Molecular and General Genetics MGG

, Volume 224, Issue 3, pp 347–356 | Cite as

Direct screening of a small genome : Estimation of the magnitude of plant gene expression changes during adaptation to high salt

  • Gabriele Meyer
  • Jürgen M. Schmitt
  • Hans J. Bohnert


Mesembryanthemum crystallinum (common ice plant), a facultative halophyte with a genome size of 393 000 kb, was used to estimate the magnitude of changes in gene expression in response to environmental stress by excess salt. Such treatment induces a water-conserving pathway of carbon assimilation (CAM) which is, at least in part, transcriptionally controlled. From a genomic library, 200 phage containing approximately 3200 kb (0.8% of the genome) were randomly selected. The inserts in these clones could be divided into four classes ranging from highly repetitive DNA (class I clones) to single-copy DNA (class IV clones). The inserts of the 166 clones of classes II to IV were digested with various restriction enzymes and the fragments were analyzed by hybridization with radioactively labelled mRNA isolated from stressed and unstressed leaves. We found that a total of ∼ 140 DNA fragments hybridized with the RNA probe. Among those, several differentially regulated transcripts were observed. Stress-dependent fluctuation of mRNA abundance was verified by Northern analyses: one mRNA, not detectable in unstressed leaves, appeared in stressed leaves, while steady-state levels of three transcripts decreased during stress. All regulated signals are derived from low abundance mRNAs, which may be missed during screening of cDNA libraries. We conclude from these results that, for the entire genome, on the order of more than one hundred genes are differentially regulated in response to salt stress.

Key words

Genome organization Environmental stress Changes in gene expression Mesembryanthemum crystallinum 


