Biomedical subjects

B Z Shilo

Publications and source records attributed to B Z Shilo.

At least 55 records · Page 3Linked to original sources

PubMed1992

Roles of receptor tyrosine kinases in Drosophila development.

Communication between cells is a fundamental component of development and morphogenesis. Identification of the molecules mediating cell-cell communication is crucial for elucidation of the molecular basis of these processes. Receptor tyrosine kinases (RTKs) appear to play a central role in this context by transmitting into cells information dictating their fate. The functions of RTKs in Drosophila are extremely diverse, and include maternal determination of embryonic polarity (torso and torpedo), determination of neuroblast identity (faint little ball), and guidance of tracheal cell migration in the embryo (breathless). During compound eye development, RTKs affect the number of photoreceptor clusters (Ellipse) and the determination of photoreceptor R7 identity (sevenless). The phenotypes of mutations in RTK loci serve as a starting point for understanding processes dictating cell identity at the level of the whole organism. Recently, they have also begun to provide a basis for selection of second-site suppressor mutations, encoding additional elements in their signal transduction pathway. Common themes between the functions, regulation, and signal transduction pathways of Drosophila RTKs are drawn.

Animals↗

PubMed1992

breathless, a Drosophila FGF receptor homolog, is essential for migration of tracheal and specific midline glial cells.

A Drosophila homolog of the fibroblast growth factor (FGF) receptor was isolated and structurally characterized. After EMS mutagenesis or imprecise excisions of marked P elements inserted upstream to the gene, a phenotypic series of mutations in the locus was isolated. The mutants exhibit defects in the two embryonic tissues in which the receptor is expressed: the tracheal system and the midline. The tracheal cells fail to migrate in severe mutants and remain within the tracheal pits. Hypomorphic alleles exhibit partial migration of all tracheal branches; thus, the locus was termed breathless (btl). In the midline of the mutant embryos, the posterior pair of midline glial cells begins to migrate anteriorly, but fails to reach the posterior commissure. Abnormalities in cell migration appear to be a common denominator for the btl defects in these two disparate tissues. In hypomorphic mutants the cells exhibit partial migration but follow the normal tracts, suggesting that the presence of this receptor is essential for the ability of the migrating cells to recognize external guiding cues.

Amino Acid Sequence↗

PubMed1992

Dissection of the faint little ball (flb) phenotype: determination of the development of the Drosophila central nervous system by early interactions in the ectoderm.

The complex embryonic phenotype of mutations in the faint little ball (flb) locus, encoding the Drosophila EGF receptor homolog (DER), was dissected by temperature shifts of a temperature-sensitive allele. We show that the phenotype can be resolved into at least five components, which are temporally and spatially distinct. Most notably, the central nervous system (CNS) phenotype is determined at two separate phases. A severe collapse results from early defects in the DER-expressing ectodermal cells from which neuroblasts and midline glial cells deaminate. We thus suggest that DER activity is crucial for interactions that occur in the ectoderm at an early stage, and determine the fate of neuronal and glial cell lineages. This finding explains how a severe CNS phenotype is generated in flb embryos, in spite of the absence of expression of the protein in neuronal cells. In a second phase, during germ band retraction, the flb function is required specifically in the three pairs of midline glial cells (MG). In the absence of a functional DER protein, these cells die or fail to differentiate correctly, resulting in a fused commissure phenotype.

Animals↗

PubMed1991

Interallelic complementation among DER/flb alleles: implications for the mechanism of signal transduction by receptor-tyrosine kinases.

The large number of available embryonic lethal alleles in the Drosophila EGF receptor homolog (DER)/faint little ball locus allowed us to test the possibility of positive or negative interactions among different DER alleles. These interactions were monitored by examining the embryonic cuticular phenotypes of different heteroallelic combinations. Several positive interactions were identified, while negative interactions were restricted to a single allele. This is the first example of positive interactions within the same cell type among alleles of a receptor tyrosine kinase gene. The basis for these interactions is likely to arise from the mechanism of signal transduction by receptor tyrosine kinases, which involves receptor aggregation. A combination of two different DER mutant proteins defective in temporally distinct stages of the signal transduction process, may thus form a functional heterodimer. The mutation sites in four alleles showing positive interactions were localized. They identify regions within the protein which are likely to be important for these temporally distinct signal transduction processes.

Alleles↗

PubMed1991

The Drosophila FGF-R homolog is expressed in the embryonic tracheal system and appears to be required for directed tracheal cell extension.

