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PubMed1994

Inhibition of the DNA-binding activity of Drosophila suppressor of hairless and of its human homolog, KBF2/RBP-J kappa, by direct protein-protein interaction with Drosophila hairless.

We have purified the sequence-specific DNA-binding protein KBF2 and cloned the corresponding cDNA, which is derived from the previously described RBP-J kappa gene, the human homolog of the Drosophila Suppressor of Hairless [Su(H)] gene. Deletion studies of the RBP-J kappa and Su(H) proteins allowed us to define a DNA-binding domain conserved during evolution. Because Su(H) mutant alleles exhibit dose-sensitive interactions with Hairless (H) loss-of-function mutations, we have investigated whether the RBP-J kappa or Su(H) proteins directly interact with the H protein in vitro. We show here that H can inhibit the DNA binding of both Su(H) and RBP-J kappa through direct protein-protein interactions. Consistent with this in vitro inhibitory effect, transcriptional activation driven by Su(H) in transfected Drosophila S2 cells is inhibited by H. These results support a model in which H acts, at least in part, as a negative regulator of Su(H) activity. This model offers a molecular view to the antagonistic activities encoded by the H and Su(H) genes for the control of sensory organ cell fates in Drosophila. We further propose that a similar mechanism might occur in mammals.

Amino Acid Sequence↗
PubMed2004

Anomalies in the expression profile of interspecific hybrids of Drosophila melanogaster and Drosophila simulans.

When females of Drosophila melanogaster and males of Drosophila simulans are mated, the male progeny are inviable, whereas the female progeny display manifold malformations and are sterile. These abnormalities result from genetic incompatibilities accumulated since the time the lineages of the species diverged, and may have their origin in aberrant gene transcription. Because compensatory changes within species may obscure differences at the regulatory level in conventional comparisons of the expression profile between species, we have compared the gene-expression profile of hybrid females with those of females of the parental species in order to identify regulatory incompatibilities. In the hybrid females, we find abnormal levels of messenger RNA for a large fraction of the Drosophila transcriptome. These include a gross underexpression of genes preferentially expressed in females, accompanying gonadal atrophy. The hybrid females also show significant overexpression of male-biased genes, which we attribute to incompatibilities in the regulatory mechanisms that normally act to control the expression of these genes in females. The net result of the multiple incompatibilities is that the gene-expression profiles of the parental females are more similar to each other than either is to that of the hybrid.

Animals↗
PubMed1991

Molecular transfer of a species-specific behavior from Drosophila simulans to Drosophila melanogaster.

Drosophila males modulate the interpulse intervals produced during their courtship songs. These song cycles, which are altered by mutations in the clock gene period, exhibit a species-specific variation that facilitates mating. We have used chimeric period gene constructs from Drosophila melanogaster and Drosophila simulans in germline transformation experiments to map the genetic control of their song rhythm difference to a small segment of the amino acid encoding information within this gene.

Amino Acid Sequence↗
PubMed2000

Cubitus interruptus requires Drosophila CREB-binding protein to activate wingless expression in the Drosophila embryo.

CREB-binding protein (CBP) serves as a transcriptional coactivator in multiple signal transduction pathways. The Drosophila homologue of CBP, dCBP, interacts with the transcription factors Cubitus interruptus (CI), MAD, and Dorsal (DL) and functions as a coactivator in several signaling pathways during Drosophila development, including the hedgehog (hh), decapentaplegic (dpp), and Toll pathways. Although dCBP is required for the expression of the hh target genes, wingless (wg) and patched (ptc) in vivo, and potentiates ci-mediated transcriptional activation in vitro, it is not known that ci absolutely requires dCBP for its activity. We used a yeast genetic screen to identify several ci point mutations that disrupt CI-dCBP interactions. These mutant proteins are unable to transactivate a reporter gene regulated by ci binding sites and have a lower dCBP-stimulated activity than wild-type CI. When expressed exogenously in embryos, the CI point mutants cannot activate endogenous wg expression. Furthermore, a CI mutant protein that lacks the entire dCBP interaction domain functions as a negative competitor for wild-type CI activity, and the expression of dCBP antisense RNAs can suppress CI transactivation in Kc cells. Taken together, our data suggest that dCBP function is necessary for ci-mediated transactivation of wg during Drosophila embryogenesis.

