Transvection and chromosomal trans-interaction effects.
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Cloning and characterization of a copy of Tirant transposable element in Drosophila melanogaster.
A Tirant element, inserted at the 5' end of the mitochondrial glutamine synthetase (mt-gs) gene in a mutant allele giving rise to a recessive female sterility phenotype, was cloned and utilized to characterize this novel retrotransposable element of the Drosophila melanogaster genome. The 5.3 kb element present in the fs(2) PM11-19 mt-gs allele possesses a 417 bp long terminal repeat (LTR) at both ends. There is a serine tRNA binding site downstream of the 5' LTR sequence and a polypurine tract upstream of the 3' LTR end. The insertion leads to the duplication of a host-site CGCG sequence. In situ hybridization to salivary glands chromosomes showed evidence of the mobile nature of the element. The DNA sequencing of the cloned 5.3 kb element revealed that Tirant possesses an open reading frame (ORF) that shows similarity with the envelope protein encoded by the gypsy and 297 retrotransposons. In addition, the cloned element appears to be a subgenomic fragment of a not yet identified complete element, because only the integrase domain of the reverse transcriptase gene is found.
Amino Acid Sequence↗
Chromosome movement: dynein-out at the kinetochore.
Cell biologists have long speculated that a minus end-directed motor localized at kinetochores contributes to the poleward movement of chromosomes during mitosis. Two recent studies provide direct evidence that cytoplasmic dynein can perform this function.
Animals↗
Polymorphism and linkage analysis of the prothoracicotropic hormone gene in the silkmoth, Bombyx mori.
We looked for polymorphism of the prothoracicotropic hormone gene locus (Ptth) among inbred strains of the silkmoth, Bombyx mori, by in vitro DNA amplification (polymerase chain reaction), and found three alleles, PtthA, PtthB and PtthC. The PtthA allele contained a third intron consisting of 680 bp and a fourth intron of 350 bp. PtthB contained the same size third intron but a longer fourth intron of 490 bp, while PtthC had a longer third intron of 1080 bp and a shorter fourth intron of 350 bp. In 29 strains which we examined, 9 strains had PtthA, 8 strains had PtthB, and 2 strains had PtthC. The other 10 strains had heterogeneous genotypes with the same 3 alleles. Linkage analyses using these polymorphisms showed that Ptth was linked to the or gene on chromosome 22, and was independent of other chromosome markers. To determine the map position, we obtained an F1 hybrid between the w41 strain (PtthC+or+mw) and the w22 strain (PtthB or mw), and backcrossed the F1 hybrid to females of the w22 strain. From the segregation of or, mw and Ptth in 861 progeny, recombination values were calculated as 18.5% between or and mw, 23.9% between mw and Ptth, and 7.3% between Ptth and or. Because or and mw have already been mapped at positions 22-8.9 cM and 22-25.2 cM, respectively, we mapped Ptth at 22-2.5 cM. The Ptth locus is independent of any loci which are known to control growth and molting.
Animals↗
Microsatellites reveal extensive geographical, ecological and genetic contacts between invasive and indigenous whitefly biotypes in an insular environment.
Human-mediated bioinvasions provide the opportunity to study the early stages of contact between formerly allopatric, divergent populations of a species. However, when invasive and resident populations are morphologically similar, it may be very difficult to assess their distribution in the field, as well as the extent of ecological overlap and genetic exchanges between invasive and resident populations. We here illustrate the use of data obtained from a set of eight microsatellite markers together with Bayesian clustering methods to document invasions in a group of major tropical pests, Bemisia tabaci, which comprises several morphologically indistinguishable biotypes with different agronomic impacts. We focus on the island of La Réunion, where an invasive biotype (B) has recently been introduced and now interacts with the resident biotype (Ms). The temporal and spatial distribution, host-plant range and genetic structure of both biotypes are investigated. We showed (i) that, without prior information, clustering methods separate two groups of individuals that can safely be identified as the B and Ms biotypes; (ii) that the B biotype has invaded all regions of the island, and showed no signs of genetic founder effect relative to the Ms biotype; (iii) that the B and Ms biotypes coexist in sympatry throughout most of their geographical ranges, although they tend to segregate into different host plants; and finally (iv) that asymmetrical and locus-specific introgression occurs between the two biotypes when they are in syntopy.
