Abstract
The promoter region of the Agrobacterium tumefaciens T-cyt gene was linked in a translational fusion to the coding DNA of the reporter gene uidA (for β-glucuronidase or GUS protein; EC 3.2.1.31) and to nos 3′ flanking DNA. The chimaeric gene was introduced by Agrobacterium transformation into potato (Solanum tuberosum L. cv. Désirée). In nine transgenic lines, the average GUS levels were highest in extracts from stems and roots of in vitro grown plants (ca. 11 000 GUS activity units per pmol MU per mg protein per min) but lower in leaves of the in vitro grown plants (ca. 7000 units). GUS activity was intermediate in stems and roots of plants grown in soil as well as in in vitro crown galls (ca. 3000 units). Activity was low in tubers, irrespective of whether these developed in vitro or in soil (both ca. 100 units), and lowest of all in leaves of soil-grown plants (ca. 10–15 units). However, in shoot cultures reestablished from soil-grown plants, GUS activity in the leaves increased to that determined in the original shoot cultures. Hence, plant culture conditions strongly influenced the expression of the T-cyt-uidA-nos gene. In particular, it was silenced in leaves of soil-grown plants. The results are compared with previous analyses of the promoter region of the wild-type T-cyt gene and with the growth properties of a large number of crown gall cell lines and crown-gall-derived plants, including over forty S. tuberosum cv. Désirée cell lines isolated in the present study that were transformed with the wild-type T-cyt gene and six promoter-mutated derivatives. A number of implications are discussed for crown gall formation and for control of expression of plant genes which contain Activator or G-box type 5′ expression control sequences.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Akiyoshi, DE, Klee, H, Amasino, RM, Nester, EW, Gordon, MP: The T-DNA of Agrobacterium tumefaciens codes for an enzyme of cytokinin biosynthesis. Proc Natl Acad Sci USA 81: 5994–5998 (1984).
Amasino, RM, Powell, ALT, Gordon, MP: Changes in T-DNA methylation and expression are associated with phenotypic variation and plant regeneration in a crown gall line. Mol Gen Genet 197: 437–446 (1984).
Barry, GF, Rogers, SG, Fraley, RT, Brand, L: Identification of a cloned cytokinin biosynthetic gene. Proc Natl Acad Sci USA 81: 4776–4780 (1984).
Bevan, MW: Binary Agrobacterium vectors for plant transformation. Nucl Acids Res 12: 8711–8721 (1984).
Binns, AN: The biology and molecular biology of plant cells infected by Agrobacterium tumefaciens. In: Miflin, BJ (ed), Plant Molecular and Cell Biology vol 1, pp. 133–160. Oxford University Press, Oxford (1984).
Buchmann, I, Marner, F-J, Schroeder, G, Waffenschmidt, S, Schroeder, J: Tumour genes in plants: T-DNA encoded cytokinin biosynthesis. EMBO J 4: 853–859 (1985).
Burrell, MM, Twell, D, Karp, A, Ooms, G: Expression of Ti T-DNA in differentiated tissues of potato (Solanum tuberosum cv. Maris Bard). Plant Mol Biol 5: 213–222 (1985).
Chen, ZL, Schuler, MA, Beachy, RN: Functional analysis of regulatory elements on a plant embryo-specific gene. Proc Natl Acad Sci USA 83: 8560–8564 (1986).
Cleland, RE: Is plant development regulated by changes in the concentration of growth substances or by changes in the sensitivity to growth substances? Changes in hormone concentration are important too. Trends Biochem Sci 8: 354–357 (1983).
Davies, PJ: The plant hormone concept: transport, concentration and sensitivity. In: Davies, PJ (ed), The Plant Hormones and their Role in Plant Growth and Development, pp. 12–23. Martinus Nijhoff, Dordrecht (1987).
Dean, C, Jones, J, Favreau, M, Dunsmuir, P, Bedbrook, J: Influence of flanking sequences on variability in expression levels of an introduced gene in transgenic tobacco plants. Nucl Acids Res 16: 9267–9283 (1988).
Dean, C, Pickersky, E, Dunsmuir, P: Structure, evolution and regulation of RbcS genes in higher plants. Ann Rev of Plant Physiol Plant Mol Biol 40: 415–439 (1989).
de Pater BS: Plant expression signals of the Agrobacterium T-cyt gene. Thesis, Leiden University, Netherlands (1987).
de, Pater, BS, Klinkhamer, MP, Amesz, PA, de, Kam, RJ, Memelink, J, Hoge, JHC, Schilperoort, RA: Plant expression signals of the Agrobacterium T-cyt gene. Nucl Acids Res 15: (20) 8267–8282 (1987).
de, Pater, BS, de, Kam, RJ, Hoge, JHC, Schilperoort, RA: Effects of mutations in the TATA box region of the Agrobacterium T-cyt gene on its transcription in plant tissues. Nucl Acids Res 15: 8283–8292 (1987).
Ferl, RJ, Laugher, BH: In vivo detection of regulatory factors binding sites of Arabidopsis thaliana Adh. Plant Mol Biol 12: 357–366 (1989).
Fluhr, R, Kuhlemeier, C, Nagy, F, Chua, N: Organ-specific and light-induced expression of plant genes. Science 232: 1106–1112 (1986).
Giuliano, G, Pichersky, E, Malik, US, Timko, MP, Scolnik, PA, Cashmore, AR: An evolutionarily conserved protein binding sequence upstream of a plant light regulated gene. Proc Natl Acad Sci USA 85: 7089–7093 (1988).
