Jun mediates Frizzled-induced R3/R4 cell fate distinction and planar polarity determination in the Drosophila eye.

Jun acts as a signal-regulated transcription factor in many cellular decisions, ranging from stress response to proliferation control and cell fate induction. Genetic interaction studies have suggested that Jun and JNK signaling are involved in Frizzled (Fz)-mediated planar polarity generation in the Drosophila eye. However, simple loss-of-function analysis of JNK signaling components did not show comparable planar polarity defects. To address the role of Jun and JNK in Fz signaling, we have used a combination of loss- and gain-of-function studies. Like Fz, Jun affects the bias between the R3/R4 photoreceptor pair that is critical for ommatidial polarity establishment. Detailed analysis of jun(-) clones reveals defects in R3 induction and planar polarity determination, whereas gain of Jun function induces the R3 fate and associated polarity phenotypes. We find also that affecting the levels of JNK signaling by either reduction or overexpression leads to planar polarity defects. Similarly, hypomorphic allelic combinations and overexpression of the negative JNK regulator Puckered causes planar polarity eye phenotypes, establishing that JNK acts in planar polarity signaling. The observation that Dl transcription in the early R3/R4 precursor cells is deregulated by Jun or Hep/JNKK activation, reminiscent of the effects seen with Fz overexpression, suggests that Jun is one of the transcription factors that mediates the effects of fz in planar polarity generation.

Tirant is a new member of the gypsy family of retrotransposons in Drosophila melanogaster.

In this paper, we propose a consensus sequence for a putative complete Tirant retrotransposon. Several defective copies, as well as relevant sequences available in databases have been analyzed. The putative complete Tirant element is 8533 bp long, and presents all the structural features of a retrovirus-like transposable element of the gypsy family. It contains three ORFs (open reading frames) that encode putative products resembling the retroviral Gag, Pol, and Env proteins. Southern blot analyses show that complete and defective Tirant elements are widespread in Drosophila melanogaster. The different hybridization patterns observed in several natural populations of this species suggest that Tirant is an active element.

Dnop56, a Drosophila gene homologous to the yeast nucleolar NOP56 gene.

Small nucleolar RNAs (snoRNAs) are trans-acting factors involved in maturation of rRNA and have been classified into Box C/D and Box H/ACA families. Most of the snoRNAs occur as ribonucleoprotein complexes with snoRNA-associated proteins (snoRNPs). All Box C/D snoRNAs in yeast form complexes with Nop1p, Nop56p and Nop58p. Similarly, it has been reported that Box H/ACA-containing snoRNAs form complexes with yeast Gar1p. Nop56p and Nop58p homologs have been described in several species. Here we report the isolation and molecular characterization of the Dnop56 genes from D. melanogaster and D. subobscura which show a very similar structure. Drosophila Nop56p proteins contain lysine-rich regions at their carboxy-terminus, and show a high degree of similarity to other Nop56p proteins from different organisms. Phylogenetic relationships among these proteins and other snoRNPs have been established.

The Drosophila STE20-like kinase misshapen is required downstream of the Frizzled receptor in planar polarity signaling.

The Drosophila misshapen (msn) gene is a member of the STE20 kinase family. We show that msn acts in the Frizzled (Fz) mediated epithelial planar polarity (EPP) signaling pathway in eyes and wings. Both msn loss- and gain-of-function result in defective ommatidial polarity and wing hair formation. Genetic and biochemical analyses indicate that msn acts downstream of fz and dishevelled (dsh) in the planar polarity pathway, and thus implicates an STE20-like kinase in Fz/Dsh-mediated signaling. This demonstrates that seven-pass transmembrane receptors can signal via members of the STE20 kinase family in higher eukaryotes. We also show that Msn acts in EPP signaling through the JNK (Jun-N-terminal kinase) module as it does in dorsal closure. Although at the level of Fz/Dsh there is no apparent redundancy in this pathway, the downstream effector JNK/MAPK (mitogen-activated protein kinase) module is redundant in planar polarity generation. To address the nature of this redundancy, we provide evidence for an involvement of the related MAP kinases of the p38 subfamily in planar polarity signaling downstream of Msn.

