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Cloning, characterization and expression of a chloroplastic fructose-1,6-bisphosphatase from Porteresia coarctata conferring salt-tolerance in transgenic tobacco

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Abstract

A gene coding for the chloroplastic fructose 1,6-bisphosphatase (PcCFR) in Porteresia coarctata Tateoka (Roxb.), a halophytic wild rice, has been isolated along with its rice (Oryza sativa; var. indica) homologue (OsCFR), cloned and sequenced. Comparison between the nucleotide and deduced amino acid sequences of these two revealed a difference in five amino acid residues, namely Glu14, Thr24, Ala48, Ala163 and Arg296 in OsCFR which have been found to be replaced by Ser14, Ile24, Ser48, Ser163 and Lys296 in PcCFR respectively. The purified recombinant PcCFR is found to retain its enzymatic activity in presence of up to 500 mM NaCl in vitro as opposed to OsCFR, which is inactivated even at lower salt concentration. The six in vitro point mutant proteins of PcCFR showed varied degree of sensitivity towards high salt, with the maximum OsCFR-like effect in the triple mutant S14A-S48A-S163A suggesting a possible concerted role of all three serine residues in the in vitro salt tolerance property of PcCFR protein. Transgenic tobacco plants with chloroplast targeted PcCFR and OsCFR gene(s) have been developed under constitutive expression of CaMV 35S promoter and NOS terminator. The PcCFR transgenics showed better plant growth during exposure to salt stress in comparison to either the OsCFR or the empty vector transformed plants. The PcCFR transgenics also revealed enhanced photosynthetic efficiency coupled with protection to both photodamage of PSII and chlorophyll degradation through better reactive oxygen species scavenging at higher concentration of NaCl during late salt-stress growth.

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Acknowledgments

The PcCFR gene was isolated during a short visit (ALM) to the laboratory of Dr. Autar Mattoo at USDA, Beltesville, USA. ALM is a Raja Ramanna Fellow of the Department of Atomic Energy, Government of India. This work has been supported by research Grants (to ALM) from the Department of Biotechnology and the Department of Atomic Energy, Government of India.

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  1. Division of Plant Biology, Bose Institute, Centenary Campus, P-1/12 CIT Scheme VII M, Kolkata, 700054, India

    Jolly Chatterjee, Barunava Patra, Rajeswari Mukherjee, Papri Basak, Sritama Mukherjee, Sudipta Ray, Sanghamitra Bhattacharyya, Susmita Maitra, Krishnarup GhoshDastidar, Shilpi Ghosh, Sonali Sengupta & Arun Lahiri Majumder

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Correspondence to Arun Lahiri Majumder.

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Chatterjee, J., Patra, B., Mukherjee, R. et al. Cloning, characterization and expression of a chloroplastic fructose-1,6-bisphosphatase from Porteresia coarctata conferring salt-tolerance in transgenic tobacco. Plant Cell Tiss Organ Cult 114, 395–409 (2013). https://doi.org/10.1007/s11240-013-0334-y

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