Animals↗
Mapping candidate genes for Drosophila melanogaster resistance to the parasitoid wasp Leptopilina boulardi.
Drosophila melanogaster resistance against the parasitoid wasp Leptopilina boulardi is under the control of a single gene (Rlb), with two alleles, the resistant one being dominant. Using strains bearing deletions, we previously demonstrated that the 55E2-E6; 55F3 region on chromosome 2R is involved in the resistance phenomenon. In this paper, we first restricted the Rlb containing region by mapping at the molecular level the breakpoints of the Df(2R)Pc66, Df(2R)P34 and Df(2R)Pc4 deficiencies, using both chromosomal in situ hybridization and Southern analyses. The resistance gene was localized in a 100 kb fragment, predicted to contain about 10 different genes. Male recombination genetic experiments were then performed, leading to identification of two possible candidates for the Rlb gene. Potential involvement of one of this genes, edl/mae, is discussed.
Animals↗
Genetic factors on the second and third chromosomes responsible for the variation of amylase activity and inducibility in Drosophila melanogaster.
Using second- or third-chromosome substitution lines of Drosophila melanogaster, the genetic variation of inducibility and amylase specific activities in three media (starch, normal and glucose) were investigated. Genetic factors on both the second and third chromosomes were responsible for the variation in amylase specific activity and inducibility. In glucose medium, the genetic variance of amylase specific activity estimated for the second-chromosome substitution lines was larger than that for the third-chromosome substitution lines; however, for starch medium and inducibility, the variance was larger for the third-chromosome substitution lines. High correlations for the second-chromosome substitution lines and low correlations for the third-chromosome substitution lines were observed for amylase specific activities in different media. These results suggest that the genetic factor(s) responsible for inducibility or amylase activity variation in an induced medium such as starch should be on the third chromosome and those in the non-induced medium such as glucose should be on the second chromosome. The functional roles of the factors on the second and third chromosomes would be the repression and induction of amylase, respectively.
Animals↗
Linkage disequilibrium mapping of molecular polymorphisms at the scabrous locus associated with naturally occurring variation in bristle number in Drosophila melanogaster.
We evaluated the hypothesis that the Drosophila melanogaster second chromosome gene scabrous (sca), a candidate sensory bristle number quantitative trait locus (QTL), contributes to naturally occurring variation in bristle number. Variation in abdominal and sternopleural bristle number was quantified for wild-derived sca alleles in seven genetic backgrounds: as homozygous second chromosomes (C2) in an isogenic background, homozygous lines in which approximately 20 cM including the sca locus had been introgressed into the isogenic background (sca BC), as C2 and sca BC heterozygotes and hemizygotes against a P element insertional sca allele and a P-induced sca deficiency in the same isogenic background, and as sca BC heterozygotes against the wild-type sca allele of isogenic strain. Molecular restriction map variation was determined for a 45 kb region including the sca locus, and single-stranded conformational polymorphism (SSCP) was examined for the third intron and parts of the third and fourth exons. Associations between each of the 27 molecular polymorphisms and bristle number were evaluated within each genotype and on the first principal component score determined from all seven genotypes, separately for each sex and bristle trait. Permutation tests were used to assess the empirical significance thresholds, accounting for multiple, correlated tests, and correlated markers. Three sites in regulatory regions were associated with female-specific variation in abdominal bristle number, one of which was an SSCP site in the region of the gene associated with regulation of sca in embryonic abdominal segments.
Animals↗
A new widely distributed nuclear protein involved in the regulation of transcription catalyzed by RNA polymerase II.
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Amino Acid Sequence↗
The Drosophila claret segregation protein is a minus-end directed motor molecule.
A product encoded at the claret locus in Drosophila is needed for normal chromosome segregation in meiosis in females and in early mitotic divisions of the embryo. The predicted amino-acid sequence of the segregation protein was shown recently to be strikingly similar to Drosophila kinesin heavy chain. We have expressed the claret segregation protein in bacteria and have found that the bacterially expressed protein has motor activity in vitro with several novel features. The claret motor is slow (4 microns min-1), unlike either kinesin or dyneins. It has the directionality, the ability to generate torque and the sensitivity to inhibitors reported previously for dyneins. The finding of minus-end directed motor activity for a protein with sequence similarity to kinesin suggests that the dynein and kinesin motor domains are ancestrally related. The minus-end directed motor activity of the claret motor is consistent with a role for this protein in producing chromosome movement along spindle microtubules during prometaphase and/or anaphase.
