Innovation in life sciences

Evaluation and Licensing Opportunities For further information on this technology and evaluation / licensing opportunities please contact: Dr Jan Chojecki ajsc@pbltechnology.com Tel: +44 (0)1603 456500 Fax: +44 (0)1603 456552 Tech ID: 01.266

Stress Resistant Plants

Expression of a cyanobacterial flavodoxin in plants results in enhanced stress resistance and increased yield in field trials

Researchers at IBR (Universidad Nacional de Rosario and CONICET, Argentina), in collaboration with scientist at the Universidad Nacional de Zaragoza (Spain) have expressed a bacterial flavodoxin in chloroplasts that results in enhanced tolerance Patent Literature to the toxic effects of reactive oxygen species generated by multiple sources of US Patent No. 6,781,034 environmental stress. Transgenic tobacco plants containing the flavodoxin show Australian Patent No. 2002334130 tolerance to methyl viologen (paraquat), UV-B radiation, extreme temperatures, European Patent No. EP1442127 drought stress and irradiation. Necrotic injury caused by bacterial, viral and fungal infections is also considerably reduced when compared to wild type plants. Field trials have now demonstrated the potential of this technology. Transgenic plants expressing the flavodoxin in the plastids, especially the chloroplasts were compared with those expressing in the cytosol and with non-transformed wild type. Despite unseasonally mild conditions, resulting in a greater survival of wild type than expected, the transformants, especially those expressing the flavodoxin in the plastids showed significantly higher dry weight yield and survival than wild type.

Genotype
Wild Type Plastid (pfld5-8) Cytosol (cfld1-4)

n 114 131 114

Ave. Dry Weight (g/m2) 9.6 15.5 10.8

Median Dry Weight (g/m2) 9.6 15.6 10.9

SD 1.1 1.3 1.6

% Survival 72 87 77

Analysis of variance showed pfld5-8 differed from wt and cfld1-4 with p<0.001, cfld1-4 differed from wt with p<0.01

TEC Release: October 2002

Updated December 2009

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Innovation in life sciences

New work by the inventors and co-workers has now been published in The Plant Cell (Tognetti et al, August 2006). This paper characterizes how the bacterial flavodoxin compensates for deficiencies on photoelectron transport that occur in various stress situations. Chloroplast ferredoxin plays a pivotal role in plant cell metabolism, delivering reducing equivalents to various essential oxidoreductive pathways, however its levels decline in stress conditions. The Plant Cell paper shows that the cyanobacterial flavodoxin can mediate plant ferredoxin-dependent reactions both in vitro and, using transgenic tobacco, in-vivo. The transgenic plants showed increased tolerance to multiple sources of stress including : redox-cycling herbicides extreme temperatures high irradiation water deficit UV radiation

Increased Tolerance of Transgenic Tobacco Plants Expressing Bacterial Flavodoxin in Chloroplasts to Different Sources of Environmental Stress. (A) Two-weeks old seedlings were illuminated for 18 h at 500 mol quanta m-2 s-1 and 40oC in MS agar. (B) Seven-weeks old plants were cultured for 20 days at 500 mol quanta m2 -1 s and 9oC in MS agar. (C) Leaves from 2-months old plants (node 8) grown in soil were exposed for 18 h to a focused light beam of 2,000 mol quanta m-2 s-1. (D) Two-months old plants grown in soil were exposed for 20 min to UV-C radiation and photographed after 3 days. (E) Two-months old specimens grown in soil were subjected to a 3-day water deprivation regime. (F) Seven-weeks old plants cultured in hydroponia were exposed to UV-AB radiation for 24 h. Homozygous transformants of the T3 generation (flavodoxin expressed in plastids, pfld, or in the cytosol, cfld) and their wild-type (WT) siblings were exposed to the stress treatments as described in Tognetti et al The Plant Cell (2006) The panels illustrate characteristic phenotypes, with the arrows indicating sites of tissue damage (C and D).

References:
Functional Replacement of Ferredoxin by a Cyanobacterial Flavodoxin in Tobacco Confers Broad Range Stress Tolerance. Tognetti V, Palatnik J, Fillat M, Melzer M, Hajirezaei M-R, Valle E and Carillo N. The Plant Cell (2006): 18(8); 2035-2050. Enhanced plant tolerance to iron starvation by functional substitution of chloroplast ferredoxin with a bacterial flavodoxin. Tognetti VB, Zurbriggen MD, Morandi EN, Fillat MF, Valle EM, Hajirezaei M-R and Carrillo N. PNAS (2007): 104 (27); 11495-11500. Stress-inducible flavodoxin from photosynthetic microorganisms. The mystery of flavodoxin loss from the plant genome. Zurbriggen MD,Tognetti VB and Carrillo N. IUBMB Life (2007): 59(4-5):355-60. Detoxification of 2,4-dinitrotoluene by Transgenic Tobacco Plants Expressing a Bacterial Flavodoxin. Tognetti VB, Monti MR, Valle EM, Carrillo N and Smania AM. Environmental Science and Technology (2007): 41 (11); 4071-4076. Chloroplast-generated reactive oxygen species play a major role in localized cell death during the non-host interaction between tobacco and Xanthomonas campestris pv. vesicatoria. Zurbriggen MD, Carrillo N, Tognetti VB, Melzer M, Peisker M, Hause B and Hajirezaei M-R. Plant J (2009): 60 (6); 962 - 973.

TEC Release: October 2002

Updated December 2009

www.pbltechnology.com