Biomedical subjects

H Biessmann

Publications and source records attributed to H Biessmann.

At least 19 recordsLinked to original sources

PubMed1999

Silencing at Drosophila telomeres: nuclear organization and chromatin structure play critical roles.

Transgenes inserted into the telomeric regions of Drosophila melanogaster chromosomes exhibit position effect variegation (PEV), a mosaic silencing characteristic of euchromatic genes brought into juxtaposition with heterochromatin. Telomeric transgenes on the second and third chromosomes are flanked by telomeric associated sequences (TAS), while fourth chromosome telomeric transgenes are most often associated with repetitious transposable elements. Telomeric PEV on the second and third chromosomes is suppressed by mutations in Su(z)2, but not by mutations in Su(var)2-5 (encoding HP1), while the converse is true for telomeric PEV on the fourth chromosome. This genetic distinction allowed for a spatial and molecular analysis of telomeric PEV. Reciprocal translocations between the fourth chromosome telomeric region containing a transgene and a second chromosome telomeric region result in a change in nuclear location of the transgene. While the variegating phenotype of the white transgene is suppressed, sensitivity to a mutation in HP1 is retained. Corresponding changes in the chromatin structure and inducible activity of an associated hsp26 transgene are observed. The data indicate that both nuclear organization and local chromatin structure play a role in this telomeric PEV.

Animals↗

PubMed1999

Moose, a new family of LTR-retrotransposons in the mosquito Anopheles gambiae.

A novel LTR retrotransposable element called moose has been cloned and characterized from the malaria transmitting mosquito, Anopheles gambiae. This element has all the characteristic features of LTR retroelements and is related to retroelements from other insects and nematodes, belonging to a subgroup of retroelements distinct from the copia/Ty1 and gypsy/Ty3 groups. The moose element appears to be active in A. gambiae, and strong RNA expression is detected in the male and female gonads. The use of this retroelement as a potential vector for germ line transformation is discussed.

Amino Acid Sequence↗

PubMed1999

Molecular cloning and tissue-specific expression of the mutator2 gene (mu2) in Drosophila melanogaster.

We present here the molecular cloning and characterization of the mutator2 (mu2) gene of Drosophila melanogaster together with further genetic analyses of its mutant phenotype. mu2 functions in oogenesis during meiotic recombination, during repair of radiation damage in mature oocytes, and in proliferating somatic cells, where mu2 mutations cause an increase in somatic recombination. Our data show that mu2 represents a novel component in the processing of double strand breaks (DSBs) in female meiosis. mu2 does not code for a DNA repair enzyme because mu2 mutants are not hypersensitive to DSB-inducing agents. We have mapped and cloned the mu2 gene and rescued the mu2 phenotype by germ-line transformation with genomic DNA fragments containing the mu2 gene. Sequencing its cDNA demonstrates that mu2 encodes a novel 139-kD protein, which is highly basic in the carboxy half and carries three nuclear localization signals and a helix-loop-helix domain. Consistent with the sex-specific mutant phenotype, the gene is expressed in ovaries but not in testes. During oogenesis its RNA is rapidly transported from the nurse cells into the oocyte where it accumulates specifically at the anterior margin. Expression is also prominent in diploid proliferating cells of larval somatic tissues. Our genetic and molecular data are consistent with the model that mu2 encodes a structural component of the oocyte nucleus. The MU2 protein may be involved in controlling chromatin structure and thus may influence the processing of DNA DSBs.

Amino Acid Sequence↗

PubMed1998

Directional gene silencing induced by a complex subtelomeric satellite from Drosophila.