The Drosophila homolog of the vertebrate fibroblast growth factor receptor (FGF-R) was isolated by low-stringency hybridization. In contrast to the diversity of this subclass of receptor tyrosine kinases in vertebrates, the Drosophila genome appears to encode only a single homolog. Nucleotide sequence analysis demonstrates that the Drosophila FGF-R homolog (DFGF-R) protein has a conserved sequence, size, and organization. The extracellular region encodes three immunoglobulin-like domains, and the cytoplasmic kinase domain exhibits a high degree of similarity to the vertebrate FGF-Rs with the typical split kinase and comparably sized juxtamembrane and carboxy-terminal regions. The DFGF-R was mapped to position 70C on the third chromosome, and two overlapping chromosomal deficiencies that remove the gene were identified. Developmental Northern blots show that the gene has a single transcript of 4.3 kb and is expressed at all stages of development. Localization of the transcript and protein in embryos has shown that the gene is predominantly expressed in a restricted set of tissues: the developing tracheal system and the delaminating midline glial and neural cells. In embryos homozygous for a deletion of several genes including the DFGF-R locus, the initial formation of the tracheal pits is not affected. However, the extension of tracheal cell processes leading to the formation of the elaborate tree structure is blocked. The DFGF-R protein may thus participate in receiving spatial cues that guide tracheal cell outgrowth.

Amino Acid Sequence↗

PubMed1990

Enhancement of tyrosine kinase activity of the Drosophila epidermal growth factor receptor homolog by alterations of the transmembrane domain.

The Drosophila epidermal growth factor receptor homolog (DER) displays sequence similarity to both the epidermal growth factor (EGF) receptor and the neu vertebrate proteins. We have examined the possibility of deregulating the tyrosine kinase activity of DER by introducing structural changes which mimic the oncogenic alterations in the vertebrate counterparts. Substitution of valine by glutamic acid in the transmembrane domain, in a position analogous to the oncogenic mutation in the rat neu gene, elevated the in vivo kinase activity of DER in Drosophila Schneider cells sevenfold. A chimera containing the oncogenic neu extracellular and transmembrane domains and the DER kinase region, also showed a threefold elevated activity relative to a similar chimera with normal neu sequences. Double truncation of DER in the extracellular and cytoplasmic domains, mimicking the deletions in the v-erbB oncogene, did not however result in stimulation of in vivo kinase activity. The chimeric constructs were also expressed in monkey COS cells, and similar results were obtained. The ability to enhance the DER kinase activity by a specific structural modification of the transmembrane domain demonstrates the universality of this activation mechanism and strengthens the notion that this domain is intimately involved in signal transduction. These results also support the inclusion of DER within the tyrosine-kinase receptor family.

Amino Acid Sequence↗

PubMed1990

Localization of the DER/flb protein in embryos: implications on the faint little ball lethal phenotype.

Antibodies were raised against the Drosophila EGF receptor homolog (DER) and used for immunohistochemical analyses of Drosophila embryos. We found that DER is localized in a wide array of embryonic tissues, displaying a dynamic pattern of expression. DER appears to be expressed in all cells at the cellular blastoderm and gastrula stages. In extended-germ-band embryos, it is found predominantly in the mesoderm and the head. Finally, in retracted-germ-band embryos, DER immunoreactivity is most pronounced at sites of somatic muscle attachments and along the ventral midline of the CNS. We have thus observed that DER is expressed in the diverse tissues which are affected in the DER faint little ball (flb) embryonic lethal phenotype. The different pattern and extent of expression in each tissue suggests that the disparate aspects of the flb phenotype may result from different mechanisms of DER function. To understand the basis for the CNS phenotype of DER/flb mutants, we have closely followed the collapse of the CNS in mutant embryos. Our observations on the evolution of the final CNS phenotype, in combination with the temporo-spatial pattern of appearance of DER in the ventral neuroepithelium, suggest that this receptor participates in the second phase of neuron-glia interactions, namely in stabilization of the ladder-like CNS scaffolding formed by outgrowth of pioneer axonal processes along the glial pre-pattern.

Animals↗

PubMed1990

Biochemical properties of the Drosophila EGF receptor homolog (DER) protein.

Antibodies recognizing the Drosophila EGF receptor homolog (DER) were used to define the biochemical properties of the protein. We show that DER is a glycosylated protein, with different processing patterns in Drosophila and mammalian cells, and confirm its localization in the plasma membrane with the predicted orientation. The receptor produced in transfected cells displays only a basal activity of in vivo tyrosine kinase autophosphorylation, providing a sensitive assay for ligands which may trigger DER. EGF and TGF alpha, which trigger the mammalian EGF receptor, did not increase the kinase activity of DER.