Hänischten Cate, CL, Ennik, E, Roest, S, Ramulu, KS, Dihkhuis, P, De, Groot, B: Regeneration and characterization of plants from potato root lines transformed by Agrobacterium rhizogenes. Theor Appl Genet 75: 452–459 (1988).
Hooykaas PJJ, Schilperoort RA: The molecular genetics of crown gall tumour genesis. Adv Genet 22: 210–283.
Jefferson, RA, Kavanagh, TA, Bevan, MW: GUS fusions: beta glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J 6: 3901–3908 (1987).
Jefferson, RA: Assaying chimaeric genes in plants. The GUS gene fusion system. Plant Mol Biol Rep 5: 4, 387–405 (1987).
John, MC, Amasino, RM: Extensive changes in DNA methylation patterns accompany activation of a silent T-DNA ipt gene in Agrobacterium tumefaciens transformed plant cells. Mol Cell Biol 9: 4298–4303 (1989).
Letham, DS, Palni, LMS: The biosynthesis and metabolism of cytokinins. Ann Rev Plant Physiol 34: 163–197 (1983).
Maniatis, T, Fritsch, EF, Sambrook, J: Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1982).
Matthysse, AG, Scott, TK: Functions of hormones at the whole plant level of organization. In: Scott, TK (ed), Hormonal Regulation of Development II. Encyclopedia of Plant Physiology vol 10, pp. 217–243 Springer-Verlag, Berlin (1984).
McKendree, WC, Paul, AL, Delisle, AJ, Ferl, RJ: In vivo and in vitro characterization of protein interactions with the Dyad G-box of the Arabidopsis Agh gene. Plant Cell 2: 207–214 (1990).
Mouras, A, Negrutiu, I, Dessaux, Y: Phenotypic and genetic variations in crown-gall tumour cells of tobacco. Theor Appl Genet 74: 253–260 (1987).
Nester, EW, Gordon, MP, Amasino, RM, Yanofsky, MF: Crown gall: a molecular and physiological analysis. Ann Rev Plant Physiol 35: 387–413 (1984).
Nissen, P: Dose responses of cytokinins. Physiol Plant 74: 450–456 (1988).
Ooms, G, Hooykaas, PTT, van, Veen, RJM, van, Beelen, P, Regensberg, TJG, Schilperoort, RA: Octopine Ti-plasmid deletion mutants of Agrobacterium tumefaciens with emphasis on the right side of the Ti-region. Plasmid 7: 15–29 (1982).
Ooms, G, Karp, A, Roberts, J: From tumour to tuber: tumour cell characteristics and chromosome numbers of crown gall-derived tetraploid potato plants (Solanum tuberosum cv. Maris Bard). Theor Appl Genet 66: 169–172 (1983).
Ooms, G, Lenton, JR: T-DNA genes to study plant development: precocious tuberization and enhanced cytokinins in A. tumefaciens transformed potato. Plant Mol Biol 5: 205–213 (1985).
Ooms, G, Risiott, R, Kendall, A, Keys, A, Lawlor, D, Smith, S, Turner, J, Young, A: Phenotypic changes in T-cyt-transformed potato plants are consistent with enhanced sensitivity of specific cell types to normal regulation by root-derived cytokinin. Plant Mol Biol 17: 727–743 (1991).
Peerbolte, R, Leenhouts, K, Hooykaas-van Slogteren, GMS, Wullems, GJ, Schilperoort, RA: Clones from a shooty tobacco crown gall tumour II. Irregular T-DNA structures and organization, T-DNA methylation and conditional expression of genes. Plant Mol Biol 7: 285–300 (1986).
Sanders, PR, Winter, JA, Barnason, AR, Rogers, SG, Fraley, RT: Comparison of cauliflower mosaic virus 35S and nopaline synthase promoters in transgenic plants. Nucl Acids Res 15: 1543–1558 (1987).
Tobin, E, Silverthorne, J: Light regulation of gene expression in higher plants. Ann Rev Plant Physiol 36: 569–593 (1985).
Trewavas, A: Is plant development regulated by changes in the concentration of growth substances or by changes in the sensitivity to growth substances? Sensitivity is the limiting factor. Trends Biochem Sci 8: 354–357 (1983).
Turgeon, R, Wood, HN, Braun, AC: Studies on the recovery of crown gall tumour cells. Proc Natl Acad Sci USA 73: 3562–3564 (1976).
Twell, D, Ooms, G: The 5′ flanking DNA of a patatin gene directs tuber specific expression of a chimaeric gene in potato. Plant Mol Biol 9: 365–375 (1987).
van den, Elzen, PJM, Lee, KY, Towsend, J, Bedbrook, JR: Simple binary vectors for DNa transfer to plant cells. Plant Mol Biol 5: 149–154 (1985).
Wullems, GJ, Molendijk, L, Ooms, G, Schilperoort, RA: Retention of tumour markers in F1 progeny plants from in vitro induced octopine and nopaline tumour tissues. Cell 24: 719–727 (1981).
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Dymock, D., Risiott, R., de Pater, S. et al. Regulation of Agrobacterium tumefaciens T-cyt gene expression in leaves of transgenic potato (Solanum tuberosum L. cv. Désirée) is strongly influenced by plant culture conditions. Plant Mol Biol 17, 711–725 (1991). https://doi.org/10.1007/BF00037056
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00037056