The telomeric regions in Drosophila cause transcriptional silencing of integrated transgenes. A complex satellite has recently been identified in the subterminal region of the left arm of chromosome 2 that is a good candidate for the source of the observed telomeric silencing, because genetically marked transposable elements that have inserted into this subtelomeric array show repression and variegation of the reporter gene. We asked whether this satellite can also cause transcriptional repression in ectopic chromosomal positions by placing it upstream of a mini-white reporter gene in P element constructs used for germ line transformation. The transgenes are shielded from external influences at the integration site using SU(HW) binding sites at either end. It was found that the satellite represses transcription of the reporter gene in an orientation dependent and an array length dependent manner. The satellite does not, however, induce variegation under the conditions used. The repressed transgenes do not respond to typical modifiers of centromeric position effect variegation, such as Su(var)2055, Su(var)2-11, Su(var)3-11, and Su(var)3-61, or to the addition of a Y chromosome. However, as with the original variegating telomeric insertion, suppression in the transgenes is relieved by Su(z)25, suggesting that suppression induced by the subtelomeric satellite retains aspects of telomeric silencing in ectopic positions.

ATP-Binding Cassette Transporters↗

PubMed1998

DNA organization and length polymorphism at the 2L telomeric region of Anopheles gambiae.

The molecular structure of the telomeric region at the left arm of the second chromosome of the mosquito Anopheles gambiae has been determined in the transformed strain G418 that contains a pUChsneo transgene attached at the 2L chromosome end, and in the Pink eye laboratory strain (PE). Both strains contain the same complex satellite positioned distal to a unique region. FIGE mapping of the telomeric region of the PE strain revealed distinct DNA fragment lengths that segregated with individual chromosomes. Genomic DNA fragments were cloned from the 2L telomeric region, which accounted for about half of 2L chromosomes in the PE population. In all three cases studied, long fragments of different middle repetitive sequences were found attached to the distal ends of the 2L satellite. We propose that random fragments of DNA may be occasionally added during recombination between complex satellite repeats at the chromosome ends.

Animals↗

PubMed1997

Drosophila telomere elongation.

Drosophila melanogaster has an unusual telomere elongation mechanism. Instead of short repeats that are synthesized by telomerase, long retrotransposons, HeT-A and TART, transpose to the ends of chromosomes. This mechanism generates tandem arrays of these elements at the chromosome ends, in which all elements are oriented with their oligo(A) tails towards the centromere. Structural features of HeT-A and TART elements may provide clues as to their transposition mechanism. Drosophila telomere length polymorphism is mainly due to terminal retrotransposon arrays that differ between chromosome tips and that change with time. In addition, stable terminal chromosome deletions can be generated that do not contain terminal HeT-A and TART arrays, suggesting that, unlike the equivalent terminal repeats in yeast and humans, the presence and length of terminal arrays in Drosophila may not be critical for cell cycle progression.

Animals↗

PubMed1997

Telomere maintenance without telomerase.

Telomeres are nucleoprotein structures at the ends of eukaryotic chromosomes that perform a number of vital functions. They allow a cell to distinguish between natural chromosome ends and chromosome breaks in order to delay the cell cycle and repair the broken end. Telomeres also compensate for the inability of DNA polymerase to replicate the chromosome completely. In most eukaryotes a special reverse transcriptase, telomerase, adds telomeric DNA repeats to the chromosome ends using an internal RNA template. However, evidence is accumulating for alternative elongation mechanisms in a variety of eukaryotes. In the yeast Saccharomyces cerevisiae, and possibly in humans, both of which normally use telomerase, a different mechanism can be used for chromosome length maintenance when telomerase is inactive or inactivated. Yeast apparently uses recombination for this purpose; the mechanism in humans is not known. Some insect and plant species, on the other hand, do not use telomerase as their primary mechanism for maintaining chromosome length. Drosophila makes use of specific retrotransposons for this purpose, while other dipterans use recombination. We summarize here the current knowledge of these alternative telomere elongation mechanisms.

Animals↗

PubMed1997

Chromosome end elongation by recombination in the mosquito Anopheles gambiae.

