in Drosophila melanogaster

P element-mediated transformation has been used to investigate the regulation of expression of the sn-glycerol-3-phosphate dehydrogenase gene of Drosophila melanogaster. A 13-kb construct containing the eight exons and associated introns, 5 kb of the 5' region, and 3 kb downstream from the structural gene produced normal levels of enzyme activity and rescued the poor viability of flies lacking the enzyme. All the regulatory elements essential for normal enzyme expression were located in a fragment that included the exons and introns and 1-kb upstream noncoding sequence. Deletions of the 1.6-kb second intron reduced activity to 25%. Transformants with fusion constructs between the sn-glycerol-3-phosphate dehydrogenase gene and the beta-galactosidase gene from E. coli revealed three elements that affected expression. A (CT)9 repeat element at the 5' end of the second intron increased expression in both larvae and adults, particularly at emergence. A second regulatory element, which includes a (CT)7 repeat, was located 5' to the TATA box and had similar effects on the gene's expression. A third, undefined, enhancer was located in the second intron, between 0.5 and 1.8 kb downstream of the translation initiation codon. This element increases enzyme activity to a similar extent in larvae and adults but has little effect when the enhancer at the 5' end of the intron is present.     

Amyrel, a paralogous gene of the amylase gene family in Drosophila melanogaster and the Sophophora subgenus

We describe a gene from Drosophila melanogaster related to the alpha-amylase gene Amy. This gene, which exists as a single copy, was named Amyrel. It is strikingly divergent from Amy because the amino acid divergence is 40%. The coding sequence is interrupted by a short intron at position 655, which is unusual in amylase genes. Amyrel has also been cloned in Drosophila ananassae, Drosophila pseudoobscura, and Drosophila subobscura and is likely to be present throughout the Sophophora subgenus, but, to our knowledge, it has not been detected outside. Unexpectedly, there is a strong conservation of 5' and 3' flanking regions between Amyrel genes from different species, which is not the case for Amy and which suggests that selection acts on these regions. In contrast to the Amy genes, Amyrel is transcribed in larvae of D. melanogaster but not in adults. However, the protein has not been detected yet. Amyrel evolves about twice as fast as Amy in the several species studied. We suggest that this gene could result from a duplication of Amy followed by accelerated and selected divergence toward a new adaptation.     

Size variation within the second hypervariable region of the surface envelope gene of the bovine lentivirus BIV in experimentally and naturally infected cattle

The bovine lentivirus also known as the bovine immunodeficiency-like virus (BIV) has conserved and hypervariable regions in the surface envelope (SU) gene. Size variation between isolates can be as large as 200 bp, mostly occurring in the second hypervariable (V2) gene region of the SU gene. The V2 region was cloned and sequenced from both experimentally and naturally infected cattle. Temporal evaluation of provirus from an experimentally inoculated cow showed two different-sized variants that appeared over time. The variation appeared to result from a recombinational event resulting in an apparent direct repeat. Cloned proviral nucleotide sequence diversity increased over time. Virus that was cultured and then cloned and sequenced showed progressive change from the inoculum virus, but culturing reduced the diversity of the clones as compared with direct amplification of provirus from leukocyte samples from the cow. The quasispecies phenomenon was evident in clones sequenced from a cow naturally infected with BIV. Of 10 clones examined from the V2 region, 6 different-size clones were present with nine different patterns of sequence rearrangement. Sequence length of different clones varied by as much as 43 amino acids (aa), with 21- and 15-aa direct repeats accounting for most of the size variation. Similar to other lentiviruses, BIV appears to mutate rapidly, which may be important in viral persistence and pathogenesis.     

Behavioral response to naturally occurring stress in cyclothymia and dysthymia.

