GrainGenes

II.2. The isolation and characterisation of photorespiratory mutants of barley Hordeum vulgare. L. cv. Maris mink

P.J. Lea*, N.P. Hall, A.C. Kendall, A.J. Keys, B.J. Miflin, J.C. Turner and R.M. Wallsgrove. Department of Biochemistry, Rothamsted Experimental Station, Harpenden, Herts AL5 2JQ, U.K. *Department of Biological Sciences, The University of Lancaster, Lancaster, LA1 4Y Q, U.K.

The evolution of C02 in the light exhibited by a range of plants (termed C3) is known as photorespiration (Ogren and Chollet, 1982). The flux of carbon along the photorespiratory pathway appears to be solely regulated by the proportion of C02 to O2 available to the enzyme RUBP carboxylase/oxygenase. In 1979 Somerville and Ogren devised an entirely novel selection procedure (subsequently described in detail, Somerville and Ogren, 1982; Somerville, 1984) which entailed growing mutated plants in air enriched to 1% C02. The plants were then transferred to normal air and any individuals that showed symptoms of stress were returned to 1% C02 and grown on to subsequent generations. Using this system Somerville and his colleagues identified seven different nuclear mutations in Arabidopsis thaliana.

At Rothamsted a similar selection procedure with azide treated M2 barley seeds (Kleinhofs et al., 1978) has been employed. Since 1979, 61 mutant with the air-sensitive phenotype have been selected from over 120,000 M2 seeds. However, only five of the mutants have been positively identified and three subjected to genetic analysis (see Bright et al., 1984, for review). Plants are given the prefix RPr (Rothamsted Photorespiration) followed by the year of isolation and the number in that year.

RPr 79/2. Dicarboxylate Transport dct
The mutant is unable to transport 2-oxoglutarate in to the chloroplast and is similar to that described in A. thaliana (Somerville and Ogren, 1983). In vivo the chloroplasts are unable to convert glutamine to glutamate although they contain normal levels of ferredoxin dependent glutamate synthase (Lea al. 1984).

RPr 79/4. Catalase Cat 2
The mutant plants contain less than 10% of the peroxisomal catalase activity (Kendal et al., 1983; Parker and Lea, 1983) and two major bands of catalase activity are missing when leaf extracts are subject to starch gel electrophoresis. The plants synthesise excess glutathione when exposed to air (Smith et al., 1985). We initially suggested that the mutation was in the Cat 3 gene, but following the data of Tsaftaris et al. (1983) we have redesignated it as the Cat 2 gene.

RPr 82/1 and RPr 82/9. Ferredoxin Dependent Glutamate Synthase gluS
Two separate lines of plants contain less than 5% ferredoxin dependent glutamate synthase activity and no detectable enzyme protein (Lea et al., 1984). Leaves accumulate glutamine, malate and ammonia when they are exposed to air. The mutants are similar to that described by Somerville and Ogren (1980).

All three mutations appear to involve a single recessive nuclear gene. Approximately 25% of the F2 progeny are air-sensitive and there is an exact correlation between this sensitivity and the enzyme deficiency. F1 progeny grow normally in air and for RPr 79/4 and RPr 82/9 contain approximately 50% of the normal catalase and glutamate synthase activities respectively.

Two further mutants; RPr 83/202 unable to convert glycine to serine and RPr 84/90 lacking phosphoglycollate phosphatase have also been identified. Rpr 83/202 is also a dwarf mutant and does not produce flowering heads. Studies on Rpr 84/90 are currently being carried out.

References:

Bright, S.W.J., P.J. Lea, P. Arruda, N.P. Hall, A.C. Kendall, A.J. Keys, J.S.H. Kueh, M.L. Parker, S.E. Rognes, T.C. Turner, R.M. Wallsgrove and B.J. Miflin. 1984. Manipulation of key pathways in photorespiration and amino acid metabolism by mutation and selection. In The Genetic Manipulation of Plants and its Application to Agriculture (eds. P.J. Lea and G.R. Stewart) Oxford University Press, Oxford. pp. 141-169.

Kendall, A.C., A.J. Keys, J.C. Turner, P.J. Lea and B.J. Miflin. 1983. The isolation and characterisation of a catalase deficient mutant of barley (Hordeum vulgare L.). Planta 159:505-511.

Kleinhofs, R.L. Warner, F.J. Muehlbauer and R.A. Nilan. 1978. Induction and selection of specific gene mutations in Hordeum and Pisum. Mutation Research 51:29-35.

Lea, P.J., N.P. Hall, A.C. Kendall, A.J. Keys, B.J. Miflin, J.C. Turner and R.M. Wallsgrove. 1984. The isolation and characterisation of photorespiratory mutants of barley Hordeum vulgare. In Advances in Photosynthesis Research (e d. C. Sybesma), Martinus Nijhoff Dr. W. Junk, The Hague. Vol III, pp. 841-844.

Ogren, W.L. and R. Chollet. 1982. Photorespiration. In Photosynthesis: Development, Carbon Metabolism and Plant Productivity (ed. Govindjee), Academic Press, New York. Vol II, pp. 191-230.

Parker, M.L. and P.J. Lea. 1983. Ultrastructure of the mesophyll cells of leaves of a catalase-deficient mutant of barley (Hordeum vulgare L.) Planta 159:512-517.

Smith I.K., A.C. Kendall, A.J. Keys, J.C. Turner, and P.J. Lea. 1985. Increased levels of glutathione in a catalase deficient mutant of barley (Hordeum vulgare L.). Plant Science (in press).

Somerville, C.R. 1984. The analysis of photosynthetic carbon dioxide fixation and photorespiration by mutant selection. In. Oxford Surveys of Plant Molecular and Cell Biology (ed. B.J. Miflin), Oxford University Press, Oxford. Vol I, pp. 103-131.

Somerville, C.R. and W.L. Ogren. 1980. Inhibition of photosynthesis in Arabidopis mutants lacking leaf glutamate synthase activity. Nature 286:257-259.

Somerville, C.R. and W.L. Ogren. 1982. Genetic modification of photorespiration. Trends. Biochem. Sci. 7:171-174.

Somerville, S.C. and W.L. Ogren. 1983. An Arabidopis thaliana mutant defective in chloroplast dicarboxylate transport. Proc. Natl. Acad. Sci. USA. 80:1290-1294.

Tsaftaris, A.S., A.M. Bosabalidis and J.G. Scandalios. 1983. Cell-type-specific gene expression and acatalasemic peroxisomes in a null Cat 2 catalase mutant of maize. Proc. Natl. Acad. Sci. USA. 80:4455-4459.

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