One of the functions of telomeres is to counteract the terminal nucleotide loss associated with DNA replication. While the vast majority of eukaryotic organisms maintain their chromosome ends via telomerase, an enzyme system that generates short, tandem repeats on the ends of chromosomes, other mechanisms such as the transposition of retrotransposons or recombination can also be used in some species. Chromosome end regression and extension were studied in a medically important mosquito, the malaria vector Anopheles gambiae, to determine how this dipteran insect maintains its chromosome ends. The insertion of a transgenic pUChsneo plasmid at the left end of chromosome 2 provided a unique marker for measuring the dynamics of the 2L telomere over a period of about 3 years. The terminal length was relatively uniform in the 1993 population with the chromosomes ending within the white gene sequence of the inserted transgene. Cloned terminal chromosome fragments did not end in short repeat sequences that could have been synthesized by telomerase. By late 1995, the chromosome ends had become heterogeneous: some had further shortened while other chromosomes had been elongated by regenerating part of the integrated pUChsneo plasmid. A model is presented for extension of the 2L chromosome by recombination between homologous 2L chromosome ends by using the partial plasmid duplication generated during its original integration. It is postulated that this mechanism is also important in wild-type telomere elongation.

Animals↗

PubMed1996

Catecholamine metabolism and in vitro induction of premature cuticle melanization in wild type and pigmentation mutants of Drosophila melanogaster.

The major pathway leading to adult cuticle melanization in Drosophila melanogaster has been investigated by a combination of biochemical and genetic approaches. By comparing catecholamine pools in newly emerged flies and in frass (excreta) collected 1 to 4 days after eclosion from wild type with those obtained from several pigmentation mutants, the major flow of catecholamines through the pathway to an unidentified final catabolite was determined. We also demonstrate that incubation with dopamine in vitro induces premature melanization in wild type unpigmented pharate adults several hours before the developmentally programmed onset of melanization, supporting the hypothesis that the availability of catecholamines may be the limiting factor determining the onset of melanization and that the major enzymatic activities that act downstream of dopa decarboxylase in the pathway are deposited into the cuticle before pigmentation begins. In vitro melanization studies with various pigmentation mutants that are associated with critical enzymatic steps in Drosophila catecholamine metabolism are consistent with their proposed function and suggest a central role of N-beta-alanyldopamine in adult cuticle pigmentation.

Alleles↗

PubMed1996

Molecular characterization of the Anopheles gambiae 2L telomeric region via an integrated transgene.

A Drosophila P-element derivative (pUChsneo) integrated into the telomeric region of the left arm of the second chromosome of Anopheles gambiae was used to clone the proximally flanking An. gambiae sequences. Molecular analyses revealed that the pUChsneo construct was partially duplicated and had integrated into a subterminal minisatellite. This satellite has a repeat unit of 820 bp and is located exclusively at the tip of 2L. No sequence similarity to subterminal minisatellites from other dipterans was detected, but some structural features such as tandem subrepeats are shared. The end of the chromosome was mapped with respect to restriction sites in pUChsneo at approximately generation 100 after the integration event. Considering inevitable terminal nucleotide loss due to incomplete DNA replication, we conclude that the chromosome end must have undergone a dramatic elongation process since it was mapped in generation 23.

Animals↗

PubMed1995

DNA organization and polymorphism of a wild-type Drosophila telomere region.

Telomeres at the ends of linear chromosomes of eukaryotes protect the chromosome termini from degradation and fusion. While telomeric replication/elongation mechanisms have been studied extensively, the functions of subterminal sequences are less well understood. In general, subterminal regions can be quite polymorphic, varying in size from organism to organism, and differing among chromosomes within an organism. The subterminal regions of Drosophila melanogaster are not well characterized today, and it is not known which and how many different components they contain. Here we present the molecular characterization of DNA components and their organization in the subterminal region of the left arm of chromosome 2 of the Oregon RC wild-type strain of D. melanogaster, including a minisatellite with a 457bp repeat length. Two distinct polymorphic arrangements at 2L were found and analyzed, supporting the Drosophila telomere elongation model by retrotransposition. The high incidence of terminal chromosome deficiencies occurring in natural Drosophila populations is discussed in view of the telomere structure at 2L.

Animals↗

PubMed1995

The unusual telomeres of Drosophila.