Animals↗
PubMed1995

Structure and expression of histone H3.3 genes in Drosophila melanogaster and Drosophila hydei.

We demonstrate that in Drosophila melanogaster the histone H3.3 replacement variant is encoded by two genes, H3.3A and H3.3B. We have isolated cDNA clones for H3.3A and cDNA and genomic clones for H3.3B. The genes encode exactly the same protein but are widely divergent in their untranslated regions (UTR). Both genes are expressed in embryos and adults; they are expressed in the gonads as well as in somatic tissues of the flies. However, only one of them, H3.3A, shows strong testes expression. The 3' UTR of the H3.3A gene is relatively short (approximately 250 nucleotides (nt)). H3.3B transcripts can be processed at several polyadenylation sites, the longest with a 3' UTR of more than 1500 nt. The 3' processing sites, preferentially used in the gonads and somatic tissues, are different. We have also isolated the Drosophila hydei homologues of the two H3.3 genes. They are quite similar to the D. melanogaster genes in their expression patterns. However, in contrast to their vertebrate counterparts, which are highly conserved in their noncoding regions, the Drosophila genes display only limited sequence similarity in these regions.

Animals↗
PubMed1998

The Drosophila EGF receptor controls the formation and specification of neuroblasts along the dorsal-ventral axis of the Drosophila embryo.

The segmented portion of the Drosophila embryonic central nervous system develops from a bilaterally symmetrical, segmentally reiterated array of 30 unique neural stem cells, called neuroblasts. The first 15 neuroblasts form about 30-60 minutes after gastrulation in two sequential waves of neuroblast segregation and are arranged in three dorsoventral columns and four anteroposterior rows per hemisegment. Each neuroblast acquires a unique identity, based on gene expression and the unique and nearly invariant cell lineage it produces. Recent experiments indicate that the segmentation genes specify neuroblast identity along the AP axis. However, little is known as to the control of neuroblast identity along the DV axis. Here, I show that the Drosophila EGF receptor (encoded by the DER gene) promotes the formation, patterning and individual fate specification of early forming neuroblasts along the DV axis. Specifically, I use molecular markers that identify particular neuroectodermal domains, all neuroblasts or individual neuroblasts, to show that in DER mutant embryos (1) intermediate column neuroblasts do not form, (2) medial column neuroblasts often acquire identities inappropriate for their position, while (3) lateral neuroblasts develop normally. Furthermore, I show that active DER signaling occurs in the regions from which the medial and intermediate neuroblasts will later delaminate. In addition, I demonstrate that the concomitant loss of rhomboid and vein yield CNS phenotypes indistinguishable from DER mutant embryos, even though loss of either gene alone yields minor CNS phenotypes. These results demonstrate that DER plays a critical role during neuroblast formation, patterning and specification along the DV axis within the developing Drosophila embryonic CNS.

Animals↗
PubMed2003

A mouse homologue of Drosophila pins can asymmetrically localize and substitute for pins function in Drosophila neuroblasts.

Asymmetric cell division is a fundamental mechanism used to generate cellular diversity in invertebrates and vertebrates. In Drosophila, asymmetric division of neuroblasts is achieved by the asymmetric segregation of cell fate determinants Prospero and Numb into the basal daughter cell. Asymmetric segregation of cell fate determinants requires an apically localized protein complex that includes Inscuteable, Pins, Bazooka, DmPar-6, DaPKC and Galphai. Pins acts to stabilize the apical complex during neuroblast divisions. Pins interacts and colocalizes with Inscuteable, as well as maintaining its apical localization. We have isolated a mouse homologue of pins (Pins) and characterized its expression profile. Mouse PINS shares high similarity in sequence and structure with Pins and other Pins-like proteins from mammals. Pins is expressed in many mouse tissues but its expression is enriched in the ventricular zone of the developing central nervous systems. PINS localizes asymmetrically to the apical cortex of mitotic neuroblasts when ectopically expressed in Drosophila embryos. Like Pins, its N-terminal tetratricopeptide repeats can directly interact with the asymmetric localization domain of Insc, and its C-terminal GoLoco-containing region can direct localization to the neuroblast cortex. We further show that Pins can fulfill all aspects of pins function in Drosophila neuroblast asymmetric cell divisions. Our results suggest a conservation of function between the fly and mammalian Pins homologues.

