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. 1987 Mar 25;15(6):2549–2561. doi: 10.1093/nar/15.6.2549

20-Hydroxyecdysone regulates cytoplasmic actin gene expression in Drosophila cultured cells.

J L Couderc, L Hilal, M L Sobrier, B Dastugue
PMCID: PMC340668  PMID: 2436146

Abstract

The steroid hormone 20-hydroxyecdysone (20-OHE) induces, in Kc cultured Drosophila melanogaster cells, important morphological transformations and specific changes of enzymatic activities and of protein synthesis. These changes are accompanied by an increase of synthesis and an accumulation of actin. Specific probes were used to reveal transcripts of each actin gene in mRNA populations isolated from cells at various times of 20-OHE treatment. Only the two cytoplasmic actin genes 5C and 42A are expressed in Kc cells and the hormone induces the accumulation of transcripts of these two genes. We have also taken advantage of S1 mapping and extension procedures to identify the 5' ends of the actin mRNAs from these two genes and to compare their respective levels of expression. The 5C gene is more expressed than the 42A one in untreated and in hormone treated cells. The 5C gene encodes three RNAs that differ in their 3' end. The two genes are interrupted by an intervening sequence immediately upstream of ATG initiation codon but not at the same position. The transcription rate for the two genes is increased up to five fold upon 20-OHE treatment, demonstrating a direct effect of the steroid hormone at the transcriptional level for these genes.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aviv H., Leder P. Purification of biologically active globin messenger RNA by chromatography on oligothymidylic acid-cellulose. Proc Natl Acad Sci U S A. 1972 Jun;69(6):1408–1412. doi: 10.1073/pnas.69.6.1408. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bergsma D. J., Chang K. S., Schwartz R. J. Novel chicken actin gene: third cytoplasmic isoform. Mol Cell Biol. 1985 May;5(5):1151–1162. doi: 10.1128/mcb.5.5.1151. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bingham P. M., Levis R., Rubin G. M. Cloning of DNA sequences from the white locus of D. melanogaster by a novel and general method. Cell. 1981 Sep;25(3):693–704. doi: 10.1016/0092-8674(81)90176-8. [DOI] [PubMed] [Google Scholar]
  4. Breathnach R., Chambon P. Organization and expression of eucaryotic split genes coding for proteins. Annu Rev Biochem. 1981;50:349–383. doi: 10.1146/annurev.bi.50.070181.002025. [DOI] [PubMed] [Google Scholar]
  5. Clarke M., Spudich J. A. Nonmuscle contractile proteins: the role of actin and myosin in cell motility and shape determination. Annu Rev Biochem. 1977;46:797–822. doi: 10.1146/annurev.bi.46.070177.004053. [DOI] [PubMed] [Google Scholar]
  6. Couderc J. L., Cadic A. L., Sobrier M. L., Dastugue B. Ecdysterone induction of actin synthesis and polymerization in a Drosophila melanogaster cultured cell line. Biochem Biophys Res Commun. 1982 Jul 16;107(1):188–195. doi: 10.1016/0006-291x(82)91687-4. [DOI] [PubMed] [Google Scholar]
  7. Couderc J. L., Dastugue B. Ecdysterone-induced modifications of protein synthesis in a Drosophila melanogaster cultured cell line. Biochem Biophys Res Commun. 1980 Nov 17;97(1):173–181. doi: 10.1016/s0006-291x(80)80151-3. [DOI] [PubMed] [Google Scholar]
  8. Couderc J. L., Sobrier M. L., Giraud G., Becker J. L., Dastugue B. Actin gene expression is modulated by ecdysterone in a Drosophila cell line. J Mol Biol. 1983 Mar 5;164(3):419–430. doi: 10.1016/0022-2836(83)90059-1. [DOI] [PubMed] [Google Scholar]
  9. Courgeon A. M. Action of insect hormones at the cellular level. Morphological changes of a diploid cell line of Drosophila melanogaster, treated with ecdysone and several analogues in vitro. Exp Cell Res. 1972 Oct;74(2):327–336. doi: 10.1016/0014-4827(72)90384-9. [DOI] [PubMed] [Google Scholar]
  10. Echalier G., Ohanessian A. Isolement, en cultures in vitro, de lignées cellulaires diploïdes de Drosophila melanogaster. C R Acad Sci Hebd Seances Acad Sci D. 1969 Mar 31;268(13):1771–1773. [PubMed] [Google Scholar]
  11. Fyrberg E. A., Bond B. J., Hershey N. D., Mixter K. S., Davidson N. The actin genes of Drosophila: protein coding regions are highly conserved but intron positions are not. Cell. 1981 Apr;24(1):107–116. doi: 10.1016/0092-8674(81)90506-7. [DOI] [PubMed] [Google Scholar]
  12. Fyrberg E. A., Mahaffey J. W., Bond B. J., Davidson N. Transcripts of the six Drosophila actin genes accumulate in a stage- and tissue-specific manner. Cell. 1983 May;33(1):115–123. doi: 10.1016/0092-8674(83)90340-9. [DOI] [PubMed] [Google Scholar]
