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P Cherbas

Publications and source records attributed to P Cherbas.

28 records · Page 2Linked to original sources
PubMed1989

Diverse expression of overlapping genes: the Drosophila Eip28/29 gene and its upstream neighbors.

The Eip28/29 gene of Drosophila is known to be regulated by the steroid hormone ecdysone in Kc cells and other cell lines. An investigation of Eip28/29 gene expression in intact animals has led to the discovery of two new genes on its 5' flank. Together these three genes generate at least seven distinct transcripts with diverse patterns of developmental expression. gonadal (gdl) is transcribed on the same strand as Eip28/29 and is expressed in two modes. gdlM transcripts, observed exclusively in the testes, are 1200 and 1500 N long, differing by their polyadenylation site but probably otherwise identical; gdlF transcripts are 900 and 1200 N long and share their terminal exons with the two gdlM transcripts. In adults they are exclusively ovarian, but they are also present in early embryos and, at a lower abundance, in Kc cells. In each mode, the longer transcript results from use of a polyadenylation site within the 5' exon of Eip28/29. The shared region includes 3' noncoding sequences of gdl transcripts and 5' flanking DNA and 5' noncoding sequences of Eip28/29. gdl expression in Kc cells is, however, unaffected by ecdysone. z600 is more distal to Eip28/29 but still contained at least in part within the 2 kb upstream of that gene. Its 600 N transcript is expressed predominantly during the first few hours of embryogenesis. Finally, the Eip28/29 transcripts are present at low levels during most developmental stages.

Animals↗
PubMed1988

26-[125I]iodoponasterone A is a potent ecdysone and a sensitive radioligand for ecdysone receptors.

The effects of ecdysone, the steroid molting hormone of arthropods, are of considerable interest both to insect physiologists and to those studying steroid-regulated gene expression. Yet progress in understanding ecdysone receptors has been inhibited by the lack of a suitable highly radioactive hormone analog with high affinity for the receptor. Here we report that the synthetic ecdysteroid 26-iodoponasterone A is one of the most active ecdysones known, inducing half-maximal morphological transformation in Drosophila Kc167 cells when present at 0.5 nM. 26-[125I]Iodoponasterone A can be prepared at a specific activity of 2175 Ci/mmol (1 Ci = 37 GBq) by reaction of the precursor 26-mesylinokosterone with carrier-free Na125I. The radiolabeled material binds to Kc167 cell ecdysone receptors specifically and with affinity (Kd ca. 3.8 X 10(-10) M). Thus, 26-[125I]iodoponasterone A appears to be a superior radioligand for ecdysone receptors on grounds both of affinity and of specific activity. Its ready availability should greatly facilitate studies of these receptors.

Animals↗
PubMed1986

Structure of the Eip28/29 gene, an ecdysone-inducible gene from Drosophila.

The EIPs 28 and 29 are a family of polypeptides identified originally by their ecdysone inducibility in Drosophila cell lines. At least two family members, 28III and 29III, appear to be primary translation products. Here we describe a unique Eip28/29 gene that must encode both primary products. The Eip28/29 gene is unique because the cloned genomic DNA hybridizes to both EIP 28 and 29 messenger RNAs under stringent conditions, but does not anneal detectably to other genomic sequences even under mild conditions. Furthermore the diverse products of this gene are not alleles because flies homozygous for the chromosomal region (71CD) containing the Eip28/29 gene produce mRNAs that translate to yield all the EIPs 28 and 29. We report here the sequence of a 2855-nucleotide region encompassing the Eip28/29 gene. By comparisons with complementary DNA sequences and by nuclease protection experiments we have derived a complete structure for the Eip28/29 transcription unit. The primary transcript is 2146 nucleotides long and is processed by the removal of three introns to yield the predominant mature transcript in tissue culture cells (979 nucleotides). This transcript probably corresponds to the 28III mRNA. Neither the start of the transcription unit nor the structure of the predominant transcript is affected by the hormone ecdysone. The genomic sequence reveals a series of heptanucleotide and octanucleotide repeats of unknown function that fall at about 50-nucleotide intervals within the first 150 nucleotides upstream from the transcription unit. In addition this sequence, when combined with previously published data, suggests that the consensus cap site sequence in Drosophila may be extended to include 13 nucleotides centered on the heptanucleotide core previously recognized by Snyder et al. (1982).