Amino Acid Sequence↗
PubMed1992

Overexpressed Drosophila src 64B is phosphorylated at its carboxy-terminal tyrosine, but is not catalytically repressed, in cultured Drosophila cells.

Little is known about the regulation of non-receptor tyrosine kinases in invertebrates. We have studied the relationship between the phosphorylation state of the Drosophila src 64B (Dsrc) gene product, p62D, and its tyrosine kinase activity in Drosophila Schneider 2 cells, using wild-type and mutated Dsrc constructs that were overexpressed by transient transfection. Phosphopeptide mapping showed that the putative regulatory C-terminal tyrosine (Tyr-547) of p62D was phosphorylated in vivo. In contrast to vertebrate src family kinases overexpressed in fibroblasts, wild-type p62D overexpressed in Schneider 2 cells was phosphorylated at additional tyrosines outside of the C-terminus. These tyrosines corresponded to the major in vitro autophosphorylation sites. Overexpression of wild-type p62D or several catalytically active p62D mutants significantly increased the phosphorylation of numerous Schneider cell proteins on tyrosine, while expression of catalytically inactive mutants of p62D had no such effect. Thus, in contrast to the repression of src family kinase activity in fibroblasts, p62D is catalytically active when overexpressed in Drosophila cells, perhaps because of substoichiometric C-terminal tyrosine phosphorylation. These results raise the possibility that fly development will be sensitive to ectopic expression of p62D.

Amino Acid Sequence↗
PubMed1993

Characterization and evolution of the Adh genomic region in Drosophila guanche and Drosophila madeirensis.

We have sequenced the genomic region of the Adh (alcohol dehydrogenase) gene of Drosophila guanche and Drosophila madeirensis. Two genes, Adh and Adh-dup, whose main features are shared by other Drosophila species, are contained in this region. Interspecies comparisons of this genomic region of these two species with the region in D. subobscura have been made. Our data are in agreement with the phylogenetic relationship of the three species and particularly with the very close proximity of D. madeirensis and D. subobscura. The correct alignment of the noncoding as well as the coding sequences allows us to infer how this region has evolved. The degree of divergence of D. madeirensis and D. subobscura Adh sequences is low and replacement substitutions are as frequent as silent substitutions. In addition, heterogeneous divergence rates for coding and noncoding sequences have been observed. Our data also reveal that a recent evolutionary event is responsible for the branching of the three species.

Alcohol Dehydrogenase↗
PubMed1990

Discrepancy in divergence of the mitochondrial and nuclear genomes of Drosophila teissieri and Drosophila yakuba.

Restriction sites were compared in the mitochondrial DNA (mtDNA) molecules from representatives of two closely related species of fruit flies: nine strains of Drosophila teissieri and eight strains of Drosophila yakuba. Nucleotide diversities among D. teissieri strains and among D. yakuba strains were 0.07% and 0.03%, respectively, and the nucleotide distance between the species was 0.22%. Also determined was the nucleotide sequence of a 2305-nucleotide pair (ntp) segment of the mtDNA molecule of D. teissieri that contains the noncoding adenine + thymine (A + T)-rich region (1091 ntp) as well as the genes for the mitochondrial small-subunit rRNA, tRNA(f-met), tRNA(gln), and tRNA(ile), and portions of the ND2 and tRNA(Val) genes. This sequence differs from the corresponding segment of the D. yakuba mtDNA by base substitutions at 0.1% and 0.8% of the positions in the coding and noncoding regions, respectively. The higher divergence due to base substitutions in the A + T-rich region is accompanied by a greater number of insertions/deletions than in the coding regions. From alignment of the D. teissieri A + T-rich sequence with those of D. yakuba and Drosophila virilis, it appears that the 40% of this sequence that lies adjacent to the tRNA(ile) gene has been highly conserved. Divergence between the entire D. teissieri and D. yakuba mtDNA molecules, estimated from the sequences, was 0.3%; this value is close to the value (0.22%) obtained from the restriction analysis, but 10 times lower than the value estimated from published DNA hybridization results.(ABSTRACT TRUNCATED AT 250 WORDS)

Africa↗
PubMed1990

Evolution of the autosomal chorion cluster in Drosophila. IV. The Hawaiian Drosophila: rapid protein evolution and constancy in the rate of DNA divergence.