Animals↗

PubMed1989

The Drosophila EGF receptor homolog (DER) gene is allelic to faint little ball, a locus essential for embryonic development.

Recessive lethal mutations in the genetic locus of the Drosophila EGF receptor homolog (DER) were isolated. Identification of mutations in the gene is based on assays of DER protein autophosphorylation activity. Most DER alleles show little or no in vivo autophosphorylation. The ability to monitor these activities in vivo and in vitro offers a preliminary insight into the functional defects in the different mutant proteins. The identification of the DER locus was also confirmed by partial rescue of the mutant phenotype with a DER P-element construct. Homozygous DER mutants display a complex embryonic phenotype. Most notably, the anterior structures deteriorate, ventral denticle bands are missing, the germ band does not retract, and the central nervous system shows a collapse of commissure and midline pattern. Mutations in DER were shown to be allelic to the previously described locus faint little ball.

Alleles↗

PubMed1986

Alternative 5' exons and tissue-specific expression of the Drosophila EGF receptor homolog transcripts.

cDNA clones of the Drosophila epidermal growth factor receptor homolog (DER) gene were isolated and sequenced. The deduced amino acid sequence shows a similar degree of homology to the human epidermal growth factor receptor and to the rat and human neu proteins; the most striking difference is the addition of a third cysteine-rich extracellular domain in DER. The structure of the cDNA indicates the use of alternative 5' exons. Thus, the gene encodes three putative proteins differing at their N termini. The distribution of DER transcripts was analyzed by in situ hybridization. Transcripts are uniformly distributed in embryos, larval transcripts are primarily localized to proliferating tissues of the imaginal discs and brain cortex, and adult transcripts are detected mainly in the brain and ganglia. All three splicing alternatives show similar tissue distribution during development.

Age Factors↗

PubMed1986

Tissue localization of Drosophila melanogaster ras transcripts during development.

Three ras homologs have been identified in Drosophila melanogaster. Here we describe the tissue distribution of their transcripts as analyzed by in situ hybridization. The RNAs of the three genes show a similar distribution at every developmental stage examined. In embryos, the transcripts are uniformly distributed. In larvae, ras transcripts are restricted to dividing cells (e.g., imaginal disks, gonads, and brain). At the adult stage, several tissues contain ras transcripts. The strongest hybridization signals are localized to the adult ovaries and to the cortex of the brain and ganglia, which at this stage are comprised of differentiated, nondividing cells. The tissue distribution of ras transcripts in D. melanogaster suggests that the ras proteins have multiple roles during development which may be related to both the proliferative and differentiated states of the tissues.

Animals↗

PubMed1986

The Drosophila epidermal growth factor receptor homolog: structure, evolution, and possible functions.

A unique gene termed DER (Drosophila epidermal growth factor [EGF] receptor homolog) was isolated from Drosophila melanogaster and mapped to position 57F on the right arm of the second chromosome. The deduced amino acid sequence showed that the DER protein is 1409 amino acids long. In homology it is similar to the human EGF receptor and to rat and human neu proteins. The most striking difference between the Drosophila and human homologs is DER's additional 166 amino acids. The extra sequence is rich in cysteine residues and is another duplication of one of the two cysteine-rich regions, which are a hallmark of the EGF receptor and related proteins. Analysis of several cDNA clones of DER revealed variability at the 5' end of the coding region, demonstrating the presence of at least three splicing alternatives. All three transcripts have a similar tissue distribution during development: they are uniformly distributed in embryos, localized primarily in proliferating tissues in larvae, and found mainly in the brain cortex and the thoracic and abdominal ganglia in adults. The DER protein thus seems to have multiple roles during development, and it may represent a "universal" transducer of signals into the cell.

Animals↗

PubMed1985

Characterization of functional domains of p21 ras by use of chimeric genes.

Comparison of the predicted amino acid sequences of different members of the ras family in vertebrates has shown that the N-terminal 120 residues are highly conserved while the C terminus is variable. To test the possible role of the variable residues in cell transformation, chimeras were constructed containing the N-terminal 111 amino acids of the human Ha-ras EJ oncogene and the C terminus of two Drosophila ras genes. We show that one of these constructs which has only 20 conserved residues between positions 121 and 189, can transform rat-1 cells, and the transformed cells are capable of inducing lethal tumors in rats. The second construct containing the C terminus of another Drosophila ras gene exhibits a transforming capacity as well, but only after linkage to a viral transcriptional promoter. These results show that the majority of residues within the C terminus can be replaced without abolishing the transforming potential of p21 ras.

