• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of molcellbPermissionsJournals.ASM.orgJournalMCB ArticleJournal InfoAuthorsReviewers
Mol Cell Biol. Jan 1987; 7(1): 149–159.
PMCID: PMC365051

Identification of the sequences controlling cyclic AMP regulation and cell-type-specific expression of a prestalk-specific gene in Dictyostelium discoideum.

Abstract

We have cloned and analyzed a developmentally and spatially regulated prestalk cell-specific gene from Dictyostelium discoideum. The gene encodes a protein highly homologous to the lysosomal cysteine proteinases cathepsin H and cathepsin B. Amino acid comparisons between these enzymes showed that the active-site amino acids were conserved, as were amino acids known to be important for catalysis and residues which form the intramolecular cysteine bridges. We have constructed a series of internal deletions, duplications, and linker scanner mutations within the region 300 base pairs 5' to the cap site. Analysis of expression of the mutations in transformants identified a approximately 35-base pair GC-rich region containing a dAdC/dGdT palindromic repeat and a G-rich box which is homologous to the 3' GT half of the palindromic repeat. Deletion or disruption of the G box resulted in a approximately 50-fold drop in the level of expression of the gene fusion in transformants in response to cyclic AMP in single-cell culture but did not affect the temporal pattern of regulation or control by cyclic AMP. The expression of such constructs during normal multicellular differentiation paralleled that of the endogenous gene; however, the level of RNA from the constructs was only approximately 10-fold lower than that of constructs containing the G box. Deletion of the 3' half of the palindromic sequence and the G box region resulted in a dramatic decrease in the level of transcription, although the constructs still showed proper temporal expression. These results suggest that this 35-base-pair region acts as an important part of the regulatory region for cell type and cyclic AMP regulation.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (2.2M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Images in this article