Autosomal chorion genes s18, s15, and s19 are shown to diverge at extremely rapid rates in closely related taxa of Hawaiian Drosophila. Their nucleotide divergence rates are at least as fast as those of intergenic regions that are known to evolve more extensively between distantly related species. Their amino acid divergence rates are the fastest known to date. There are two nucleotide replacement substitutions for every synonymous one. The molecular basis for observed length and substitution mutations is analyzed. Length mutations are strongly associated with direct repeats in general, and with tandem repeats in particular, whereas the rate for an average transition is twice that for an average transversion. The DNA sequence of the cluster was used to construct a phylogenetic tree for five taxa of the Hawaiian picture-winged species group of Drosophila. Assignment of observed base substitutions occurring in various branches of the tree reveals an excess of would-be homoplasies in a centrally localized 1.8-kb segment containing the s15 gene. This observation may be a reflection of ancestral excess polymorphisms in the segment. The chorion cluster appears to evolve at a constant rate regardless of whether the central 1.8-kb segment is included or not in the analysis. Assuming that the time of divergence of Drosophila grimshawi and the planitibia subgroup coincides with the emergence of the island of Kauai, the overall rate of base substitution in the cluster is estimated to be 0.8% million years, whereas synonymous sites are substituted at a rate of 1.2% million years.

Amino Acid Sequence↗
PubMed2002

The actin loci in the genus Drosophila: establishment of chromosomal homologies among five nearctic species of the Drosophila obscura group by in situ hybridization.

The actin genes of five nearctic species of the Drosophila obscura group were mapped by in situ hybridization, using the 5C actin gene of D. melanogaster as a probe. In all species but D. azteca eight actin loci were observed variously dispersed over all five (A- E) chromosomal elements. In D. azteca ten actin hybridization sites were found; four of which most probably originated by duplications or by transposition events. Although the five nearctic species differ from all other Drosophila species of the D. obscura group so far studied in the number of loci as well as in the chromosomal distribution and location of the actin loci, the uniformity of the main pattern with six actin loci throughout the genus Drosophila reinforces the hypothesis that the chromosomal elements have maintained their essential identities during the course of evolution. Our findings are in accordance with the conclusion that the nearctic D. obscura species have differentiated from a common ancestor of the palearctic species and that they belong to two distinct subgroups, the pseudoobscura and the affinis subgroups.

Actins↗
PubMed2005

Phylogeny and molecular evolution of the Drosophila hydei subgroup (Drosophila repleta group) inferred from the Xanthine dehydrogenase gene.

The hydei subgroup (Drosophila repleta group) consists of seven species divided into two complexes: bifurca and hydei, whose phylogenetic relationships are not well understood. To evaluate the molecular phylogeny of this subgroup, we analyzed 2085 bp of coding sequence of the Xanthine dehydrogenase gene in six available species of the hydei subgroup, with Drosophila buzzatii and Drosophila mulleri as an outgroup. For phylogenetic reconstruction we adopted a maximum-likelihood framework, based on the adjustment of descriptive models of nucleotide substitution to real data. We employed distance-based and weighted parsimony methods to construct candidate phylogenies. In all cases, we obtained only one completely resolved tree with strong statistical support for each node, that shows a phylogeny that is partially discordant with the proposed systematics of the subgroup. This tree suggests that the two species complexes are paraphyletic, as opposed to classic phylogenies using morphologic and cytologic traits. This discordance is discussed in relation to its implication for the evolutionary history of the hydei subgroup.

Animals↗
PubMed2004

Expression of expanded polyglutamine protein induces behavioral changes in Drosophila (polyglutamine-induced changes in Drosophila).

Spinocerebellar ataxia type-3 or Machado-Joseph disease (SCA3/MJD) is an autosomal dominant neurodegenerative disease caused by triplet nucleotide expansion. The expansion of the polyglutamine tract near the C terminus of the MJD1 gene product, ataxin-3, above a threshold of 40 glutamine repeats causes neuronal loss and degeneration. The expanded ataxin-3 forms aggregates, and nuclear inclusions, within neurons, possibly due to the misfolding of mutant proteins. Here we report upon the behavioral test changes related to truncated and expanded forms of MJD protein (MJDtr) in Drosophila, and show that expanded MJDtr, when expressed in the nervous system, causes characteristic locomotor dysfunction and anosmia. This phenomenon has not been previously reported in humans or in transgenic Drosophila models. In addition, the in vivo expression of the antiapoptotic gene bcl-2 showed no evidence of ameliorating the deleterious effect of MJDtr-Q78s, either in the eye or in the nervous system. The study shows that such Drosophila transgenic models express olfactory dysfunction and ataxic behavior as observed in human patients.