Outlines an approach to epidemiologic-type life-stress studies that involves the intensive study of smaller groups of well-defined disordered individuals across the course of the response. On the basis of their previous research, the authors suggest that cyclothymia may be characterized by poor regulation of biologic and behavioral variation and that this condition may be exaggerated by stress. To extend the evidence on poor regulation generated in laboratory studies of cortisol functioning, 31 cyclothyme patients and 24 normal controls (median age 19.3 yrs) were asked to provide daily ratings of moods and events over 28 consecutive days. Data show that Ss with cyclothymia showed prolonged duration of recovery of behavioral levels compared to controls. Findings support a notion of weak inhibitory modulation in certain CNS systems that control biologic and behavioral variables related to affective disorder. It is suggested that the prolonged duration of behavioral change in cyclothymia is unlikely to be the consequence of concurrent depression and that the nature of prolonged recovery to stress in cyclothymia reflects the operation of an endogenous regulatory process, as opposed to a cognitive appraisal bias. (33 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Maternal Nanos regulates zygotic gene expression in germline progenitors of Drosophila melanogaster

Maternal Nanos (Nos) protein is required for germline development in Drosophila embryos. Here we show that Nos regulates zygotic gene expression in the germline progenitors, or pole cells. In order to probe the gene expression in pole cells, we screened ten enhancer-trap lines which showed beta-gal expression in pole cells. All of these enhancer-trap markers were fully activated in pole cells after their migration to the embryonic gonads. In the pole cells lacking Nos, the expression of nine out of ten enhancer-trap markers was affected. Among nine markers, five (Type-A) were prematurely expressed in the pole cells during the course of their migration. The expression of other four markers (Type-B) initiated correctly after pole-cell migration, but their expression was significantly reduced. Thus, we conclude that the maternal Nos plays a dual role in zygotic gene regulation in pole cells: to define the stages of expression for Type-A markers, and to enhance expression for Type-B markers. Contrary to our results, "Heller and Steinmann-Zwicky (1998)" have recently reported that no premature expression of Type-A markers occurs in the pole cells of embryos derived from nos mutant females. This discrepancy is due to the difference in the nos mutant alleles used for these analyses. We used the much stronger allele, nosBN.     

Divergent and conserved features in the spatial expression of the Drosophila pseudoobscura esterase-5B gene and the esterase-6 gene of Drosophila melanogaster

The regulatory regions of homologous genes encoding esterase 6 (Est-6) of Drosophila melanogaster and esterase 5B (Est-5B) of Drosophila pseudoobscura show very little similarity. We have undertaken a comparative study of the pattern of expression directed by the Est-5B and Est-6 5'-flanking DNA to attempt to reveal conserved elements regulating tissue-specific expression in adults. Esterase regulatory sequences were linked to a lacZ reporter gene and transformed into D. melanogaster embryos. Est-5B, 5' upstream elements, give rise to a beta-galactosidase expression pattern that coincides with the wild-type expression of Est-5B in D. pseudoobscura. The expression patterns of the Est-5B/lacZ construct are different from those of a fusion gene containing the upstream region of Est-6. Common sites of expression for both kinds of constructs are the third segment of antenna, the maxillary palps, and salivary glands. In vitro deletion mutagenesis has shown that the two genes have a different organization of regulatory elements controlling expression in both the third segment of antenna and maxillary palps. The results suggest that the conservation of the expression pattern in genes that evolved from a common ancestor may not be accompanied by preservation of the corresponding cis-regulatory elements.     

pRJ5: a naturally occurring Staphylococcus aureus plasmid expressing constitutive macrolide-lincosamide-streptogramin B resistance contains a tandem duplication in the leader region of the ermC gene

The 2.55 kb Staphylococcus aureus plasmid, pRJ5, confers constitutive resistance to macrolide-lincosamide-streptogramin B (MLS) antibiotics. pRJ5 is nearly identical to the inducible MLS resistance plasmid pT48, and has homology with the S. aureus plasmids pE194 and pSN2. The HindIII-C and/or Hind-B fragments were required for stable maintenance of the plasmid and probably carry palA. Plasmids pRJ5 and pT48 were shown to belong to the same incompatibility group, Inc12 (L). DNA sequencing showed that pRJ5 contains a 28 bp direct tandem duplication in the leader/attenuator region of ermC. This is likely to change the secondary structure of the methylase mRNA, allowing constitutive expression of ermC. The type of mutation found on plasmid pRJ5 is different from those observed in similar 2.5 kb constitutive MLS-resistance plasmids isolated from other Gram-positive bacteria, including staphylococci.