Abstract
The expression of the tobacco (Nicotiana tabacum) retrotransposon Tntl has previously been shown to be strongly regulated and driven from the 5′ long terminal repeat (LTR). We report here that the Tntl LTR can promote activity of the β-glucuronidase (GUS) reporter gene in two heterologous species of the Brassicaceae family, namely rapessed (Brassica napus) and Arabidopsis thaliana. The translational LTR-GUS fusion was active in transient expression studies performed with tobacco and rapeseed protoplasts, indicating that the LTR sequences are recognized in heterologous species. Our results also showed that Tntl LTR-promoted GUS expression in transgenic Arabidopsis is strongly regulated, and that, in contrast to tobacco, hormonal activation plays a significant role in the expression of the Tntl LTR in Arabidopsis. LTR sequences were shown to be more effective than the CaMV 35S enhancer region in transient expression studies performed with tobacco or rapessed protoplasts; and substitution of the LTR sequences upstream from the major transcriptional start with the CaMV 35S enhancer region gave high levels of expression in transgenic tobacco and Arabidopsis leaves, suggesting that a Tntl element with similar substitutions in its 5′ LTR might be suited for gene-tagging experiments in heterologous species.
Similar content being viewed by others
References
Balcells L, Swinburne J, Coupland G: Transposons as tools for the isolation of plant genes. Trends Biotechnol 9: 31–36 (1991).
Bevan M: Binary Agrobacterium vectors for plant transformation. Nucl Acids Res 12: 8711–8721 (1984).
Boeke JD, Corces VG: Transcription and reverse transcription of retrotransposons. Annu Rev Microb 43: 403–434 (1989).
Boeke JD, Garfinkel DJ, Styles CA, Fink GR: Ty elements transpose through an RNA intermediate. Cell 40: 491–500 (1985).
Bourgin JP, Missonier C: Vegetative propagation and cold preservation of haploid plants of Nicotiana tabacum and Nicotiana paniculata. Haploid Inf Serv 8: 7 (1973).
Bourgin JP, Chupeau Y, Missonier C: Plant regeneration from mesophyll protoplasts of several Nicotiana species. Physiol Plant 45: 288–292 (1979).
Bradford MM: A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72: 248–254 (1976).
Carozzi NB, Warren GW, Desai N, Jayne SM, Lotstein R, Rice DA, Evola S, Koziel MG: Expression of a chimeric CaMV 35S Bacillus thuringiensis insecticidal protein gene in transgenic tobacco. Plant Mol Biol 20: 539–548 (1992).
Casacuberta JM, Grandbastien MA: Characterisation of LTR sequences involved in the protoplast specific expression of the tobacco Tntl retrotransposon. Nucl Acids Res 21: 2087–2093 (1993).
Chupeau Y, Bourgin JP, Missonier C, Dorion N, Morel G: Préparation et culture de protoplastes de divers Nicotiana. CR Acad Sci Paris (D) 278: 1565–1568 (1974).
Flavell AJ, Dunbar E, Anderson R, Pearce SR, Hartley R, Kumar A: Tyl-copia group retrotransposons are ubiquitous and heterogeneous in higher plants. Nucl Acids Res 20: 3639–3644 (1992).
Grandbastien MA, Spielmann A, Caboche M: Tntl, a mobile retroviral-like element of tobacco isolated by plant cell genetics. Nature 337: 376–380 (1989).
Grandbastien MA: Retroelements in higher plants. Trends Genet 8: 103–108 (1992).
Guerche P, Bellini C, LeMoullec JM, Caboche M: Use of a transient expression assay for the optimization of direct gene transfer into tobacco mesophyll protoplasts by electroporation. Biochimie 69: 621–628 (1987).
Guerineau F, Brooks L, Meadows J, Lucy A, Mullineaux P: Sulfonamide resistance gene for plant transformation. Plant Mol Biol 15: 127–136 (1990).
Hirochika H: Activation of tobacco retrotransposons during tissue culture. EMBO J 12: 2521–2528 (1993).
Jefferson RA: Assaying chimeric genes in plants: the GUS gene fusion system. Plant Mol Biol Rep 5: 387–405 (1987).
Jefferson RA, Kavanagh TA, Bevan MW: GUS fusions: β-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J 6: 3901–3907 (1987).
Johns MA, Mottinger J, Freeling M: A low copy number, copia-like transposon in maize EMBO J 4: 1093–1102 (1985).
Maniatis T, Fritsch EF, Sambrook J: Molecular Cloning: A Laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY (1982).
Murashige T, Skoog F: A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15: 473–497 (1962).
Pouteau S, Huttner E, Grandbastien MA, Caboche M: Specific expression of the tobacco Tntl retrotransposon in protoplasts. EMBO J 10: 1911–1918 (1991).
Rogers SG, Horsch RB, Fraley RT: Gene transfer in plants: Production of transformed plants using Ti plasmid vectors. Meth Enzymol 118: 627–640 (1986).
Rouan D: Transfert direct de gènes chez les Brassica: application à la résistance au virus de la mosaïque du chou-fleur. Thèse, Toulouse, France (1991).
Rouan D, Guerche P, Transformation and regeneration of oilseed rape protoplasts. In: Lindsey K (ed) Plant Tissue Culture Manual, B4: 1–24. Kluwer Academic Publishers. Dordrecht, Netherlands (1991).
Smyth DR: Plant retrotransposons. In: Verma DPS (ed) Control of Plant Gene Expression, pp 1–15 CRC Press, Boca Raton, FL (1993).
Sneddon A, Flavell AJ: The transcriptional control regions of the copia retrotransposon. Nucl Acids Res 17: 4125–4035 (1989).
Valvekens D, Van Montagu M, Van Lijsebettens M: Agrobacterium tumefaciens-mediated transformation of Arabidopsis thaliana root explants by using kanamycin selection. Proc Natl Acad Sci USA 85: 5536–5540 (1988).
Voytas DF, Cummings MP, Konieczny A, Ausubel F, Rodermel C: Copia-like retrotransposons are ubiquitous among plants. Proc Natl Acad Sci USA 89: 7124–7128 (1993).
Yanisch-Perron C, Vieira J, Messing J: Improved M13 phage cloning vectors and host strans: Nucleotide sequences of the M13mp18 and pUC19 vectors. Gene 33: 103–119 (1985).
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Pauls, P.K., Kunert, K., Huttner, E. et al. Expression of the tobacco Tnt1 retrotransposon promoter in heterologous species. Plant Mol Biol 26, 393–402 (1994). https://doi.org/10.1007/BF00039548
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00039548