Edward M. Berger

Temporal regulation of microRNA expression in Drosophila melanogaster mediated by hormonal signals and broad-Complex gene activity.

lin-4 and let-7 are founding members of an extensive family of genes that produce small transcripts, termed microRNAs (miRNAs). In Caenorhabditis elegans, lin-4 and let-7 control the timing of postembryonic events by translational repression of target genes, permitting progression from early to late developmental programs. To identify Drosophila melanogaster miRNAs that could play similar roles in the control of developmental timing, we characterized the developmental expression profile of 24 miRNAs in Drosophila, and found 7 miRNAs that are either upregulated or downregulated in conjunction with metamorphosis. The upregulation of three of these miRNAs (mir-100, mir-125, and let-7), and the downregulation of a fourth (mir-34) requires the hormone ecdysone (Ecd) and the activity of the Ecd-inducible gene Broad-Complex. Interestingly, mir-125 is a putative homologue of lin-4. mir-100, -125, and let-7 are clustered within an 800-bp region on chromosome 2L, suggesting that these three miRNAs may be coordinately regulated via common cis-acting elements during metamorphosis. In S2 cells, Ecd and the juvenile hormone analog methoprene exert opposite effects on the expression of these four miRNAs, indicating the participation of both these hormones in the temporal regulation of mir-34, -100, -125, and let-7 expression in vivo.

Ecdysterone induces the transcription of four heat-shock genes in Drosophila S3 cells and imaginal discs

Ecdysterone stimulates a 2- to 15-fold increase in the synthesis of four small heat-shock proteins (hsp) in Drosophila line S3 cells. This is accompanied by a rapid and coincident increase in the abundance of small hsp transcripts. A parallel in vivo situation is now described in imaginal discs isolated from pupariating larvae. In neither system are the high-molecular-weight heat-shock proteins, or their RNAs, induced by hormone. Nor does induction occur in variant cell lines which have acquired resistance to ecdysterone. Other genes within or contiguous to the small hsp cluster are not induced by either ecdysterone or high temperature. Induction by ecdysterone appears to be a primary hormone response.

GENE-ENZYME VARIATION IN THREE SYMPATRIC SPECIES OF LITTORINA

Gene-enzyme variation was examined, electrophoretically, at three non-specific esterase loci in 15 sympatric populations of the prosobranch gastropods: Littorina littorea L. obtusata, and L. saxatilis. Gene frequencies, determined by assuming a correspondence between bands on gels and alleles, revealed marked differences between the species. These differences appear to be correlated with the mechanisms and capabilities for larval disposal in these species.

Small heat shock proteins in Drosophila may confer thermal tolerance

The four small heat shock proteins in Drosophila melanogaster are genetically linked and simultaneously synthesized, both in response to high temperature and, developmentally, during puparium formation. In tissue culture cells their synthesis is inducible by the molting hormone, ecdysterone. We show here that accompanying their induction and accumulation, the cells and animals acquire thermal tolerance.

Ecdysone-induced changes in morphology and protein synthesis in Drosophila cell cultures.

Abstract Permanent cell lines derived from embryonic Drosophila melanogaster show pronounced morphological and biochemical changes in response to the addition of β-ecdysone, the moulting hormone, or two of its analogs. Schneiders line 3 cells, and the French line Kc, morphologically differentiate into what appear to be neural-like cells. During this period levels of acetylcholinesterase activity increase 10- to 30-fold, and DNA synthesis ceases. The level of choline acetyltransferase activity remains unchanged in line 3, and is very low. Schneiders line 2 cells show no morphological transformation in response to β-ecdysone, but undergo a marked change in the pattern of protein synthesis. The six major ecdysone proteins are not “heat shock” proteins.

Polymorphism at the α-glycerophosphate dehydrogenase locus in Drosophila melanogaster. I. Properties of adult allozymes

A biochemical comparison was made between α-glycerophosphate dehydrogenase allozymes from Drosophila melanogaster. Enzymes extracted from the three major genotypes were indistinguishable in terms of their pH optima and thermal stabilities. Distinctive differences were observed for three parameters; temperature dependence of specific activity, temperature dependence of Km, and reaction rate constancy over a physiological temperature range. These results are discussed in terms of a model of balancing selection and the existence of spatial and temporal allele frequency clines in natural populations.

