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Telomere length in Drosophila is maintained by targeted transposition of three non-LTR retrotransposons: HeT-A, TART and TAHRE (HTT), but understanding the regulation of this process is hindered by our poor knowledge of HTT associated proteins. We have identified new protein components of the HTT array: Chromator (Chro), the TRF2/DREF complex and the sumoylation machinery. Chro was localized on telomeric HTT arrays by immunostaining, where it may interact with Prod directly, as indicated by yeast two-hybrid interaction, co-IP, and colocalization on polytene chromosomes. The TRF2/DREF complex may promote the open structure of HTT chromatin. The protein interactions controlling HTT chromatin structure and telomere length may be modulated by sumoylation.

Haematophagous insects are frequently carriers of parasitic diseases, including malaria. The mosquito Anopheles gambiae is the major vector of malaria in sub-Saharan Africa and is thus responsible for thousands of deaths daily. Although the role of olfaction in A. gambiae host detection has been demonstrated, little is known about the combinations of ligands and odorant binding proteins (OBPs) that can produce specific odor-related responses in vivo. We identified a ligand, indole, for an A. gambiae odorant binding protein, AgamOBP1, modeled the interaction in silico and confirmed the interaction using biochemical assays. RNAi-mediated gene silencing coupled with electrophysiological analyses confirmed that AgamOBP1 binds indole in A. gambiae and that the antennal receptor cells do not respond to indole in the absence of AgamOBP1. This case represents the first documented instance of a specific A. gambiae OBP-ligand pairing combination, demonstrates the significance of OBPs in odor recognition, and can be expanded to the identification of other ligands for OBPs of Anopheles and other medically important insects.

At the heart of the odor recognition process in all animals are G-protein-coupled receptors, which are seven-transmembrane domain proteins that initiate G-protein-mediated signaling cascades when activated by their ligands. Odorant receptors (ORs) are a large, diverse family of proteins with some 80 members in the mosquito Anopheles gambiae. With the assumption that more sensilla on female antennae are tuned to human odors than on male antennae, comparison of specific OR mRNA levels in male and female antennae can provide an indication as to which receptors may be stimulated by host odors. We have used RT PCR and quantitative real-time PCR (qRT PCR) to investigate sex-biased expression levels of 80 A. gambiae ORs in male and female antennae and maxillary palps. On the basis of prevalence of expression in female antennae and on a strong female relative to male expression bias we identified a short list of ORs that are likely involved in host odor recognition by female mosquitoes.

Drosophila telomeres comprise DNA sequences that differ dramatically from those of other eukaryotes. Telomere functions, however, are similar to those found in telomerase-based telomeres, even though the underlying mechanisms may differ. Drosophila telomeres use arrays of retrotransposons to maintain chromosome length, while nearly all other eukaryotes rely on telomerase-generated short repeats. Regardless of the DNA sequence, several end-binding proteins are evolutionarily conserved. Away from the end, the Drosophila telomeric and subtelomeric DNA sequences are complexed with unique combinations of proteins that also modulate chromatin structure elsewhere in the genome. Maintaining and regulating the transcriptional activity of the telomeric retrotransposons in Drosophila requires specific chromatin structures and, while telomeric silencing spreads from the terminal repeats in yeast, the source of telomeric silencing in Drosophila is the subterminal arrays. However, the subterminal arrays in both species may be involved in telomere-telomere associations and/or communication., ((c) 2007 Wiley Periodicals, Inc.)

Drosophila melanogaster telomeres have two DNA domains: a terminal array of retrotransposons and a subterminal repetitive telomere-associated sequence (TAS), a source of telomere position effect (TPE). We reported previously that deletion of the 2L TAS array leads to dominant suppression of TPE by stimulating in trans expression of a telomeric transgene. Here, we compared the transcript activities of a w transgene inserted between the retrotransposon and TAS arrays at the 2L telomere in genotypes with different lengths of the 2L TAS. In contrast to individuals bearing a wild-type 2L homologue, flies with a TAS deficiency showed a significant increase in the level of telomeric w transcript during development, especially in pupae. Moreover, we identified a read-through w transcript initiated from a retrotransposon promoter in the terminal array. Read-through transcript levels also significantly increased with the presence of a 2L TAS deficiency in trans, indicating a stimulating force of the TAS deficiency on retrotransposon promoter activity. The read-through transcript contributes to total w transcript, although most w transcript originates at the w promoter. While silencing of transgenes in nonhomologous telomeres is suppressed by 2L TAS deficiencies, suggesting a global effect, the overall level of HeT-A transcripts is not increased under similar conditions.