The telomeres of most eukaryotes contain short, simple repeats that are highly conserved. Drosophila, on the other hand, does not have such sequences, but carries at the ends of its chromosomes one or more LINE-like retrotransposable elements. Instead of elongation by telomerase, incomplete DNA replication at the termini of Drosophila chromosomes is counterbalanced by transposition of these elements at high frequency specifically to the termini. These transposable elements are not responsible for distinguishing telomeric ends in Drosophila from broken chromosome ends; the structure performing this function is not yet known. Proximal to the terminal array of transposable elements are regions of tandem repeats that are structurally, and probably functionally, analogous to the subterminal regions in other eukaryotes.

Animals↗

PubMed1994

Mapping a mutator, mu2, which increases the frequency of terminal deletions in Drosophila melanogaster.

A mutator, mu2, in Drosophila melanogaster has been identified recently that potentiates the recovery of terminal deficiencies. The deleted chromosomes behave as if they had been capped; that is, they are protected from degradation and from fusion with other chromosome fragments. The mutator maps near the telomere on the left arm of chromosome 3. Using the selectable marker Aprt, 150 deficiencies for region 62 of the cytological map have been recovered. These deficiencies identify the map position of mu2 as 62B11-C1. A yeast artificial chromosome spanning this region has been subcloned into lambda phage, and the positions of deficiency breakpoints on either side of the mu2 gene have been identified within the subclones. These positions limit the location of the left end of the gene to a 23 kb region. In the course of these experiments, three additional, presumptive mutant alleles were identified, suggesting that other mutator alleles remain undiscovered in many standard laboratory stocks.

Adenine Phosphoribosyltransferase↗

PubMed1994

Comparison of two active HeT-A retroposons of Drosophila melanogaster.

HeT-A elements are Drosophila melanogaster LINE-like retroposons that transpose to broken chromosome ends by attaching themselves with an oligo(A) tail. Since this family of elements is believed to be involved in the vital function of telomere elongation in Drosophila, it is important to understand their transposition mechanism and the molecular aspects of activity. By comparison of several elements we have defined here the unit length of HeT-A elements to be approximately 6 kb. Also, we studied an active HeT-A element that had transposed very recently to the end of a terminally deleted X chromosome. The 12 kb of newly transposed DNA consisted of a tandem array of three different HeT-A elements joined by oligo(A) tails to each other and to the chromosome end broken in the yellow gene. Such an array may have transposed as a single unit or resulted from rapid successive transpositions of individual HeT-A elements. By sequence comparison with another recently transposed HeT-A element, conserved domains in the single open reading frame (ORF), encoding a gag-like polypeptide, of these elements were defined. We conclude that for transposition an intact ORF is required in cis, while the reverse transcriptase is not encoded on the HeT-A element but is provided in trans. This would make HeT-A elements dependent on an external reverse transcriptase for transposition and establish control of the genome over the activity of HeT-A elements. This distinguishes the Drosophila HeT-A element, which has been implicated in Drosophila telomere elongation, from the other, 'selfish' LINE-like elements.

Amino Acid Sequence↗

PubMed1994

Telomeric repeat sequences.

Chromosomes not only carry transcribed genes and their regulatory DNA sequences, but also contain regions that are required for the stability and maintenance of the chromosome as a unit. These include centromeres, telomeres and origins of replication. It is clear for replication origins and centromeres that the positions of these chromosomal organelles are determined by sites of the appropriate DNA sequences, but also that functional performance requires one or more contributing proteins. Telomeres are also structurally complex, with one or more DNA components, including simple telomeric repeats and more complex telomere-associated sequences, as well as one or more specific proteins that recognize these sequences. Accumulating evidence suggests that the simple telomeric repeats are required in most, but not all species, although they are not sufficient to determine the chromosomal position of a telomere.

Animals↗

PubMed1993

The genomic organization of HeT-A retroposons in Drosophila melanogaster.