Amino Acid Sequence↗

PubMed1985

Developmental changes in expression of the Drosophila melanogaster epidermal growth factor receptor gene.

A Drosophila gene homologous to the human EGF receptor gene has recently been isolated and sequenced. Two transcripts, 7.6- and 7.1-kb long, encoded by this gene were identified in Drosophila melanogaster. The transcripts are present at low abundance in the maternal RNA stored in unfertilized eggs and in 2-hr-old embryos. The abundance of both transcripts increases sharply between 2 and 5 hr after egg deposition, it remains high throughout embryogenesis, and decreases again in the larval and pupal stages. In adult flies the two transcripts are expressed differentially. The 7.1-kb transcript is present during adulthood at the same level detected previously in pupal stage, but the abundance of the 7.6-kb transcript decreases substantially and it remains low during adulthood.

Animals↗

PubMed1985

The Drosophila EGF receptor gene homolog: conservation of both hormone binding and kinase domains.

Chicken v-erB probe was used to isolate a unique clone of Drosophila melanogaster DNA. It maps by in situ hybridization to position 57F on chromosome 2. A complete nucleotide sequence of the coding region has been obtained. The putative Drosophila EGF receptor protein is similar in overall organization to the human homolog. It shows three distinct domains: an extracellular putative EGF binding domain, a hydrophobic transmembrane region, and a cytoplasmic kinase domain. The overall amino acid homology is 41% in the extracellular domain and 55% in the kinase domain. Two cysteine-rich regions, a hallmark of the human ligand-binding domain, have also been conserved. Fusion of the coding sequences of the kinase and extracellular domains generating the receptor gene must have occurred over 800 million years ago.

Amino Acid Sequence↗

PubMed1984

The Drosophila ras oncogenes: structure and nucleotide sequence.

Three Drosophila genes homologous to the Ha-ras probe were isolated and mapped to positions 85D, 64B, and 62B on chromosome 3. Two of these genes (termed Dras 1 and Dras 2) were sequenced. In the case of Dras 1, which contains multiple introns, a cDNA clone was isolated and sequenced. In the case of Dras2, the nucleotide sequence fo the genomic clone was determined. Each gene codes for a protein with a predicted molecular weight of 21.6 kd. Alignment of the amino acid sequence of Dras 1 with the vertebrate Ha-ras protein shows that at the amino terminus and central portion (residues 1-121 and 137-164) the two proteins are remarkably similar, and have an overall homology of 75%. The Dras 2 gene lacks significant homology to the vertebrate counterpart at the extreme amino terminus and is homologous only between positions 28-120 and 139-161 (overall homology of 50%). This result suggests that the N terminus of p21 forms a distinct regulatory or functional domain. At the carboxy terminus, the major region of variability among the vertebrate ras proteins, the two Drosophila sequences also display considerable variability. However, both appear to be more similar to exon 4B of the Ki-ras gene.

Amino Acid Sequence↗

PubMed1984

Expression of the src and abl cellular oncogenes during development of Drosophila melanogaster.

Transcription of the Drosophila homologs of the abl and src cellular oncogenes, termed Dash and Dsrc , respectively, was studied. Despite the fact that Dash and Dsrc share sequence homology, they have distinct patterns of transcription during development. The Dash transcript, 6.2 kilobases long, is found in maternal RNA stored in unfertilized eggs and in embryos up to 4 h after egg laying. In contrast, Dsrc transcripts are 3.2, 4.8, and 5.2 kilobases long. They are detected in eggs and embryos and to a lesser extent in larvae and adult flies. The relative amount of each of the three Dsrc transcripts changes during the different stages of development.

Animals↗

PubMed1983

Nucleotide sequences of the Drosophila src and abl homologs: conservation and variability in the src family oncogenes.

Two cloned Drosophila sequences, Dash and Dsrc, were previously isolated by hybridization to the viral oncogenes v-abl and v-src. We report that the Drosophila DNA sequences are more than 50% homologous to 700 base pairs of the vertebrate oncogenes that are essential for kinase activity of the v-abl and the v-src gene products. Alignment of the predicted amino acid sequences in the homologous regions shows amino acid sequence identities of 74% between Dash and the 5' portion of v-abl and of 54% between Dsrc and 3' portion of v-src. Comparison of the Drosophila and vertebrate sequences identifies amino acids that may be essential for the distinct functions of the c-src and the c-abl gene products and places the gene duplication event that has generated the two genes prior to the Chordate-Arthropod divergence.

Amino Acid Sequence↗