Heterochromatic distribution of HeT-A- and TART-like sequences in several Drosophila species.

Drosophila melanogaster telomeres contain arrays of two non-LTR retrotransposons called HeT-A and TART. Previous studies have shown that HeT-A- and TART-like sequences are also located at non-telomeric sites in the Y chromosome heterochromatin. By in situ hybridization experiments, we mapped TART sequences in the h16 region of the long arm close to the centromere of the Y chromosome of D. melanogaster. HeT-A sequences were localized in two different regions on the Y chromosome, one very close to the centromere in the short arm (h18-h19) and the other in the long arm (h13-h14). To assess a possible heterochromatic location of TART and HeT-A elements in other Drosophila species, we performed in situ hybridization experiments, using both TART and HeT-A probes, on mitotic and polytene chromosomes of D. simulans, D. sechellia, D. mauritiana, D. yakuba and D. teissieri. We found that TART and HeT-A probes hybridize at specific heterochromatic regions of the Y chromosome in all Drosophila species that we analyzed.

Cis-effects of heterochromatin on heterochromatic and euchromatic gene activity in Drosophila melanogaster.

Chromosomal rearrangements that juxtapose heterochromatin and euchromatin can result in mosaic inactivation of heterochromatic and euchromatic genes. This phenomenon, position effect variegation (PEV), suggests that heterochromatic and euchromatic genes differ in their regulatory requirements. This report describes a novel method for mapping regions required for heterochromatic genes, and those that induce PEV of a euchromatic gene. P transposase mutagenesis was used to generate derivatives of a translocation that variegated for the light+ (lt+) gene and carried the euchromatic white+ (w+) gene on a transposon near the heterochromatin-euchromatin junction. Cytogenetic and genetic analyses of the derivatives showed that P mutagenesis resulted in deletions of several megabases of heterochromatin. Genetic and molecular studies showed that the derivatives shared a euchromatic breakpoint but differed in their heterochromatic breakpoint and their effects on seven heterochromatic genes and the w+ gene. Heterochromatic genes differed in their response to deletions. The lt+ gene was sensitive to the amount of heterochromatin at the breakpoint but the heterochromatic 40Fa gene was not. The severity of variegated w+ phenotype did not depend on the amount of heterochromatin in cis, but varied with local heterochromatic environment. These data are relevant for considering mechanisms of PEV of both heterochromatic and euchromatic genes.

Developmental genetical analysis and molecular cloning of the abnormal oocyte gene of Drosophila melanogaster.

Studies of the abnormal oocyte (abo) gene of Drosophila melanogaster have previously been limited to the analysis of a single mutant allele, abnormal oocyte1 (abo1). The abo1 mutation causes a maternal-effect lethality that can be partially rescued zygotically by the abo+ allele and by increasing the dosage of specific regions of heterochromatin denoted ABO. This report describes the properties of abo2, a new P-element-induced allele that allowed us to reexamine the nature of maternal-effect defect. Comparisons of the phenotype of progeny of abo1/abo1 and abo1/abo2 females show that the preblastoderm lethality previously described as a component of the abo mutant maternal effect results from a recessive fertilization defect associated with the abo1 chromosome. We demonstrate here that the abo-induced maternal effect lethality occurs predominately late in embryogenesis after cuticle deposition but before hatching. The phenocritical period for zygotic rescue by heterochromatin coincides with this period of late embryogenesis. We have used the abo2 mutation to map and molecularly clone the gene. We show that the abo gene is located in the 32C cytogenetic interval and identify the putative abo transcript from mRNA isolated from adult females. Using germline transformation, we show that a 9-kb genomic fragment to which the transcript maps, partially fulfills requirement for maternal and zygotic abo+ function.

Gynecomastia and the complete circumareolar approach in the surgical management of skin redundancy.

Gynecomastia is a benign enlargement of the male breast due to a physiological or pathological factor that interferes with the balance between estrogens and androgens in the serum. Gynecomastia itself requires no treatment unless the persistent enlargement of the male breast is a source of embarrassment and/or distress for the adolescent or adult man. The indications for the surgical treatment of gynecomastia are founded on two main objectives: (1) the restoration of male chest shape and (2) diagnostic evaluation of suspected breast lesions. The diagnostic evaluation begins with an adequate history and a thorough breast examination helped by laboratory tests and instrumental research. Several approaches for surgical treatment have been described in the literature. Some problems arise in patients who have significant enlargement and ptosis of the breast that will require skin reduction and in some patients requiring nipple-areola complex reduction. The authors believe that the complete circumareolar technique with purse-string suture creates the best aesthetic results, with fewer complications, in patients with moderate and severe ptotic glandular breast enlargements that have skin redundancy combined with areolar enlargement. From 1995 through 1999, a total of 10 male patients with moderate to severe gynecomastia were treated surgically using a complete circumareolar approach. All patients achieved a good aesthetic contour of the chest. Only two patients required a revision of the circumareolar scar to correct postoperative enlargement.

