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J A Kassis

Publications and source records attributed to J A Kassis.

At least 19 recordsLinked to original sources
PubMed1999

The DNA-binding polycomb group protein pleiohomeotic mediates silencing of a Drosophila homeotic gene.

Polycomb group (PcG) proteins repress homeotic genes in cells where these genes must remain inactive during development. This repression requires cis-acting silencers, also called PcG response elements. Currently, these silencers are ill-defined sequences and it is not known how PcG proteins associate with DNA. Here, we show that the Drosophila PcG protein Pleiohomeotic binds to specific sites in a silencer of the homeotic gene Ultrabithorax. In an Ultrabithorax reporter gene, point mutations in these Pleiohomeotic binding sites abolish PcG repression in vivo. Hence, DNA-bound Pleiohomeotic protein may function in the recruitment of other non-DNA-binding PcG proteins to homeotic gene silencers.

Animals↗
PubMed1998

The Drosophila Polycomb group gene pleiohomeotic encodes a DNA binding protein with homology to the transcription factor YY1.

Genes of the Polycomb group (PcG) of Drosophila encode proteins necessary for the maintenance of transcriptional repression of homeotic genes. PcG proteins are thought to act by binding as multiprotein complexes to DNA through Polycomb group response elements (PREs); however, specific DNA binding has not been demonstrated for any of the PcG proteins. We have identified a sequence-specific DNA binding protein that interacts with a PRE from the Drosophila engrailed gene. This protein (PHO) is a homolog of the ubiquitous mammalian transcription factor Yin Yang-1 and is encoded by pleiohomeotic, a known member of the PcG. We propose that PHO acts to anchor PcG protein complexes to DNA.

Amino Acid Sequence↗
PubMed1998

Enhancer of Polycomb is a suppressor of position-effect variegation in Drosophila melanogaster.

Polycomb group (PcG) genes of Drosophila are negative regulators of homeotic gene expression required for maintenance of determination. Sequence similarity between Polycomb and Su(var)205 led to the suggestion that PcG genes and modifiers of position-effect variegation (PEV) might function analogously in the establishment of chromatin structure. If PcG proteins participate directly in the same process that leads to PEV, PcG mutations should suppress PEV. We show that mutations in E(Pc), an unusual member of the PcG, suppress PEV of four variegating rearrangements: In(l)wm4, B(SV), T(2;3)Sb(V) and In(2R)bw(VDe2). Using reversion of a Pelement insertion, deficiency mapping, and recombination mapping as criteria, homeotic effects and suppression of PEV associated with E(Pc) co-map. Asx is an enhancer of PEV, whereas nine other PcG loci do not affect PEV. These results support the conclusion that there are fewer similarities between PcG genes and modifiers of PEV than previously supposed. However, E(Pc) appears to be an important link between the two groups. We discuss why Asx might act as an enhancer of PEV.

Animals↗
PubMed1997

Rescue of Drosophila engrailed mutants with a highly divergent mosquito engrailed cDNA using a homing, enhancer-trapping transposon.

Specific fragments of Drosophila regulatory DNA can alter the insertional specificity of transposable elements causing them to 'home' to their parent gene. We used this property to insert a transposon-encoded functional coding region near a defective one and rescue a null mutation. This approach differs from homologous recombination in that the endogenous defective coding region is left in place and the genomic DNA is altered by the addition of the therapeutic transposon. We constructed a P-element-based transposon in which an engrailed cDNA from Anopheles gambiae (a mosquito) is expressed from a Drosophila engrailed minimal promoter. The promoter fragment used includes 2.6 kb of regulatory DNA that causes transposons to home to the endogenous Drosophila engrailed gene at high frequencies. We inserted this transposon onto a Drosophila chromosome that produces no functional engrailed proteins. When this transposon integrated near the engrailed promoter, adult viability was restored to engrailed mutant flies showing that the highly divergent mosquito engrailed protein can replace the Drosophila engrailed protein at all stages of development. Insertion of this transposon into the adjacent invected gene, which is transcribed in a pattern similar to engrailed, led to only embryonic rescue, suggesting an important difference in the regulation of these two genes.