Members of the Drosophila HeT-A family of transposable elements are LINE-like retroposons that are found at telomeres and in centric heterochromatin. We recently characterized an active HeT-A element that had transposed to a broken chromosome end fewer than nine generations before it was isolated. The sequence arrangement of this element, called 9D4, most likely represents the organization of an actively transposing member of the HeT-A family. Here we assess the degree of divergence among members of the HeT-A family and test a model of telomere length maintenance based on HeT-A transposition. The region containing the single open reading frame of this element appears to be more highly conserved than the non-coding regions. The HeT-A element has been implicated in the Drosophila telomere elongation process, because frequent transpositions to chromosome ends are sufficient to counter-balance nucleotide loss due to incomplete DNA replication. The proposed elongation model and the hypothetical mechanism of HeT-A transposition predict a predominant orientation of HeT-A elements with their oligo (A) tails facing proximally at chromosome ends, as well as the existence of irregular tandem arrays of HeT-A elements at chromosome ends resulting from transposition of new HeT-A elements onto chromosome ends with existing elements. Twenty-nine different HeT-A fragments were isolated from directional libraries that were enriched in terminal DNA fragments. Sequence analyses of these fragments and comparisons with the organization of the HeT-A element, 9D4, fit these two predictions and support the model of Drosophila telomere elongation by transposition of HeT-A elements.

Animals↗

PubMed1992

Activation of maternal centrosomes in unfertilized sea urchin eggs.

Centrosomes are undetectable in unfertilized sea urchin eggs, and normally the sperm introduces the cell's microtubule-organizing center (MTOC) at fertilization. However, artificial activation or parthenogenesis triggers microtubule assembly in the unfertilized egg, and this study explores the reappearance and behavior of the maternal centrosome. During activation with A23187 or ammonia, microtubules appear first at the cortex; centrosomal antigen is detected diffusely throughout the entire cytoplasm. Later, the centrosome becomes more distinct and organizes a radial microtubule shell, and eventually a compact centrosome at the egg center organizes a monaster. In these activated eggs, centrosomes undergo cycles of compaction and decompaction in synchrony with the chromatin, which also undergoes cycles of condensation and decondensation. Parthenogenetic activation with heavy water (50% D2O) or the microtubule-stabilizing drug taxol (10 microM) induces numerous centrosomal foci in the unfertilized sea urchin egg. Within 15 min after incubation in D2O, numerous fine centrosomal foci are detected, and they organize a connected network of numerous asters which fill the entire egg. Taxol induces over 100 centrosomal foci by 15 min after treatment, which organize a corresponding number of asters. The centrosomal material in either D2O- or taxol-treated eggs aggregates with time to form fewer but denser foci, resulting in fewer and larger asters. Fertilization of eggs pretreated with either D2O or taxol shows that the paternal centrosome is dominant over the maternal centrosome. The centrosomal material gradually becomes associated with the enlarged sperm aster. These experiments demonstrate that maternal centrosomal material is present in the unfertilized egg, likely as dispersed undetectable material, which can be activated without paternal contributions. At fertilization, paternal centrosomes become dominant over the maternal centrosomal material.

Ammonia↗

PubMed1992

Frequent transpositions of Drosophila melanogaster HeT-A transposable elements to receding chromosome ends.

HeT-A elements are a new family of transposable elements in Drosophila that are found exclusively in telomeric regions and in the pericentric heterochromatin. Transposition of these elements onto broken chromosome ends has been implicated in chromosome healing. To monitor the fate of HeT-A elements that had attached to broken ends of the X chromosome, we examined individual X chromosomes from a defined population over a period of 17 generations. The ends of the X chromosomes with new HeT-A additions receded at the same rate as the broken ends before the HeT-A elements attached. In addition, some chromosomes, approximately 1% per generation, had acquired new HeT-A sequences of an average of 6 kb at their ends with oligo(A) tails at the junctions. Thus, the rate of addition of new material per generation matches the observed rate of terminal loss (70-75 bp) caused by incomplete replication at the end of the DNA molecule. One such recently transposed HeT-A element which is at least 12 kb in length has been examined in detail. It contains a single open reading frame of 2.8 kb which codes for a gag-like protein.

Amino Acid Sequence↗