Animals↗
PubMed1989

Genetics of sexual isolation between two sibling species, Drosophila simulans and Drosophila mauritiana.

Drosophila simulans and Drosophila mauritiana are sibling species that show substantial sexual isolation in one of their two reciprocal hybridizations. Genetic analysis reveals that in females this isolation is caused by at least one recessive gene on each autosome, while the X chromosome has little or no effect. Our results, combined with those of previous studies, show that in Drosophila the genetics of sexual isolation differs from that of postzygotic reproductive isolation, which invariably involves large effects of the X chromosome.

Animals↗
PubMed1993

A Drosophila G-protein alpha subunit, Gf alpha, expressed in a spatially and temporally restricted pattern during Drosophila development.

Heterotrimeric guanine nucleotide binding proteins (G proteins) couple receptors for extracellular signals to intracellular second messenger-generating systems. Previous studies have shown that G-protein alpha subunits (G alpha) are expressed in a spatially and temporally restricted manner during Drosophila embryogenesis and, thus, may be responsible for mediating developmental interactions. We have used the polymerase chain reaction to search for specific G alpha subunits that function primarily during development and not in the adult fly. Using poly(A)+ RNA isolated from early pupae (day 1), we have isolated a cDNA coding for a fly G alpha subunit, designated Gf alpha. This subunit is 30-38% identical to previously described vertebrate and Drosophila G alpha subunits and appears to define an additional family of G alpha proteins. Gf alpha transcripts are expressed primarily during embryonic, larval, and early pupal stages and only at low levels in adult flies. In situ hybridization studies indicate that Gf alpha transcripts are expressed maternally and later during embryogenesis primarily in the developing midgut and transiently in the amnioserosa. The Gf alpha gene has been characterized and mapped to position 73B of the Drosophila genome.

Amino Acid Sequence↗
PubMed1993

An autosomal factor from Drosophila arizonae restores normal spermatogenesis in Drosophila mojavensis males carrying the D. arizonae Y chromosome.

Males of Drosophila mojavensis whose Y chromosome is replaced by the Y chromosome of the sibling species Drosophila arizonae are sterile. It is shown that genetic material from the fourth chromosome of D. arizonae is necessary and sufficient, in single dose, to restore fertility in these males. In introgression and mapping experiments this material segregates as a single Mendelian factor (sperm motility factor, SMF). Light and electron microscopy studies of spermatogenesis in D. mojavensis males whose Y chromosome is replaced by introgression with the Y chromosome of D. arizonae (these males are symbolized as mojYa) revealed postmeiotic abnormalities all of which are restored when the SMF of D. arizonae is co-introgressed (these males are symbolized as mojYaSMFa). The number of mature sperm per bundle in mojYaSMFa is slightly less than in pure D. mojavensis and is even smaller in males whose fertility is rescued by introgression of the entire fourth chromosome of D. arizonae. These observations establish an interspecific incompatibility between the Y chromosome and an autosomal factor (or more than one tightly linked factors) that can be useful for the study of the evolution of male hybrid sterility in Drosophila and the genetic control of spermatogenesis.

Animals↗
PubMed2006

Localization and characterization of X chromosome inversion breakpoints separating Drosophila mojavensis and Drosophila arizonae.

Ectopic exchange between transposable elements or other repetitive sequences along a chromosome can produce chromosomal inversions. As a result, genome sequence studies typically find sequence similarity between corresponding inversion breakpoint regions. Here, we identify and investigate the breakpoint regions of the X chromosome inversion distinguishing Drosophila mojavensis and Drosophila arizonae. We localize one inversion breakpoint to 13.7 kb and localize the other to a 1-Mb interval. Using this localization and assuming microsynteny between Drosophila melanogaster and D. arizonae, we pinpoint likely positions of the inversion breakpoints to windows of less than 3000 bp. These breakpoints define the size of the inversion to approximately 11 Mb. However, in contrast to many other studies, we fail to find significant sequence similarity between the 2 breakpoint regions. The localization of these inversion breakpoints will facilitate future genetic and molecular evolutionary studies in this species group, an emerging model system for ecological genetics.

Animals↗