We report in this paper that the PROD protein, encoded by the gene proliferation disrupter (prod), is associated with the telomeric chromatin in Drosophila melanogaster. It binds to a region just upstream of the promoter of the telomere-specific retrotransposon HeT-A, which is located in the long 3'untranslated region of the element near its oligo(A) tail. Reduction of PROD in prod heterozygote flies results in elevated levels of HeT-A RNA in the ovaries, suggesting that PROD functions as a repressor of HeT-A transcriptional activity at the telomeres.

Chromosome length in Drosophila is maintained by targeted transposition of three non-long terminal repeat retrotransposons, HeT-A, TART, and TAHRE, to the chromosome ends. The length and composition of these retrotransposon arrays can vary significantly between chromosome tips and between fly stocks, but the significance and consequences of these length differences are not understood. A dominant genetic factor, Tel, has been described, which causes a severalfold elongation of the retrotransposon arrays at all telomeres. We used this strain to assess possible affects of extended telomeres on the organism. While we found no effect on life span of the adults, we could demonstrate a correlation between long telomeres and reduced fertility and fecundity in individual females, which is also reflected in abnormal oocyte development.

Telomeres are generally considered heterochromatic. On the basis of DNA composition, the telomeric region of Drosophila melanogaster contains two distinct subdomains: a subtelomeric region of repetitive DNA, termed TAS, and a terminal array of retrotransposons, which perform the elongation function instead of telomerase. We have identified several P-element insertions into this retrotransposon array and compared expression levels of transgenes with similar integrations into TAS and euchromatic regions. In contrast to insertions in TAS, which are silenced, reporter genes in the terminal HeT-A, TAHRE, or TART retroelements did not exhibit repressed expression in comparison with the same transgene construct in euchromatin. These data, in combination with cytological studies, provide evidence that the subtelomeric TAS region exhibits features resembling heterochromatin, while the terminal retrotransposon array exhibits euchromatic characteristics.

Noncoding repetitive sequences make up a large portion of eukaryotic genomes, but their function is not well understood. Large blocks of repetitive DNA-forming heterochromatin around the centromeres are required for this region to function properly, but are difficult to analyze. The smaller regions of heterochromatin at the telomeres provide an opportunity to study their DNA and protein composition. Drosophila telomere length is maintained through the targeted transposition of specific non-long terminal repeat retrotransposons to chromosome ends, where they form long tandem arrays. A subterminal telomere-associated sequence (TAS) lies immediately proximal to the terminal-retrotransposon array. Here, we review the experimental support for the heterochromatic features of Drosophila telomeres, and provide evidence that telomeric regions contain 2 distinct chromatin subdomains: TAS, which exhibits features that resemble beta heterochromatin; and the terminal array of retrotransposons, which appears euchromatic. This organization is significantly different from the telomeric organization of other eukaryotes, where the terminal telomerase-generated repeats are often folded in a t-loop structure and become part of the heterochromatin protein complex.

Chromosome ends in Drosophila melanogaster can be elongated either by terminal attachment of the telomere-specific retrotransposons HeT-A and TART or by terminal gene conversion. Here we show that a decrease in Ku70 or Ku80 gene dosage causes a sharp increase in the frequency of HeT-A and TART attachments to a broken chromosome end and in terminal DNA elongation by gene conversion. Loss of Ku80 has more pronounced effects than loss of Ku70. However, lower Ku70 concentration reduces the stability of terminally deficient chromosomes. Our results suggest a role of the end-binding Ku complex in the accessibility and length regulation of Drosophila telomeres.

Drosophila melanogaster extends its telomeres by transposition of two non-LTR retrotransposons, HeT-A and TART, to chromosome ends. We have determined the tissue-specific expression of these two elements by whole-mount in situ hybridization with digoxigenin-labeled RNA sense and antisense probes in the germ line and in a variety of larval tissues during normal development in the wild type and in tissues of mutants that cause overproliferation. Our results indicate that transcript levels, which are a key component in the process of telomere elongation in D. melanogaster, are correlated with cell proliferation in normal tissues and that RNA levels are elevated in growth-stimulated tissues.