Animals↗
PubMed1986

Alternative splicing generates two distinct Eip28/29 gene transcripts in Drosophila Kc cells.

The Drosophila Eip28/29 gene encodes two primary translation products, ecdysone-inducible polypeptide (EIP) 28III and EIP 29III. When cells of the Kc cell line are treated with the steroid hormone ecdysone, the number of Eip28/29 transcripts and the synthesis of the various forms of EIP 28 and 29 increase rapidly. We have reported the sequence of the Eip28/29 gene and of its major transcript. Here we describe a minor or short-form transcript that is about 25% of the total Eip28/29 gene transcripts in both untreated and hormone-treated cells. This transcript is formed by the use of an alternative splice donor sequence 12 nucleotides upstream from the major donor site at the end of the second exon. Evidently the relative abundance of the two products is not hormonally regulated. The short form translation product should lack only an internal dibasic tetrapeptide. The long and short forms probably represent distinct mRNAs for EIP 28III and EIP 29III, respectively.

Animals↗
PubMed1984

cDNA clones for the ecdysone-inducible polypeptide (EIP) mRNAs of Drosophila Kc cells.

The ecdysone-inducible polypeptides (EIPs) 28, 29 and 40 were identified previously as polypeptides whose synthesis is stimulated early in the ecdysone response of Drosophila Kc cells. We have now shown, using two-dimensional gels, that each of these EIPs consists of three species differing in pI, and all stimulated by ecdysone. Translations and hybrid-arrested translations indicated that the poly(A) EIP mRNAs increase 10-fold in abundance during the first 4 h of ecdysone treatment. By a differential screen of a cDNA library we have identified cDNA clones corresponding to all three EIPs. Two kinds of clones were isolated: one hybridizes to the EIP 40 mRNA(s); the second hybridizes to the mRNA(s) encoding all the EIPs 28 and 29. The EIP 28/29 and EIP 40 loci detected by these clones are each present at single sites on the polytene chromosomes and each is at or in the vicinity of an ecdysone-regulated puff.

Journal Article↗
PubMed1980

Ecdysteroid-inducible polypeptides in a Drosophila cell line.

In the Drosophila melanogaster cell line Kc-H, ecdysteroid hormone treatment causes increased relative synthesis of three ecdysteroid-inducible polypeptides (EIPs), named according to their molecular weights (in kilodaltons) EIP 40, EIP 29 and EIP 28. Increased synthesis of the EIPs is detectable within 45 min (EIP 28) or 75 min (EIPs 40 and 29), is maximal at 4-8 hr and continues for almost 2 days. During this period no other major changes in protein synthesis are discernible using one-dimensional gels. At maximum, EIP 28 synthesis is elevated at least 10 fold above its basal level, and EIPs 40 and 29 somewhat less. EIP induction is ecdysteroid-specific and is detectable in the presence of 10(-8) M 20-hydroxyecdysone. It does not occur in hormone-resistant cells. Apparently identical polypeptides are inducible in another ecdysteroid-responsive cell line, Schneider's line 3. Because EIP synthesis is an early and substantial response to ecdysteroids, this is a promising system for the study of steroid hormone action.

Animals↗
PubMedDate not supplied

The control of puffing by ions - the Kroeger hypothesis: a critical review.

We have re-examined the several papers which appear to us to represent the principal lines of evidence for what we call the Kroeger hypothesis. To do this we have stated this hypothesis in its simplest, most concrete form, a form that has been repeatedly and forceably enunciated in the literature (Kroeger, 1963a, 1965, 1966, 1967, 1968; Kroeger and Lezzi 1966; Lezzi and Frigg, 1971). The evidence suggests to us that ecdysone's effect on puffing is probably not mediated by the [K+]/[Na+]. While such a model cannot, even now, be excluded, we see little reason to believe in it. We take the general issues raised by Kroeger's ideas very seriously. Nucleoprotein complexes are exquisitely sensitive to changes in salt concentration and ionic selectivity is a well-known property of proteins (and of ion-exchangers in general, see, for example, Diamond and Wright, 1969). Thus it might not be shocking if cells utilized this specificity in some general control over chromsosome structure, perhaps a second-layer of control superimposed upon other transcriptional controls. Therefore it is our feeling that Kroeger's data merits very careful and critical study, the more so because the experiments involved are intrinsically difficult. It is in this vein that we have tried to review Kroeger's data.

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