GEM, a cluster of repetitive sequences in the Drosophila subobscura genome.

GEM is a new family of repetitive sequences detected in the D. subobscura genome. Two of the four described GEM elements encompass a heterogeneous central module, with no detectable ORF, flanked by two long inverted repeats. These elements are composed of a set of repetitive modules, which are inverted repeat (IR), direct repeat (DR), palindromic sequence (PS), long sequence (LS) and short sequence (SS). These five modules can be found either clustered or dispersed as single modules in the D. subobscura genome, in euchromatic and heterochromatic regions. In addition to the 3' region of Adh retrosequences, single IR and LS blocks were found associated with the promoter region of different genes, in particular, LS-like blocks have also been found associated with functional genes in D. melanogaster and D. virilis. Conversely, the DR block is highly similar to satellite DNAs from some other species of the obscura group. In addition, GEM elements share some structural features with IS elements described in different Drosophila species. It is likely that both GEM and IS sequences would be vestiges of an ancestral transposable element.

Drosophila tumor suppressor PTEN controls cell size and number by antagonizing the Chico/PI3-kinase signaling pathway.

The human tumor suppressor gene PTEN encodes a putative cytoskeleton-associated molecule with both protein phosphatase and phosphatidylinositol 3,4,5-trisphosphate (PIP3) 3-phosphatase activities. In cell culture, the lipid phosphatase activity of this protein is involved in regulating cell proliferation and survival, but the mechanism by which PTEN inhibits tumorigenesis in vivo is not fully established. Here we show that the highly evolutionarily conserved Drosophila PTEN homolog, DPTEN, suppresses hyperplastic growth in flies by reducing cell size and number. We demonstrate that DPTEN modulates tissue mass by acting antagonistically to the Drosophila Class I phosphatidylinositol 3-kinase, Dp110, and its upstream activator Chico, an insulin receptor substrate homolog. Surprisingly, although DPTEN does not generally affect cell fate determination, it does appear to regulate the subcellular organization of the actin cytoskeleton in multiple cell types. From these data, we propose that DPTEN has a complex role in regulating tissue and body size. It acts in opposition to Dp110 to control cell number and growth, while coordinately influencing events at the cell periphery via its effects on the actin cytoskeleton.

Molecular evolution of P transposable elements in the Genus drosophila. II. The obscura species group.

A phylogenetic analysis of P transposable elements in the Drosophila obscura species group is described. Multiple P sequences from each of 10 species were obtained using PCR primers that flank a conserved region of exon 2 of the transposase gene. In general, the P element phylogeny is congruent with the species phylogeny, indicating that the dominant mode of transmission has been vertical, from generation to generation. One manifestation of this is the distinction of P elements from the Old World obscura and subobscura subgroups from those of the New World affinis subgroup. However, the overall distribution of elements within the obscura species group is not congruent with the phylogenetic relationships of the species themselves. There are at least four distinct subfamilies of P elements, which differ in sequence from each other by as much as 34%, and some individual species carry sequences belonging to different subfamilies. P sequences from D. bifasciata are particularly interesting. These sequences belong to two subfamilies and both are distinct from all other P elements identified in this survey. Several mechanisms are postulated to be involved in determining phylogenetic relationships among P elements in the obscura group. In addition to vertical transmission, these include retention of ancestral polymorphisms and horizontal transfer by an unknown mating-independent mechanism.

saliva, a new Drosophila gene expressed in the embryonic salivary glands with homologues in plants and vertebrates.

saliva (slv) transcription begins at the salivary gland placodes and continues on throughout development as salivary glands invaginate and reach their final location and morphology. saliva is located cytogenetically in 76A/B, and encodes a 226-amino-acid protein with four hydrophobic domains. A Northern blot detects a 1.6-kb transcript throughout development. Database similarity searches reveal homology to proteins from Caenorhabditis, Lilium, Medicago and mouse.

Dishevelled activates JNK and discriminates between JNK pathways in planar polarity and wingless signaling.