Adenosine Triphosphatases↗
The Grapes checkpoint coordinates nuclear envelope breakdown and chromosome condensation.
Mutations in the embryonic Drosophila Grapes/Chk1 checkpoint result in an abbreviated interphase, chromosome condensation defects and metaphase delays. To clarify the relationship between these phenotypes, we simultaneously timed multiple nuclear and cytoplasmic events in mutant grp-derived embryos. These studies support a model in which grp disrupts an S-phase checkpoint, which results in progression into metaphase with incompletely replicated chromosomes. We also show that chromosome condensation is independent of the state of DNA replication in the early embryo. Therefore, grp condensation defects are not a direct consequence of entering metaphase with incompletely replicated chromosomes. Rather, initiation of chromosome condensation (ICC) occurs at the normal time in grp-derived embryos, but the shortened interval between ICC and metaphase does not provide sufficient time to complete condensation. Our results suggest that these condensation defects, rather than incomplete DNA replication, are responsible for the extensive metaphase delays observed in grp-derived embryos. This analysis provides an example of how the loss of a checkpoint can disrupt the timing of multiple events not directly monitored by that checkpoint. These results are likely to apply to vertebrate cells and suggest new strategies for destroying checkpoint-compromised cancer cells.
Animals↗
Dynamics of heat shock factor association with native gene loci in living cells.
Direct observation of transcription factor action in the living cell nucleus can provide important insights into gene regulatory mechanisms. Live-cell imaging techniques have enabled the visualization of a variety of intranuclear activities, from chromosome dynamics to gene expression. However, progress in studying transcription regulation of specific native genes has been limited, primarily as a result of difficulties in resolving individual gene loci and in detecting the small number of protein molecules functioning within active transcription units. Here we report that multiphoton microscopy imaging of polytene nuclei in living Drosophila salivary glands allows real-time analysis of transcription factor recruitment and exchange on specific native genes. After heat shock, we have visualized the recruitment of RNA polymerase II (Pol II) to native hsp70 gene loci 87A and 87C in real time. We show that heat shock factor (HSF), the transcription activator of hsp70, is localized to the nucleus before heat shock and translocates from nucleoplasm to chromosomal loci after heat shock. Assays based on fluorescence recovery after photobleaching show a rapid exchange of HSF at chromosomal loci under non-heat-shock conditions but a very slow exchange after heat shock. However, this is not a consequence of a change of HSF diffusibility, as shown here directly by fluorescence correlation spectroscopy. Our results provide strong evidence that activated HSF is stably bound to DNA in vivo and that turnover or disassembly of transcription activator is not required for rounds of hsp70 transcription. This and previous studies indicate that transcription activators display diverse dynamic behaviours in their associations with targeted loci in living cells. Our method can be applied to study the dynamics of many factors involved in transcription and RNA processing, and in their regulation at native heat shock genes in vivo.
Animals↗
The involvement of ovarian tumour in the intracellular localization of Sex-lethal protein.
The Drosophila ovarian tumour gene is required at multiple times in the germline for oogenesis. A second gene, Sex-lethal, controls sex determination in the soma and also has a separate germline function affecting similar oogenic stages as ovarian tumour. We demonstrate that ovarian tumour is not required for early Sex-lethal gene expression in the female germline, as had been previously reported. Instead, we provide evidence that ovarian tumour has a specific role in the developmentally regulated accumulation of SEX-LETHAL protein within the cytoplasm and nucleus. Furthermore, the examination of nurse cell polytene chromosomes produced by certain ovarian tumour mutations showed that SEX-LETHAL protein can associate with discrete chromosomal sites in the germline and that this pattern appears to change as the egg chamber matures. This is the first indication that SEX-LETHAL is capable of direct physical interactions with chromosomes (albeit in a mutant background) and is consistent with the developmentally regulated nuclear localization of SEX-LETHAL being important for oogenesis.
Animals↗
A preliminary linkage map of the hard tick, Ixodes scapularis.
A linkage map of the Ixodes scapularis genome was constructed, based upon segregation amongst 127 loci. These included 84 random amplified polymorphic DNA (RAPD) markers, 32 Sequence-Tagged RAPD (STAR) markers, 5 cDNAs, and 5 microsatellites in 232 F1 intercross progeny from a single, field-collected P1 female. A preliminary linkage map of 616 cM was generated across 14 linkage groups with one marker every 10.8 cM. Assuming a genome size of approximately 109 bp, the relationship of physical to genetic distance was found to be approximately 300 kb/cM in the I. scapularis genome.
