Expression of uidA gene in tobacco transplastomic plants under the control of 5'-untranslated region of genomic RNA of potato virus Y
DOI:
https://doi.org/10.26577/eb-2019-3-b12Abstract
Regulation of the expression of chloroplast genes mainly occurs at the post-transcriptional level, in which 5'-untranslated regions (5'UTR) of plastid mRNA play a significant role. In transplastomic tobacco plants, we studied the effect of the 5’UTRs of the genomic RNA of potato Y-virus (5’PVY), as well as the 5’UTR of the plastid psbA gene (5’psbA) on the expression of the bacterial reporter uidA gene encoding β-glucuronidase (GUS). Genetic constructs containing the uidA gene were created under the control of identical transcriptional regulatory elements: the promoter and terminator of the psbA gene and two different 5’UTRs — 5’psbA and 5’PVY. Тobacco chloroplasts were transformed with these constructs by bioballistic method. Transplastomic lines showed approximately 24-fold differences in GUS activity. Replacing the natural 5’psbA with the heterologous 5’PVY resulted in a significant decrease in the synthesis of GUS protein.
Key words: chloroplasts, protein synthesis, 5’UTR, transplastomic plants, reporter protein.
References
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28 Yukawa M., Kuroda H., Sugiura M. (2007) A new in vitro translation system for non-radioactive assay from tobacco chloroplasts: effect of pre-mRNA processing on translation in vitro. Plant J, vol. 49, pp. 367–376.
29 Zoschke R., Bock R. (2018) Chloroplast translation: structural and functional organization, operational control and regulation. Plant Cell, vol. 30, pp. 745–770.
2 Bock R. (2007) Plastid biotechnology: prospects for herbicide and insect resistance, metabolic engineering and molecular farming. Curr Opin Biotechnol, vol. 18, pp. 100–106.
3 Fernández-San Millán A., Ortigosa S. M., Hervás-Stubbs S., Corral-Martínez P., Seguí-Simarro J. M., Gaétan J., Veramendi J. (2008) Human papillomavirus L1 protein expressed in tobacco chloroplasts self-assembles into virus-like particles that are highly immunogenic. Plant Biotechnol J, vol. 6, pp. 427–441.
4 Gerasymenko I. M., Sheludko Y. V., Klebanovych A. A., Rudas V. A., Shakhovsky A. M., Klein T. M., Kuchuk N. V. (2016) Comparison of effectiveness of 5′-regulatory sequences in transplastomic tobacco chloroplasts. Transgenic Res, vol. 26, pp. 65–75.
5 Gray B.N., Ahner B.A, Hanson M.R. (2009) Extensive homologous recombination between introduced and native regulatory plastid DNA elements in transplastomic plants. Transgenic Res, vol. 18, pp. 559–572.
6 Hennig A., Bonfig K., Roitsch T., Warzecha H.(2007) Expression of the recombinant bacterial outer surface protein A in tobacco chloroplasts leads to thylakoid localization and loss of photosynthesis. FEBS J, vol. 274, pp. 5749-5758.
7 Herz S., Füßl M., Steiger S., Koop H.U. (2005) Development of novel types of plastid transformation vectors and evaluation of factors controlling expression. Transgenic Res, vol. 14, pp. 969–982.
8 Jefferson R.A., Kavanagh T.A., Bevan M.W. (1987) GUS fusions: beta-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J, vol. 6, pp.3901–3907.
9 Jin S., Daniell H. (2015) The Engineered Chloroplast Genome Just Got Smarter. Trends Plant Sci, vol. 20, pp. 622–640.
10 Jin S., Kanagaraj A., Verma D., Lange T., Daniell H. (2011) Release of hormones from conjugates: chloroplast expression of β-glucosidase results in elevated phytohormone levels associated with significant increase in biomass and protection from aphids or whiteflies conferred by sucrose esters. Plant physiol, vol. 155, pp. 222–235.
11 Kuroda H., Maliga P. (2001) Complementarity of the 16S rRNA penultimate stem with sequences downstream of the AUG destabilizes the plastid mRNAs. Nucleic Acids Res, vol. 29, pp. 970–975.
12 Kuroda H., Maliga P. (2002) Overexpression of the clpP 5'-untranslated region in a chimeric context causes a mutant phenotype, suggesting competition for a clpP-specific RNA maturation factor in tobacco chloroplasts. Plant physiol, vol. 129, pp. 1600-1606.