In spite of advances in medicine and public health, malaria and other mosquito-borne diseases are on the rise worldwide. Although vaccines, genetically modified mosquitoes and safer insecticides are under development, herein we examine a promising new approach to malaria control through better repellents. Current repellents, usually based on DEET, inhibit host finding by impeding insect olfaction, but have significant drawbacks. We discuss how comparative genomics, using data from the Anopheles genome project, allows the rapid identification of members of three protein classes critical to insect olfaction: odorant-binding proteins, G-protein-coupled receptors, and odorant-degrading enzymes. A rational design approach similar to that used by the pharmaceutical industry for drug development can then be applied to the development of products that interfere with mosquito olfaction. Such products have the potential to provide more complete, safer and longer lasting protection than conventional repellents, preventing disease transmission by interrupting the parasite life cycle., (Copyright 2003 Wiley Periodicals, Inc.)

The terminal DNA arrays on chromosomes of Drosophila melanogaster are composed of two families of non-LTR retrotransposons, HeT-A and TART. Available evidence suggests that chromosome length in this species and its close relatives is maintained by targeted transposition of these elements, with attachment of the elements to the chromosome end by their 3' oligo(A) tails. However, the regulation of transposition of these elements and the control of telomere length are poorly understood. Here we present the hypothesis that the forces involved in telomere length regulation in Drosophila are the underlying forces that manifest themselves as telomeric position effect (TPE). Based on recent studies of TPE, which found that expression of a reporter gene is influenced by telomere structure in cis and trans, we propose that the subtelomeric satellite (TAS) in D. melanogaster plays an important role in controlling telomere elongation. Transcription of a HeT-A element is probably initiated at a promoter in the 3' UTR of an upstream element, and TAS may repress this transcriptional activity in cis and trans. A region of HeT-A not at the extreme 3' end of the element may act as a transcriptional enhancer that may be modulated by TAS.

One model of telomeric position effect (TPE) in Drosophila melanogaster proposes that reporter genes in the vicinity of telomeres are repressed by subterminal telomere-associated sequences (TAS) and that variegation of these genes is the result of competition between the repressive effects of TAS and the stimulating effects of promoters in the terminal HeT-A transposon array. The data presented here support this model, but also suggest that TPE is more complex. Activity of a telomeric white reporter gene increases in response to deletion of some or all of the TAS on the homolog. Only transgenes next to fairly long HeT-A arrays respond to this trans-interaction. HeT-A arrays of 6-18 kb respond by increasing the number of dark spots on the eye, while longer arrays increase the background eye color or increase the number of spots sufficiently to cause them to merge. Thus, expression of a subtelomeric reporter gene is influenced by the telomere structure in cis and trans. We propose that the forces involved in telomere length regulation in Drosophila are the underlying forces that manifest themselves as TPE. In the wild-type telomere TAS may play an important role in controlling telomere elongation by repressing HeT-A promoter activity. Modulation of this repression by the homolog may thus regulate telomere elongation.

One way of controlling disease transmission by blood-feeding mosquitoes is to reduce the frequency of insect-host interaction, thus reducing the probability of parasite transmission and re-infection. A better understanding of the olfactory processes responsible for allowing mosquitoes to identify human hosts is required in order to develop methods that will interfere with host seeking. We have therefore initiated a molecular approach to isolate and characterize the genes and their products that are involved in the olfactory recognition pathway of the mosquito Anopheles gambiae, which is the main malaria vector in sub-Saharan Africa. We report here the isolation and preliminary characterization of several cDNAs from male and female A. gambiae antennal libraries that encode putative odourant binding proteins. Their conceptual translation products show extensive sequence similarity to known insect odourant binding proteins (OBPs)/pheromone binding proteins (PBPs), especially to those of D. melanogaster. The A. gambiae OBPs described here are expressed in the antennae of both genders, and some of the A. gambiae OBP genes are well conserved in other disease-transmitting mosquito species, such as Aedes aegypti and Culex quinquefasciatus.