Amino Acid Sequence↗
PubMed1994

Unusual properties of regulatory DNA from the Drosophila engrailed gene: three "pairing-sensitive" sites within a 1.6-kb region.

We have previously shown that a 2-kb fragment of engrailed DNA can suppress expression of a linked marker gene, white, in the P element vector CaSpeR. This suppression is dependent on the presence of two copies of engrailed DNA-containing P elements (P[en]) in proximity in the Drosophila genome (either in cis or in trans). In this study, the 2-kb fragment was dissected and found to contain three fragments of DNA which could mediate white suppression [called "pairing-sensitive sites" (PS)]. A PS site was also identified in regulatory DNA from the Drosophila escargot gene. The eye colors of six different P[en] insertions in the escargot gene suggest an interaction between P[en]-encoded and genome-encoded PS sites. I hypothesize that white gene expression from P[en] is repressed by the formation of a protein complex which is initiated at the engrailed PS sites and also requires interactions with flanking genomic DNA. Genes were sought which influence the function of PS sites. Mutations in some Polycomb and trithorax group genes were found to affect the eye color from some P[en] insertion sites. However, different mutations affected expression from different P[en] insertion sites and no one mutation was found to affect expression from all P[en] insertion sites examined. These results suggest that white expression from P[en] is not directly regulated by members of the Polycomb and trithorax group genes, but in some cases can be influenced by them. I propose that engrailed PS sites normally act to promote interactions between distantly located engrailed regulatory sites and the engrailed promoter.

Animals↗
PubMed1992

Altering the insertional specificity of a Drosophila transposable element.

Vectors derived from the Drosophila P element transposon are widely used to make transgenic Drosophila. Insertion of most P-element-derived vectors is nonrandom, but they exhibit a broad specificity of target sites. During experiments to identify cis-acting regulatory elements of the Drosophila segmentation gene engrailed, we identified a fragment of engrailed DNA that, when included within a P-element vector, strikingly alters the specificity of target sites. P-element vectors that contain this fragment of engrailed regulatory DNA insert at a high frequency near genes expressed in stripes.

Animals↗
PubMed1992

castor encodes a novel zinc finger protein required for the development of a subset of CNS neurons in Drosophila.

Using an enhancer detection screen, we have identified castor, a new gene required for embryonic CNS development in Drosophila. Embryos that lack castor expression have a diminished CNS axonal network and express engrailed aberrantly late in CNS development. castor is unique among the previously described genes involved in Drosophila neurogenesis in that its expression is restricted to a subset of delaminated CNS neuroblasts and to ventral midline glial precursor cells. The putative castor gene product contains a novel zinc-binding domain and multiple transcriptional activation domains, suggesting that it acts as a transcription factor necessary for the development of a subset of CNS neuronal precursors.

Amino Acid Sequence↗
PubMed1992

The Drosophila gene escargot encodes a zinc finger motif found in snail-related genes.

Two independent P-element enhancer detection lines were obtained that express lacZ in a pattern of longitudinal stripes early in germband elongation. In this paper, molecular and genetic characterization of a gene located near these transposons is presented. Sequence analysis of a cDNA clone from the region reveals that this gene has a high degree of similarity with the Drosophila snail gene (Boulay et al., 1987). The sequence similarity extends over 400 nucleotides, and includes a region encoding five tandem zinc finger motifs (72% nucleotide identity; 76% amino acid identity). This region is also conserved in the snail homologue from Xenopus laevis (76% nucleotide identity; 83% amino acid identity) (Sargent and Bennett, 1990). We have named the Drosophila snail-related gene escargot (esg), and the region of sequence conservation common to all three genes the 'snailbox'. A number of Drosophila genomic DNA fragments cross-hybridize to a probe from the snailbox region suggesting that snail and escargot are members of a multigene family. The expression pattern of escargot is dynamic and complex. Early in germband elongation, escargot RNA is expressed in a pattern of longitudinal stripes identical to the one observed in the two enhancer detection lines. Later in development, escargot is expressed in cells that will form the larval imaginal tissues, escargot is allelic with l(2)35Ce, an essential gene located near snail in the genome.