Click on the image to see a larger version.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Barklis E, Lodish HF. Regulation of dictyostelium discoideum mRNAs specific for prespore or prestalk cells. Cell. 1983 Apr;32(4):1139–1148. [PubMed]
  • Berk AJ, Sharp PA. Spliced early mRNAs of simian virus 40. Proc Natl Acad Sci U S A. 1978 Mar;75(3):1274–1278. [PMC free article] [PubMed]
  • Breathnach R, Chambon P. Organization and expression of eucaryotic split genes coding for proteins. Annu Rev Biochem. 1981;50:349–383. [PubMed]
  • Chisholm RL, Barklis E, Lodish HF. Mechanism of sequential induction of cell-type specific mRNAs in Dictyostelium differentiation. Nature. 1984 Jul 5;310(5972):67–69. [PubMed]
  • Datta S, Gomer RH, Firtel RA. Spatial and temporal regulation of a foreign gene by a prestalk-specific promoter in transformed Dictyostelium discoideum. Mol Cell Biol. 1986 Mar;6(3):811–820. [PMC free article] [PubMed]
  • Devine KM, Loomis WF. Molecular characterization of anterior-like cells in Dictyostelium discoideum. Dev Biol. 1985 Feb;107(2):364–372. [PubMed]
  • Doolittle RF. Similar amino acid sequences: chance or common ancestry? Science. 1981 Oct 9;214(4517):149–159. [PubMed]
  • Dowds BC, Loomis WF. Cloning and expression of a cDNA that comprises part of the gene coding for a spore coat protein of Dictyostelium discoideum. Mol Cell Biol. 1984 Nov;4(11):2273–2278. [PMC free article] [PubMed]
  • Firtel RA, Bonner J. Developmental control of -1-4 glucan phosphorylase in the cellular slime mold Dictyostelium discoideum. Dev Biol. 1972 Sep;29(1):85–103. [PubMed]
  • Firtel RA, Bonner J. Characterization of the genome of the cellular slime mold Dictyostelium discoideum. J Mol Biol. 1972 May 28;66(3):339–361. [PubMed]
  • Fong D, Rutherford CL. Protease activity during cell differentiation of the cellular slime mold Dictyostelium discoideum. J Bacteriol. 1978 May;134(2):521–527. [PMC free article] [PubMed]
  • Gomer RH, Datta S, Mehdy M, Crowley T, Sivertsen A, Nellen W, Reymond C, Mann S, Firtel RA. Regulation of cell-type-specific gene expression in Dictyostelium. Cold Spring Harb Symp Quant Biol. 1985;50:801–812. [PubMed]
  • Grunstein M, Hogness DS. Colony hybridization: a method for the isolation of cloned DNAs that contain a specific gene. Proc Natl Acad Sci U S A. 1975 Oct;72(10):3961–3965. [PMC free article] [PubMed]
  • Haribabu B, Dottin RP. Pharmacological characterization of cyclic AMP receptors mediating gene regulation in Dictyostelium discoideum. Mol Cell Biol. 1986 Jul;6(7):2402–2408. [PMC free article] [PubMed]
  • Kimmel AR, Firtel RA. Sequence organization in Dictyostelium: unique structure at the 5'-ends of protein coding genes. Nucleic Acids Res. 1983 Jan 25;11(2):541–552. [PMC free article] [PubMed]
  • Kirschke H, Wood L, Roisen FJ, Bird JW. Activity of lysosomal cysteine proteinase during differentiation of rat skeletal muscle. Biochem J. 1983 Sep 15;214(3):871–877. [PMC free article] [PubMed]
  • Maxam AM, Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. [PubMed]
  • McKeown M, Firtel RA. Differential expression and 5' end mapping of actin genes in Dictyostelium. Cell. 1981 Jun;24(3):799–807. [PubMed]
  • McKnight SL, Kingsbury R. Transcriptional control signals of a eukaryotic protein-coding gene. Science. 1982 Jul 23;217(4557):316–324. [PubMed]
  • Mehdy MC, Firtel RA. A secreted factor and cyclic AMP jointly regulate cell-type-specific gene expression in Dictyostelium discoideum. Mol Cell Biol. 1985 Apr;5(4):705–713. [PMC free article] [PubMed]
  • Mehdy MC, Ratner D, Firtel RA. Induction and modulation of cell-type-specific gene expression in Dictyostelium. Cell. 1983 Mar;32(3):763–771. [PubMed]
  • Nellen W, Firtel RA. High-copy-number transformants and co-transformation in Dictyostelium. Gene. 1985;39(2-3):155–163. [PubMed]
  • Nellen W, Silan C, Firtel RA. DNA-mediated transformation in Dictyostelium discoideum: regulated expression of an actin gene fusion. Mol Cell Biol. 1984 Dec;4(12):2890–2898. [PMC free article] [PubMed]
  • Oyama M, Blumberg DD. Interaction of cAMP with the cell-surface receptor induces cell-type-specific mRNA accumulation in Dictyostelium discoideum. Proc Natl Acad Sci U S A. 1986 Jul;83(13):4819–4823. [PMC free article] [PubMed]
  • Pears CJ, Mahbubani HM, Williams JG. Characterization of two highly diverged but developmentally co-regulated cysteine proteinase genes in Dictyostelium discoideum. Nucleic Acids Res. 1985 Dec 20;13(24):8853–8866. [PMC free article] [PubMed]
  • Poole SJ, Firtel RA. Genomic instability and mobile genetic elements in regions surrounding two discoidin I genes of Dictyostelium discoideum. Mol Cell Biol. 1984 Apr;4(4):671–680. [PMC free article] [PubMed]
  • Poole S, Firtel RA, Lamar E, Rowekamp W. Sequence and expression of the discoidin I gene family in Dictyostelium discoideum. J Mol Biol. 1981 Dec 5;153(2):273–289. [PubMed]
  • Proudfoot NJ, Brownlee GG. 3' non-coding region sequences in eukaryotic messenger RNA. Nature. 1976 Sep 16;263(5574):211–214. [PubMed]
  • Reymond CD, Gomer RH, Mehdy MC, Firtel RA. Developmental regulation of a Dictyostelium gene encoding a protein homologous to mammalian ras protein. Cell. 1984 Nov;39(1):141–148. [PubMed]
  • Romans P, Firtel RA. Organization of the Dictyostelium discoideum actin multigene family. Flanking sequences show subfamily homologies and unusual dyad symmetries. J Mol Biol. 1985 Jun 5;183(3):311–326. [PubMed]
  • Romans P, Firtel RA, Saxe CL., 3rd Gene-specific expression of the actin multigene family of Dictyostelium discoideum. J Mol Biol. 1985 Nov 20;186(2):337–355. [PubMed]
  • Sanger F, Nicklen S, Coulson AR. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. [PMC free article] [PubMed]
  • Saxe CL, 3rd, Firtel RA. Analysis of gene expression in rapidly developing mutants of Dictyostelium discoideum. Dev Biol. 1986 Jun;115(2):407–414. [PubMed]
  • Schaap P, van Driel RV. Induction of post-aggregative differentiation in Dictyostelium discoideum by cAMP. Evidence of involvement of the cell surface cAMP receptor. Exp Cell Res. 1985 Aug;159(2):388–398. [PubMed]
  • Seshimo H, Ryuzaki M, Yoshizato K. Specific inhibition of Triiodothyronine-induced tadpole tail-fin regression by cathepsin D-inhibitor pepstatin. Dev Biol. 1977 Aug;59(1):96–100. [PubMed]
  • Southern EM. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. [PubMed]
  • Staden R. Graphic methods to determine the function of nucleic acid sequences. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 2):521–538. [PMC free article] [PubMed]
  • WHITE GJ, SUSSMAN M. Metabolism of major cell components during slime mold morphogenesis. Biochim Biophys Acta. 1961 Oct 28;53:285–293. [PubMed]
  • Williams JG, North MJ, Mahbubani H. A developmentally regulated cysteine proteinase in Dictyostelium discoideum. EMBO J. 1985 Apr;4(4):999–1006. [PMC free article] [PubMed]
  • Young RA, Davis RW. Efficient isolation of genes by using antibody probes. Proc Natl Acad Sci U S A. 1983 Mar;80(5):1194–1198. [PMC free article] [PubMed]

Articles from Molecular and Cellular Biology are provided here courtesy of American Society for Microbiology (ASM)

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...