Genotypic and phenotypic diversity of Lactobacillus rossiae strains isolated from sourdough.

AIM: To characterize the genetic and phenotypic diversity of 33 strains of Lactobacillus rossiae. METHODS AND RESULTS: Genotypic identification was carried out by partial 16S rRNA gene sequence analysis. Genetic diversity was evaluated by RAPD-PCR analysis. Phenotypic diversity was evaluated through fermentative profile by Biolog system, proteinase and peptidase activities using synthetic substrates, and acidification capacity and amino acid profile during sourdough fermentation. The genetic analyses excluded clonal relatedness among the strains used. A large phenotypic diversity was found. It mainly concerned the capacity to use carbon sources available in sourdough during fermentation, the quotient of fermentation and the peptidase activities, especially towards proline containing synthetic substrates. The free amino acid profiles differed either for the total concentration or for the type of amino acids. With a few exceptions, proteinase activity towards wheat albumins and globulins was weak. CONCLUSIONS: Overall, no relationships between genetic and physiological analyses were found, and the strains examined showed a marked genetic and phenotypic heterogeneity. L. rossiae strains had interesting properties for application in sourdough fermentation. Although some strains combined several technological traits, the association of more strains seemed to be a requisite to get optimal sourdough characteristics. SIGNIFICANCE AND IMPACT OF THE STUDY: It represents the first characterization of the diversity within the L. rossiae species. Besides, it may represent an example of computerized analysis of genotypic and phenotypic information to select strains for improving sourdough characteristics.

Carnitine suppression of position-effect variegation in Drosophila melanogaster.

Carnitine is a well-known naturally occurring compound, very similar to butyrate, with an essential role in intermediary metabolism mainly at the mitochondrial level. Since butyrate inhibits the enzyme histone deacetylase and is capable of suppressing position-effect variegation in Drosophila melanogaster, we tested a further possible function of carnitine in the nucleus, using an assay for the suppression of position-effect variegation. We tested three physiological forms of carnitine (L-carnitine, L-propionylcarnitine, L-acetylcarnitine) for the ability to suppress two different chromosomal rearrangements, inducing variegation of the white+ and brown+ genes. The results show that the carnitine derivatives are capable of suppressing the position-effect variegation, albeit with different efficiencies. The carnitine derivatives interact lethally with Su-var(2-)1(01), a mutation that induces hyperacetylation of histones, whilst hyperacetylated histones accumulated in both the nuclei of HeLa cells and Drosophila polytene chromosomes treated with the same compounds. These results strongly suggest that the carnitine derivatives suppress position-effect variegation by a mechanism similar to that of butyrate. It is suggested that carnitines may have a functional role in the nucleus, probably at the chromatin level.

The heterochromatin protein 1 prevents telomere fusions in Drosophila.

HP1 (Heterochromatin protein 1) is a conserved, non-histone chromosomal protein that is best known for its preferential binding to pericentric heterochromatin and its role in position effect variegation in Drosophila. Using immunolocalization, we show that HP1 is a constant feature of the telomeres of interphase polytene and mitotic chromosomes. This localization does not require the presence of telomeric retrotransposons, since HP1 is also detected at the ends of terminally deleted chromosomes that lack these elements. Importantly, larvae expressing reduced or mutant versions of HP1 exhibit aberrant chromosome associations and multiple telomeric fusions in neuroblast cells, imaginal disks, and male meiotic cells. Taken together, these results provide evidence that HP1 plays a functional role in mediating normal telomere behavior in Drosophila.

The maternal effect gene, abnormal oocyte (abo), of Drosophila melanogaster encodes a specific negative regulator of histones.

The abnormal oocyte (abo) gene of Drosophila melanogaster is a peculiar maternal effect gene whose mutations cause a maternal-effect lethality that can be rescued by specific regions of heterochromatin during early embryogenesis. Here we show that abo encodes an evolutionary conserved chromosomal protein that localizes exclusively to the histone gene cluster and binds to the regulatory regions of such genes. We also show a significant increase of histone transcripts in eggs of abo mutant mothers and a partial rescue of the abo maternal-effect defect by deficiencies of the histone gene cluster. On the basis of these results, we suggest that the Abo protein functions specifically as a negative regulator of histone transcription and propose a molecular model to account for the ability of heterochromatin to partially rescue the abo maternal-effect defect. Our model proposes that increased doses of specific heterochromatic regions titrate out abnormally high levels of histones present in embryos from mutant abo mothers and that a balanced pool of histones is critical for normal embryogenesis in Drosophila.