Alleles↗
PubMed1991

A fragment of engrailed regulatory DNA can mediate transvection of the white gene in Drosophila.

We have found a fragment of engrailed regulatory DNA that has an unusual effect on expression of a linked marker gene, white, in the P element transposon CaSpeR. Normally, flies homozygous for a given CaSpeR insertion have darker eyes than heterozygotes. However, when a particular engrailed DNA fragment is included in that transposon, homozygotes often have lighter eyes than heterozygotes. Thus, engrailed DNA appears to cause white expression to be repressed in homozygotes. The suppression of white is dependent on the proximity of the two transposons in the genome-either in cis (i.e., on the same chromosome) or in trans (i.e., on homologous chromosomes). Thus, the engrailed fragment is mediating a phenomenon similar to that mediated by the zeste gene at the white locus. However, the interactions we observe do not require, nor are influenced by, mutations of zeste. We suggest that the engrailed DNA contains one or more binding sites for a protein that facilitates interactions between transposons. The normal function of these sites may be to mediate interactions between distant cis-regulatory regions of engrailed, a large locus that extends over 70 kilobases.

Animals↗
PubMed1990

A synthetic homeodomain binding site acts as a cell type specific, promoter specific enhancer in Drosophila embryos.

A DNA sequence initially defined as a consensus binding site for the Engrailed protein is also recognized by several other homeodomain proteins and mediates the transcriptional action of these regulators in transfected tissue culture cells. Here we show that these synthetic binding sites have a more restricted and specific ability to enhance transcription when assayed in transformed embryos. Several constructs with the homeodomain binding sites linked to the fushi tarazu or engrailed promoters are silent in transformed embryos. However, when linked to the hsp70 promoter, the sites specifically activate transcription in glial cells. The effect of single base pair mutations in the binding sites suggests that activation is mediated by homeodomain protein(s). We suggest that this specific pattern of expression results from combined action at sequences within the hsp70 promoter fragment and the homeodomain binding sites. Since the tissue culture transfection assay does not show such rigid constraints on promoter activation by homeodomain proteins, it appears that subsidiary phenomena apparent in the transgenic embryos contribute importantly to the specificity of action of functionally homologous homeodomain regulators.

Animals↗
PubMed1990

Spatial and temporal control elements of the Drosophila engrailed gene.

engrailed (en) is a segmentation gene expressed in a series of stripes throughout embryonic development. Here, I show that regulatory sequences for striped expression are present within the first intron of en. The 1-kb intron is able to confer striped expression early, but not late, in development. This shows that different regulatory sequences are required for en stripes at different times in development. Furthermore, stripes generated by the intron are coincident with en stripes in a wild-type background but behave differently from endogenous engrailed stripes in some segmentation mutant backgrounds. Thus, although the intron can induce apparently normal stripes, it lacks some of the regulatory sequences present within the endogenous gene. These experiments suggest that multiple regulatory programs control an expression in stripes, and each may be able to confer "normal" spatial regulation independently.

Animals↗
PubMed1989

Evolutionary conservation of homeodomain-binding sites and other sequences upstream and within the major transcription unit of the Drosophila segmentation gene engrailed.

The engrailed (en) gene functions throughout Drosophila development and is expressed in a succession of intricate spatial patterns as development proceeds. Normal en function relies on an extremely large cis-acting regulatory region (70 kilobases). We are using evolutionary conservation to help identify en sequences important in regulating patterned expression. Sequence comparison of 2.6 kilobases upstream of the en coding region of D. melanogaster and D. virilis (estimated divergence time, 60 million years) showed that 30% of this DNA occurs in islands of near perfect sequence conservation. One of these conserved islands contains binding sites for homeodomain-containing proteins. It has been shown genetically that homeodomain-containing proteins regulate en expression. Our data suggested that this regulation may be direct. The remaining conserved islands may contain binding sites for other regulatory proteins.

Animals↗
PubMed1988

Two-tiered regulation of spatially patterned engrailed gene expression during Drosophila embryogenesis.