Frizzled family proteins have been described as receptors of Wnt signaling molecules. In Drosophila, the two known Frizzled proteins are associated with distinct developmental processes. Genesis of epithelial planar polarity requires Frizzled, whereas Dfz2 affects morphogenesis by wingless-mediated signaling. Dishevelled is required in both signaling pathways. Here, we use genetic and overexpression assays to show that Dishevelled activates JNK cascades. Rescue analysis reveals different protein domain requirements in Dishevelled for the two pathways; the C-terminal DEP domain is essential to rescue planar polarity defects and induce JNK signaling. Furthermore, the planar polarity-specific dsh1 allele is mutated in the DEP domain. Our results indicate that different Wnt/Fz signals activate distinct intracellular pathways, and Dishevelled discriminates among them by distinct domain interactions.

The muscleblind gene participates in the organization of Z-bands and epidermal attachments of Drosophila muscles and is regulated by Dmef2.

We report the embryonic phenotype of muscleblind (mbl), a recently described Drosophila gene involved in terminal differentiation of adult ommatidia. mbl is a nuclear protein expressed late in the embryo in pharyngeal, visceral, and somatic muscles, the ventral nerve cord, and the larval photoreceptor system. All three mbl alleles studied exhibit a lethal phenotype and die as stage 17 embryos or first instar larvae. These larvae are partially paralyzed, show a characteristically contracted abdomen, and lack striation of muscles. Our analysis of the somatic musculature shows that the pattern of muscles is established correctly, and they form morphologically normal synapses. Ultrastructural analysis, however, reveals two defects in the terminal differentiation of the muscles: inability to differentiate Z-bands in the sarcomeric apparatus and reduction of extracellular tendon matrix at attachment sites to the epidermis. Failure to differentiate both structures could explain the partial paralysis and contracted abdomen phenotype. Analysis of mbl expression in embryos that are either mutant for Dmef2 or ectopically express Dmef2 places mbl downstream of Dmef2 function in the myogenic differentiation program. mbl, therefore, may act as a critical element in the execution of two Dmef2-dependent processes in the terminal differentiation of muscles.

muscleblind, a gene required for photoreceptor differentiation in Drosophila, encodes novel nuclear Cys3His-type zinc-finger-containing proteins.

We have isolated the embryonic lethal gene muscleblind (mbl) as a suppressor of the sev-svp2 eye phenotype. Analysis of clones mutant for mbl during eye development shows that it is autonomously required for photoreceptor differentiation. Mutant cells are recruited into developing ommatidia and initiate neural differentiation, but they fail to properly differentiate as photoreceptors. Molecular analysis reveals that the mbl locus is large and complex, giving rise to multiple different proteins with common 5' sequences but different carboxy termini. Mbl proteins are nuclear and share a Cys3His zinc-finger motif which is also found in the TIS11/NUP475/TTP family of proteins and is highly conserved in vertebrates and invertebrates. Functional analysis of mbl, the observation that it also dominantly suppresses the sE-Jun(Asp) gain-of-function phenotype and the phenotypic similarity to mutants in the photoreceptor-specific glass gene suggest that mbl is a general factor required for photoreceptor differentiation.

Structure and organization of the P element related sequences in Drosophila madeirensis.

The P element homologous sequences of the two closely related species Drosophila guanche and Drosophila subobscura represent a very special case of transposable-element derivatives. Although they have lost the regions known to be essential for P transposition by random mutations, all of them have selectively conserved the coding capacity for "P-repressor-like" proteins during the past few millions years. In both species, they are tandemly amplified in a single euchromatic gene cluster at equivalent chromosomal positions. In contrast, Drosophila madeirensis, an endemic species that is very closely related to both D. subobscura and D. guanche, harbours an additional P homologous site. Several mechanisms can be invoked to explain the generation of the new site in this species. In this work we present several molecular and cytological data in order to elucidate the possible evolutionary origin of the P derivatives of D. madeirensis.

Sequencing analysis of a 4.1 kb subtelomeric region from yeast chromosome IV identifies HXT15, a new member of the hexose transporter family.

The DNA sequence of a 4.1 kb region of Saccharomyces cerevisiae chromosome IV was determined. This region contains a single open reading frame which codes for a member of the hexose transporter family. This new gene has been named HXT15 according to yeast gene data bases.

Tirant: a new retrotransposon-like element in Drosophila melanogaster.