Heterochromatin protein 1 binds transgene arrays.

Heterochromatin protein 1 (HP1) of Drosophila and its homologs in vertebrates are key components of constitutive heterochromatin. Here we provide cytological evidence for the presence of heterochromatin within a euchromatic chromosome arm by immunolocalization of HP1 to the site of a silenced transgene repeat array. The amount of HP1 associated with arrays in polytene chromosomes is correlated with the array size. Inverted transposons within an array or increased proximity of an array to blocks of naturally occurring heterochromatin may increase transgene silencing without increasing HP1 labeling. Less dense anti-HP1 labeling is found at transposon arrays in which there is no transgene silencing. The results indicate that HP1 targets the chromatin of transposon insertions and binds more densely at a site with repeated sequences susceptible to heterochromatin formation.

Transposable elements are stable structural components of Drosophila melanogaster heterochromatin.

We determined the distribution of 11 different transposable elements on Drosophila melanogaster mitotic chromosomes by using high-resolution fluorescent in situ hybridization (FISH) coupled with charge-coupled device camera analysis. Nine of these transposable elements (copia, gypsy, mdg-1, blood, Doc, I, F, G, and Bari-1) are preferentially clustered into one or more discrete heterochromatic regions in chromosomes of the Oregon-R laboratory stock. Moreover, FISH analysis of geographically distant strains revealed that the locations of these heterochromatic transposable element clusters are highly conserved. The P and hobo elements, which are likely to have invaded the D. melanogaster genome at the beginning of this century, are absent from Oregon-R heterochromatin but clearly exhibit heterochromatic clusters in certain natural populations. Together these data indicate that transposable elements are major structural components of Drosophila heterochromatin, and they change the current views on the role of transposable elements in host genome evolution.

Cloning and molecular characterization of three ubiquitin fusion degradation 1 (Ufd1) ortholog genes from Xenopus laevis, Gallus gallus and Drosophila melanogaster.

The yeast ubiquitin fusion degradation 1 (Ufd1) protein is involved in a degradation pathway for ubiquitin fused products. The human ortholog gene (UFD1-like, UFD1L) is deleted in patients affected by the DiGeorge/velocardiofacial syndromes. We report the cloning of UFD1L orthologs from Drosophila melanogaster (dufd1l), Xenopus laevis and Gallus gallus. The 1,125-bp Drosophila cDNA encodes a protein of 316 amino acids, showing 60% identity with the human and murine proteins. The identity to the G. gallus, X. laevis, C. elegans and S. cerevisiae proteins is 95%, 83%, 32%, and 36%, respectively. Northern expression data in Drosophila indicate that dufd1l is expressed through embryonic, larval and pupal development, as well as in the adult fly.

Interaction systems between heterochromatin and euchromatin in Drosophila melanogaster.

The constitutive heterochromatin is still one of the major unsolved problems in genetics. In Drosophila melanogaster three genetic systems involving specific interactions between heterochromatic and euchromatic genetic elements are known: the Segregation Distortion, the crystal-Stellate and the abo-ABO systems. The genetic and molecular analysis of each system will allow the identification of all the components and the elucidation of the mechanisms underlying their interactions. The results of this analysis should provide insights into the biological significance of heterochromatin and into the evolutionary forces that result in the maintainance and stability of this enigmatic genetic material.

The ISWI chromatin-remodeling protein is required for gene expression and the maintenance of higher order chromatin structure in vivo.

Drosophila ISWI, a highly conserved member of the SWI2/SNF2 family of ATPases, is the catalytic subunit of three chromatin-remodeling complexes: NURF, CHRAC, and ACF. To clarify the biological functions of ISWI, we generated and characterized null and dominant-negative ISWI mutations. We found that ISWI mutations affect both cell viability and gene expression during Drosophila development. ISWI mutations also cause striking alterations in the structure of the male X chromosome. The ISWI protein does not colocalize with RNA Pol II on salivary gland polytene chromosomes, suggesting a possible role for ISWI in transcriptional repression. These findings reveal novel functions for the ISWI ATPase and underscore its importance in chromatin remodeling in vivo.