A regulatory cascade, initiated during the syncytial stage of embryogenesis, culminates in the striped pattern of engrailed gene expression at the cellular blastoderm stage. The early regulatory genes, for example the pair-rule genes, are expressed transiently and as their products decay a distinct regulatory programme involving segment polarity genes takes over. This late programme maintains and perhaps modifies the striped pattern of engrailed expression through interactions that may involve cell communication.

Animals↗
PubMed1986

Sequence conservation in the protein coding and intron regions of the engrailed transcription unit.

Engrailed (en) is a gene involved in proper segmentation of the Drosophila embryo. The predicted en protein contains a homeodomain and regions rich in polyalanine, polyglutamine, polyglutamate/aspartate and serine. We have taken an evolutionary approach to define which regions may be of fundamental importance by examining the D. virilis genomic sequence homologous to the D. melanogaster en primary transcription unit. Sequence homology begins at the first ATG of a long open reading frame yielding proteins of 584 and 552 amino acids for the D. virilis and D. melanogaster proteins, respectively. The predicted amino acid sequence can be divided into conserved and non-conserved domains. The C-terminal 30% of the protein (which includes the homeodomain) is completely conserved. In the N-terminal 70% of the protein, the overall conservation is 71%, but non-conservative amino acid changes occur in clusters and there are short stretches of highly conserved sequence. A region rich in glutamate and aspartate is conserved and has homology to an 18-amino acid sequence present in members of the myc family of proteins. Major differences in the size of the two proteins occur in regions of non-conserved repeated sequences. In the introns of the engrailed transcription units there are long stretches of conservation, suggesting this DNA may be of functional importance.

Amino Acid Sequence↗
PubMed1986

Estrogen receptors in cultured rat uterine cells: induction of progesterone receptors in the absence of estrogen receptor processing.

When cultured rat uterine cells were treated for up to 6 h with 5 nM 17 beta-estradiol, no decrease in the [3H] estradiol-binding capacity of the cells was observed (i.e. no processing). This was true whether the cells were treated directly with 5 nM [3H]estradiol or with 5 nM unlabeled 17 beta-estradiol followed by homogenization and exchange with [3H]estradiol in vitro. In additional experiments, intact cells were treated with medium containing 5 nM [3H]estradiol for 30 min, and then that medium was removed and replaced with medium containing 5 nM unlabeled 17 beta-estradiol. Receptor-bound estradiol in intact cells was totally exchangeable with estradiol in the culture medium (t1/2, approximately 90 min). Six-hour treatment of cells with 5 nM 17 beta-estradiol led to a 50% increase in the [3H]progesterone-binding capacity of the cells, while no loss of estrogen-binding capacity occurred. These results indicate that progesterone receptors can be induced by estrogen in the rat uterus in the absence of estrogen receptor processing.

Animals↗
PubMed1985

Electron microscopic heteroduplex mapping identifies regions of the engrailed locus that are conserved between Drosophila melanogaster and Drosophila virilis.

Physical localization of mutations in the engrailed (en) gene suggested that at least 70 kilobases (kb) of genomic sequences contribute to the normal function of this gene. Molecular characterization has suggested that en function is encoded in a small, 4.5-kb primary transcript. To identify functional regions within the 70 kb of the en locus of D. melanogaster, we identified sequences conserved in the D. virilis genome (estimated divergence time, 60 million years). Based on homology to D. melanogaster, we isolated en DNA from a D. virilis genomic library. Electron microscopic heteroduplex analysis indicated that in 70 kb there is 20 kb of conserved DNA in 33 different regions dispersed throughout the en locus, including two which encode parts of the major embryonic transcript. The conserved regions are in the same linear order and are spaced by similar lengths of nonconserved sequences in the D. virilis and D. melanogaster DNAs. What functional constraints have enforced conservation of sequences throughout the entire 70 kb and protected the region from divergence of size and arrangement? Our working hypothesis is that sequences necessary for the complex spatial and temporal pattern of en expression are dispersed throughout the 70-kb en locus and that selection for proper regulation restricts evolutionary divergence.

Animals↗