In this paper we report a new retrotransposon-like element of Drosophila melanogaster called Tirant. This sequence is moderately repeated in the genome of this species and it has been found to be widely dispersed throughout its distribution area. From Southern blot and in situ analyses, this sequence appears to be mobile in D. melanogaster, since its chromosome location and the hybridization patterns vary among the different strains analyzed. In this way, partial sequencing of Tirant ends suggests that it is a retrotransposon, since it is flanked by two LTRs. The presence of sequences homologous to Tirant has been also investigated in 28 species of the genus Drosophila by means of Southern analyses. These sequences were only detected in species from melanogaster and obscura groups. These data suggest that ancestral sequences of Tirant appeared after the Sophophora radiation and before the divergence of those groups.

Clinical and genetic aspects of two Spanish families with autosomal dominant retinitis pigmentosa (ADRP)

A study was made of two families with autosomal dominant retinitis pigmentosa (ADRP) from Valencia (Spain). One family (ADRP15) was found to have mutation in codon 114 of the rhodopsin gene that led to a substitution of a glycine for an aspartic acid. The second family (ADRP7) substituted an aspartic acid for valine in codon 173 of the peripherin-RDS gene. Rhodopsin is involved in 25% of ADRP cases and many mutations of this gene have been described as causing different forms of the disease, with variable severity and age at onset. ADRP has been classified as RP with a milder symptom evolution, a typical RP fundus pattern, and macular involvement occurring after the second decade of life. Peripherin-RDS gene mutations lead to RP or other retinopathies. Furthermore, two mutations in codon 172 have been described as causing macular dystrophy. In ADRP7, a mutation in neighboring codon 173 produced RP with an atypical fundus pattern and macular involvement within the first decade of life. These observations confirm the established clinical and genetic heterogeneity involved in this form of RP.

Structure and expression of clustered P element homologues in Drosophila subobscura and Drosophila guanche.

Sequence relationships and functional aspects were analysed in the P element homologues of Drosophila subobscura (Ds) and D. guanche (Dg). In both species, the P homologues are clustered at a single genomic position. They lack the characteristic terminal structures of actively transposing P elements, but they have the coding capacity for a 66-kDa 'repressor-like' protein. Two different types of cluster units (G-type and A-type) can be distinguished. The A-type unit, which is present in multiple copies, is transcribed in adult flies. In contrast, the G-type unit has a much lower copy number and is apparently not expressed. In Dg, the isolated G-type sequence carries a 420-bp insertion in the promoter region, which is probably responsible for inactivation. Sequence comparisons of different cluster units show that differentiation of the two types precedes the lineage split of these species. Substitution rates of the deduced proteins reveal two distinct subregions: high variability at the N terminus and strong sequence conservation in the rest of the protein. The variable region contains motifs characteristic of DNA-binding proteins. Adaptive diversification of the cluster units towards specific binding properties might be a plausible explanation for variability in the N-termini. Both unit types have lost the weak promoter region characteristic of P transposons. In the A-type unit, a new promoter has been formed which is apparently composed of parts of insertion sequences derived from two different mobile elements.(ABSTRACT TRUNCATED AT 250 WORDS)

Gly114Asp mutation of rhodopsin in autosomal dominant retinitis pigmentosa.

Two autosomal dominant retinitis pigmentosa families of different origin were screened for rhodopsin mutations using the method of single strand conformation polymorphism and direct sequencing. We found a CGG-CAG substitution in codon 114 of rhodopsin in both families. This change predicted the replacement of a glycine by an aspartic acid and suggested that this change is the cause of the disease in these families.

A heterochromatic P sequence in the D. subobscura genome.

The study of a heterochromatic P sequence of D. subobscura reveals that it is a degraded element, located at the centromeric region of the A chromosome (X chromosome in this species), and that it is strongly diverged from the euchromatic P sequences previously described in this species. This heterochromatic sequence is composed of some P element fragments embedded in undefined beta-heterochromatic sequences. These mosaic P sequences do not show any transcriptional activity and seem to be ancient parasites of the D. subobscura genome. Phylogenetic analyses indicate that both the euchromatic and heterochromatic P sequences of D. subobscura could come from an ancestral element which was present before the divergence of the subobscura species cluster.