  13. Gross-Bellard M., Oudet P., Chambon P. Isolation of high-molecular-weight DNA from mammalian cells. Eur J Biochem. 1973 Jul 2;36(1):32–38. doi: 10.1111/j.1432-1033.1973.tb02881.x. [DOI] [PubMed] [Google Scholar]
  14. Groudine M., Peretz M., Weintraub H. Transcriptional regulation of hemoglobin switching in chicken embryos. Mol Cell Biol. 1981 Mar;1(3):281–288. doi: 10.1128/mcb.1.3.281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Keller E. B., Noon W. A. Intron splicing: a conserved internal signal in introns of Drosophila pre-mRNAs. Nucleic Acids Res. 1985 Jul 11;13(13):4971–4981. doi: 10.1093/nar/13.13.4971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Lehrach H., Diamond D., Wozney J. M., Boedtker H. RNA molecular weight determinations by gel electrophoresis under denaturing conditions, a critical reexamination. Biochemistry. 1977 Oct 18;16(21):4743–4751. doi: 10.1021/bi00640a033. [DOI] [PubMed] [Google Scholar]
  17. Maniatis T., Hardison R. C., Lacy E., Lauer J., O'Connell C., Quon D., Sim G. K., Efstratiadis A. The isolation of structural genes from libraries of eucaryotic DNA. Cell. 1978 Oct;15(2):687–701. doi: 10.1016/0092-8674(78)90036-3. [DOI] [PubMed] [Google Scholar]
  18. McKeown M., Firtel R. A. Differential expression and 5' end mapping of actin genes in Dictyostelium. Cell. 1981 Jun;24(3):799–807. doi: 10.1016/0092-8674(81)90105-7. [DOI] [PubMed] [Google Scholar]
  19. McKnight G. S., Palmiter R. D. Transcriptional regulation of the ovalbumin and conalbumin genes by steroid hormones in chick oviduct. J Biol Chem. 1979 Sep 25;254(18):9050–9058. [PubMed] [Google Scholar]
  20. Messing J., Vieira J. A new pair of M13 vectors for selecting either DNA strand of double-digest restriction fragments. Gene. 1982 Oct;19(3):269–276. doi: 10.1016/0378-1119(82)90016-6. [DOI] [PubMed] [Google Scholar]
  21. Micard D., Sobrier M. L., Couderc J. L., Dastugue B. Purification of RNA-free plasmid DNA using alkaline extraction followed by Ultrogel A2 column chromatography. Anal Biochem. 1985 Jul;148(1):121–126. doi: 10.1016/0003-2697(85)90636-0. [DOI] [PubMed] [Google Scholar]
  22. Palmiter R. D. Magnesium precipitation of ribonucleoprotein complexes. Expedient techniques for the isolation of undergraded polysomes and messenger ribonucleic acid. Biochemistry. 1974 Aug 13;13(17):3606–3615. doi: 10.1021/bi00714a032. [DOI] [PubMed] [Google Scholar]
  23. Sanchez F., Tobin S. L., Rdest U., Zulauf E., McCarthy B. J. Two Drosophila actin genes in detail. Gene structure, protein structure and transcription during development. J Mol Biol. 1983 Feb 5;163(4):533–551. doi: 10.1016/0022-2836(83)90111-0. [DOI] [PubMed] [Google Scholar]
  24. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Snyder M., Hunkapiller M., Yuen D., Silvert D., Fristrom J., Davidson N. Cuticle protein genes of Drosophila: structure, organization and evolution of four clustered genes. Cell. 1982 Jul;29(3):1027–1040. doi: 10.1016/0092-8674(82)90466-4. [DOI] [PubMed] [Google Scholar]
  26. Sobrier M. L., Couderc J. L., Chapel S., Dastugue B. Expression of a new beta tubulin subunit is induced by 20-hydroxyecdysone in Drosophila cultured cells. Biochem Biophys Res Commun. 1986 Jan 14;134(1):191–200. doi: 10.1016/0006-291x(86)90546-2. [DOI] [PubMed] [Google Scholar]
  27. Thomas P. S. Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5201–5205. doi: 10.1073/pnas.77.9.5201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Tobin S. L., Zulauf E., Sánchez F., Craig E. A., McCarthy B. J. Multiple actin-related sequences in the Drosophila melanogaster genome. Cell. 1980 Jan;19(1):121–131. doi: 10.1016/0092-8674(80)90393-1. [DOI] [PubMed] [Google Scholar]
  29. Vandekerckhove J., Weber K. Mammalian cytoplasmic actins are the products of at least two genes and differ in primary structure in at least 25 identified positions from skeletal muscle actins. Proc Natl Acad Sci U S A. 1978 Mar;75(3):1106–1110. doi: 10.1073/pnas.75.3.1106. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Vitek M. P., Morganelli C. M., Berger E. M. Stimulation of cytoplasmic actin gene transcription and translation in cultured Drosophila cells by ecdysterone. J Biol Chem. 1984 Feb 10;259(3):1738–1743. [PubMed] [Google Scholar]
  31. Zakut R., Shani M., Givol D., Neuman S., Yaffe D., Nudel U. Nucleotide sequence of the rat skeletal muscle actin gene. Nature. 1982 Aug 26;298(5877):857–859. doi: 10.1038/298857a0. [DOI] [PubMed] [Google Scholar]

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