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  1. Bedbrook J (1981) A plant nuclear DNA preparation procedure. Plant Mol Biol Newslett 2:24Google Scholar
  2. Bishop JO, Rosbash M, Evans D (1974) Polynucleotide sequences in eukaryotic DNA and RNA that form ribonuclease-resistant complexes with polyuridylic acid. J Mol Biol 85:75–86Google Scholar
  3. Blum A (1988) Plant breeding for stress environments. CRC Press, Boca RatonGoogle Scholar
  4. Bohnert HJ, Ostrem JA, Cushman JC, Michalowski CB, Rickers J, Meyer G, DeRocher EJ, Vernon DM, Krueger M, Vazquez-Moreno L, Velten J, Höfner R, Schmitt JM (1988) Mesembryanthemum crystallinum, a higher plant model for the study of environmentally induced changes in gene expression. Plant Mol Biol Rep 6:10–28Google Scholar
  5. Cushman JC, Bohnert HJ (1989) Nucleotide sequence of the gene encoding a CAM specific isoform of phosphoenolypruvate carboxylase from Mesembryanthemum crystallinum. Nucleic Acids Res 17:6745–6746Google Scholar
  6. Cushman JC, Meyer G, Michalowski CB, Schmitt JM, Bohnert HJ (1989) Salt stress leads to differential expression of two isogenes of phosphoenolypruvate carboxylase during crassulacean acid metabolism induction in the common ice plant. Plant Cell 1:715–725Google Scholar
  7. Cushman JC, DeRocher EJ, Bohnert HJ (1990) Gene expression during adaptation to salt stress. In: Katterman FR (ed) Environmental injury to plants. Academic Press, New York, pp 173–203Google Scholar
  8. Dellaporta SL, Wood J, Hicks JB (1983) A plant DNA minipreparation: Version II. Plant Mol Biol Rep 1:19–21Google Scholar
  9. DeRocher EJ, Harkins K, Galbraith DW, Bohnert HJ (1990) Systemic endopolyploidy in ice plant is developmentally regulated and may be common to succulents with small genomes. Science 250:99–102Google Scholar
  10. Dvorak J, Ross K, Mendlinger S (1985) Transfer of salt tolerance from Elytrigia pontica (Podp.) Holub to wheat by the addition of an incomplete Elytrigia genome. Crop Sci 25:306–309Google Scholar
  11. Eckenrode VK, Arnold J, Meagher RB (1985) Comparison of the nucleotide sequence of soybean 18S rRNA with the sequence of other small subunit rRNAs. J Mol Evol 21:259–269Google Scholar
  12. Flavell RB (1980) The molecular characterization and organization of plant chromosomal DNA sequences. Annu Rev Plant Physiol 31:569–596CrossRefGoogle Scholar
  13. Flavell RB, Bennett MD, Smith DB (1974) Genome size and the proportion of repeated nucleotide sequence DNA in plants. Biochem Genet 12:257–269Google Scholar
  14. Flavell RB, Rimpau J, Smith DB, O'Dell M, Bedbrook JR (1980) The evolution of plant genome structure. In: Leaver CJ (ed) Genome organization and expression in plants. Plenum, New York, pp 35–47Google Scholar
  15. Frischauf AM, Lehrach H, Poustka A, Murray N (1983) Lambda replacement vectors carrying polylinker sequences. J Mol Biol 170:827–842Google Scholar
  16. Goldberg RB, Hoschek G, Kamalay JC, Timberlake WE (1980) Sequence complexity of nuclear and polysomal RNA in leaves of the tobacco plant. Cell 14:123–131Google Scholar
  17. Henn AM, Gorham J, Lüttge U, WynJones RG (1981) Changes of water-relation characteristics and levels of organic cytoplasmic solutes during salinity induced transition of M. crystallinum from C3-photosynthesis to Crassulacean acid metabolism. Oecologia 50:66–72Google Scholar
  18. Höfner R, Vazquez-Moreno L, Winter K, Bohnert HJ, Schmitt JM (1987) Induction of Crassulacean acid metabolism in M. crystallinum: Mass increase and de novo synthesis of PEP-carboxylase. Plant Physiol 83:915–919Google Scholar
  19. Holtum JAM, Winter K (1982) Activity of enzymes of carbon metabolism during the induction of crassulacean acid metabolism in M. crystallinum. Planta 155:8–16Google Scholar
  20. Leutwiler LS, Hough-Evans BR, Meyerowitz EM (1984) The DNA of Arabidopsis thaliana. Mol Gen Genet 194:15–23Google Scholar
  21. Maniatis T, Fritsch EF, Sambrook J (1982) Molecular cloning: A laboratory manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NYGoogle Scholar
  22. Michalowski CB, Olson SW, Piepenbrock M, Schmitt JM, Bohnert HJ (1989a) Time course of mRNA induction elicited by salt stress in the common ice plant (M. crystallinum). Plant Physiol 89:811–816Google Scholar
  23. Michalowski CB, Schmitt JM, Bohnert HJ (1989b) Expression during salt stress and nucleotide sequence of cDNA for ferredoxin-NADP+ reductase from M. crystallinum. Plant Physiol 89:817–822Google Scholar
  24. Murray MG, Cuellar RE, Thompson WF (1978) DNA sequence organization in the pea genome. Biochemistry 17:5781–5790Google Scholar
  25. Nagl W, Jeanjour M, Kling H, KuehnerS, Michels I, Mueller T, Stein B (1983) Genome and chromatin organization in higher plants. Biol Zentralbl 102:129–148Google Scholar
  26. Norlyn JD, Epstein E (1984) Variability in salt tolerance of four Triticale lines at germination and emergence. Crop Sci 24:1090–1092Google Scholar
  27. Ostrem JA, Olson SW, Schmitt JM, Bohnert HJ (1987) Salt stress increases the level of translatable mRNA for phosphoenolpyruvate carboxylase in M. crystallinum. Plant Physiol 84:1270–1275Google Scholar
  28. Ostrem JA, Vernon DM, Bohnert HJ (1990) Increased expression of a gene coding for NAD:glyceraldehyde-3-phosphate dehydrogenase during the transition from C3 photosynthesis to CAM in M. crystallinum. J Biol Chem 265:3497–3502Google Scholar
  29. Patterson TA, Dean M (1987) Preparation of high titer lambda phage lysates. Nucleic Acids Res 15:6298Google Scholar
  30. Paul MJ, Cockburn W (1989) Pinitol, a compatible solute in Mesembryanthemum crystallinum L.? J Exp Bot 40:1093–1098Google Scholar
  31. Pruitt RE, Meyerowitz EM (1986) Characterization of the genome of Arabidopsis thaliana. J Mol Biol 187:169–183Google Scholar
  32. Rickers J, Cushman JC, Michalowski CB, Schmitt JM, Bohnert HJ (1989) Expression of the CAM-form of phosphoenolypruvate carboxylase and nucleotide sequence of a full-length cDNA from Mesembryanthemum crystallinum. Mol Gen Genet 215:447–154Google Scholar
  33. Ting IP (1985) Crassulacean acid metabolism. Annu Rev Plant Physiol 37:595–622Google Scholar
  34. Treichel S (1986) The influence of NaCl on Δ-pyrroline-5-carboxyl-late reductase in proline-accumulating cell suspension cultures of M. nodiflorum and other halophytes. Physiol Plant 67:173–181Google Scholar
  35. Vernon DM, Ostrem JA, Schmitt JM, Bohnert HJ (1988) PEPCase transcript levels in Mesembryanthemum crystallinum decline rapidly upon relief from salt stress. Plant Physiol 86:1002–1004Google Scholar
  36. Winter K, von Willert DJ (1972) NaCl induzierter Crassulacean-Säurestoffwechsel bei Mesembryanthemum crystallinum. Z Pflanzenphysiol 67:166–170Google Scholar

Copyright information

© Springer-Verlag 1990

Authors and Affiliations

  • Gabriele Meyer
    • 1
  • Jürgen M. Schmitt
    • 2
  • Hans J. Bohnert
    • 1
    • 3
  1. 1.Department of BiochemistryThe University of ArizonaTucsonUSA
  2. 2.Botanisches InstitutUnversität WürzburgWürzburgGermany
  3. 3.Departments of Molecular & Cellular Biology and of Plant SciencesThe University of ArizonaTucsonUSA

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