13 Leelavathi S., Reddy V.S. (2003) Chloroplast expression of His-tagged GUS-fusions: a general strategy to overproduce and purify foreign proteins using transplastomic plants as bioreactors. Mol Breed, vol. 11, pp. 49–58.
14 Michoux F., Ahmad N., Hennig A., Nixon P.J., Warzecha H. (2003) Production of leafy biomass using temporary immersionbioreactors: an alternative platform to express proteinsin transplastomic plants with drastic phenotypes. Planta, vol. 237, pp. 903–908.
15 Morgenfeld M., Lentz E., Segretin M. E., Alfano E. F., Bravo-Almonacid F. (2014) Translational Fusion and Redirection to Thylakoid Lumen as Strategies to Enhance Accumulation of Human Papillomavirus E7 Antigen in Tobacco Chloroplasts. Mol Biotechnol, vol. 56, pp. 1021–1031.
16 Murashige T, Skoog F. (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant, vol. 15, pp.473–497.
17 Nickelsen J., Fleischmann M., Boudreau E., Rahire M., Rochaix J.D. (1999) Identification of cis-acting RNA leader elements required for chloroplast psbD gene expression in Chlamydomonas. Plant Cell, vol. 11, pp. 957–970.
18 Nizkorodova A., Beisenov D., Iskakov B. (2015) 5’-untranslated region of potato virus Y genomic RNA provides efficient mRNA translation in bacterial cells. EMBO Conference: protein synthesis and translational control, Heidelberg, 09-13 Sep. 2015, p. 243.
19 Oey M., Lohse M., Kreikemeyer B., Bock R. (2009) Exhaustion of the chloroplast protein synthesis capacity by massive expression of a highly stable protein antibiotic. Plant J, vol. 57, pp. 436–445.
20 Ruf S., Forner J., Hasse C., Kroop X., Seeger S., Schollbach L., Bock R. (2019) High-efficiency generation of fertile transplastomic Arabidopsis plants. Nat Plants, vol. 5, pp. 282–289.
21 Ruhlman T., Ahangari R., Devine A., Samsam M., Daniell H. (2007) Expression of cholera toxin B-proinsulin fusion protein in lettuce and tobacco chloroplasts: oral administration protects against development of insulitis in non-obese diabetic mice. Plant Biotechnol J, vol. 5, pp. 495–510.
22 Sambrook, J. and Russell, D.W. (2001) Molecular cloning a laboratory manual, 3rd ed. Cold Spring Harbor Laboratory Press, New York.
23 Scotti N., Cardi T. (2014) Transgene-induced pleiotropic effects in transplastomic plants. Biotechnol Lett, vol. 36, pp. 229–239.
24 Svab Z., Hajdukiewicz P., Maliga P. (1990) Stable transformation of plastids in higher plants. Proc Natl Acad Sci USA, vol. 87, pp. 8526–8530.
25 Tiller N., Bock R. (2014) The Translational Apparatus of Plastids and Its Role in Plant Development. Mol Plant, vol. 7, pp. 1105–1120.
26 Yang H., Gray B. N., Ahner B. A., Maureen R., Hanson M.R. (2013) Bacteriophage 5’ untranslated regions for control of plastid transgene expression. Planta, vol. 237, pp. 517–527.
27 Ye G. N, Hajdukiewicz P.T., Broyles D., Rodriguez D., Xu C.W., Nehra N., Staub J.M. (2001) Plastid-expressed 5-enolpyruvylshikimate-3-phosphate synthase genes provide high level glyphosate tolerance in tobacco. Plant J, vol. 25, pp.261–270.
28 Yukawa M., Kuroda H., Sugiura M. (2007) A new in vitro translation system for non-radioactive assay from tobacco chloroplasts: effect of pre-mRNA processing on translation in vitro. Plant J, vol. 49, pp. 367–376.
29 Zoschke R., Bock R. (2018) Chloroplast translation: structural and functional organization, operational control and regulation. Plant Cell, vol. 30, pp. 745–770.
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How to Cite
Stanbekova, G. E., Beisenov, D. K., Kalieva, A. A., & Iskakov, B. K. (2020). Expression of uidA gene in tobacco transplastomic plants under the control of 5’-untranslated region of genomic RNA of potato virus Y. Experimental Biology, 80(3), 126–135. https://doi.org/10.26577/eb-2019-3-b12
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МOLECULAR BIOLOGY AND GENETICS