Animals↗
Resistance to dieldrin + fipronil assorts with chromosome inversion 2La in the malaria vector Anopheles gambiae.
Cyclodiene insecticide resistance in malaria vector mosquitoes of the Anopheles gambiae species complex (Diptera: Culicidae) has been reported previously from several parts of Africa. We report resistance to dieldrin, a cyclodiene, in two laboratory strains of An. gambiae Giles sensu stricto code-named Ian P20 from Nigeria (1979) and CIG from Cote d'Ivoire (1997). Dieldrin resistance levels were high in adult female mosquitoes (40-75% survived exposure to 4% dieldrin for 1 h) and was closely linked with chromosomal paracentric inversion 2La. This inversion did not occur in Hardy-Weinberg proportions, but showed an excess of heterozygotes in both strains, which may account for the high levels of resistance. This linkage also suggests that dieldrin resistance in Ian P20 is dominant. After subsamples of strain Ian P20 were exposed for 1 h to dieldrin 4% or fipronil 2% (discriminating concentrations), the resultant mortality-rates (61% and 65%) were not significantly different. Most survivors after fipronil treatment also survived subsequent exposure to dieldrin (46/50=92%). This apparent cross-resistance between insecticides of two classes (cyclodiene and phenyl pyrazole) has implications for the management of insecticide resistance in wild populations of the An. gambiae complex.
Animals↗
Gene conversion is involved in the transfer of genetic information between naturally occurring inversions of Drosophila.
The DNA sequences of the ribosomal protein 49 (rp49) region were determined for 34 isochromosomal strains of Drosophila subobscura representing two chromosomal arrangements, the OST and the O3 + 4 gene arrangements, which differ by two overlapping inversions. The data reveal that gene conversion is a mechanism responsible for the transfer of genetic information between naturally occurring inversions of Drosophila. The estimated rate of gene transfer by gene conversion at this region, which is close to an inversion breakpoint, is lower than previous estimates obtained experimentally at the rosy (ry) gene in Drosophila melanogaster. Our data indicate that gene arrangements OST and O3 + 4 are monophyletic and rather old (0.58 and 0.73 million years old, respectively).
Animals↗
The onchocerciasis focus at Kinsuka/Kinshasa (Republic of Zaire) in 1985. I. Entomological aspect.
The entomological aspect of the onchocerciasis focus at Kinsuka/Kinshasa has not been studied since vector control was carried out in 1948. As in 1940, larvae of Simulium damnosum s.l. are located in the arm of the Zaire River flowing between Mimosa Island and the Zairian bank. They are, however, scarce. Cytotaxonomic studies showed that the S. squamosum group was the only member of the S. damnosum complex present in the region of Kinshasa. Physico-chemical analysis of the water at breeding sites determined that S. squamosum larvae develop in neutral or acid water with a conductivity of less than 50 micronhos cm-1 and a low concentration of dissolved salts. Longitudinal study showed that the 'annual biting rate' (ABR) and the 'annual transmission potential' (ATP) fluctuate from 2406-5999 bites man-1 year-1 and from 120-236 infective larvae man-1 year-1 respectively. The low transmission rate at Kinsuka is mainly due to the low level of the biting population occurring during the main river rising. The increase in Kinsuka's population has reinforced this situation by 'diluting' the parasite in the human reservoir.
Animals↗
D-Titin: a giant protein with dual roles in chromosomes and muscles.
Previously, we reported that chromosomes contain a giant filamentous protein, which we identified as titin, a component of muscle sarcomeres. Here, we report the sequence of the entire titin gene in Drosophila melanogaster, D-Titin, and show that it encodes a two-megadalton protein with significant colinear homology to the NH(2)-terminal half of vertebrate titin. Mutations in D-Titin cause chromosome undercondensation, chromosome breakage, loss of diploidy, and premature sister chromatid separation. Additionally, D-Titin mutants have defects in myoblast fusion and muscle organization. The phenotypes of the D-Titin mutants suggest parallel roles for titin in both muscle and chromosome structure and elasticity, and provide new insight into chromosome structure.
Animals↗