Juvenile hormone signaling during oogenesis in Drosophila melanogaster

Juvenile hormone (JH) participates both in the control of insect development and the establishment of reproductive maturity. In cultured Drosophila cells and in ovarian nurse cells, JH and its synthetic analog, methoprene, induce the expression of two related genes. These genes encode highly similar amino acid transport proteins that are homologous to transporters found in a variety of eukaryotes. JhI-21 is a novel Drosophila gene, and minidiscs (mnd) is a gene that was identified earlier. Two JH-inducible genes are regulated by different molecular mechanisms; JhI-21 behaves as a secondary JH-responsive gene, while mnd behaves as a primary responsive gene. Both JhI-21 and mnd transcripts show developmental profiles, which are consistent with JH regulation. Following eclosion, transcripts from JhI-21 and mnd are synthesized in ovarian nurse cells and subsequently sequestered in the mature egg. Their ectopic accumulation in ovaries can be induced by topical methoprene application. In apterous (ap4) mutant adults defective in JH secretion, mnd and JhI-21 RNA levels are severely reduced, but normal abundance is rescued to a high degree by topical methoprene treatment. Based on the evidence, we propose that during sexual maturation of Drosophila, JH provides a signal to the ovary that leads to the production of several maternally inherited mRNAs.

The juvenile hormone analogue, methoprene, inhibits ecdysterone induction of small heat shock protein gene expression

The small heat shock protein (hsp) genes of Drosophila are expressed in cultured cells in response to the moulting hormone, ecdysterone. We show here that juvenile hormone (JHIII) and the juvenile hormone analogue, methoprene, inhibit that induction in a dose-dependent manner. Heat shock induction is not inhibited. In transient expression studies using S3 line cells transfected with EcRE-CAT constructs, methoprene inhibition was found to require a 2-hr pretreatment (before ecdysterone addition), and methoprenes continued presence was essential. Farnesol, farnesyl acetate, and retinoic acid did not cause inhibition. Several models of methoprene inhibition are discussed.

Steroid and high-temperature induction of the small heat-shock protein genes in Drosophila

Transcription of the four small heat-shock protein genes of Drosophila melanogaster can be induced in cultured cells by high-temperature shock, or by physiological doses of the moulting hormone, ecdysterone. We have characterized and compared the two induction events, focusing on hsp22 and hsp23, in terms of rates of heat-shock protein synthesis, transcription rate, messenger RNA abundance and mRNA half-life. The results indicate that relative to hsp22, the rate of hsp23 synthesis is significantly greater during recovery from heat shock and during ecdysterone induction. This difference is not due to differences in transcription rate, but rather reflects differences in mRNA stability and translational efficiency. One intriguing finding is that hsp message stability is temperature-dependent; hsp transcripts are two to three times more stable at 35 degrees C than at 25 degrees C. The possible mechanism and significance of this phenomenon are discussed.

GENE-ENZYME VARIATION IN THREE SYMPATRIC SPECIES OF LITTORINA. II. THE ROSCOFF POPULATION, WITH A NOTE ON THE ORIGIN OF NORTH AMERICAN L. LITTOREA

Gene-enzyme variation was examined at ten to fifteen loci in three sympatric species of Littorina collected from Cape Cod, Massachusetts and Roscoff, Brittany. The North American and French populations of L. saxatilis and L. obtusata were, in general, quite similar in allele content and frequency. In contrast, the North American and French populations of L. littorea were genetically differentiated at a majority of their loci. This pattern of heterogeneity in the three species, along with calculated values of average heterozygosity, suggests that North American specimens of L. littorea are not recent colonists from Europe and have passed through a severe population bottleneck on the North American continent. Survey studies carried out over a four year period revealed a general pattern of temporal constancy in allele frequency in all three species.