Abstract
Functionally distinct Arabidopsis (Arabidopsis thaliana) genes that positively affect root or shoot growth when ectopically expressed were combined to explore the feasibility of enhanced biomass production. Enhanced root growth resulting from cytokinin deficiency was obtained by overexpressing CYTOKININ OXIDASE/DEHYDROGENASE3 (CKX3) under the control of the root-specific PYK10 promoter. Plants harboring the PYK10-CKX3 construct were crossed with four different transgenic lines showing enhanced leaf growth. For all combinations, the phenotypic traits of the individual lines could be combined, resulting in an overall growth increase. Unexpectedly, three out of four combinations had more than additive effects. Both leaf and root growth were synergistically enhanced in plants ectopically expressing CKX3 and BRASSINOSTEROID INSENSITIVE1, indicating cross talk between cytokinins and brassinosteroids. In agreement, treatment of PYK10-CKX3 plants with brassinolide resulted in a dramatic increase in lateral root growth that could not be observed in wild-type plants. Coexpression of CKX3 and the GROWTH-REGULATING FACTOR5 (GRF5) antagonized the effects of GRF5 overexpression, revealing an interplay between cytokinins and GRF5 during leaf cell proliferation. The combined overexpression of CKX3 and GIBBERELLIN 20-OXIDASE1 led to a synergistic increase in leaf growth, suggesting an antagonistic growth control by cytokinins and gibberellins. Only additive effects on root and shoot growth were visible in plants ectopically expressing both CKX3 and ARABIDOPSIS VACUOLAR PYROPHOSPHATASE1, hinting at an independent action mode. Our results show new interactions and contribute to the molecular and physiological understanding of biomass production at the whole plant level.
Highlights
Introduction of pBRIBRASSINOSTEROID INSENSITIVE1 (BRI1) into P10-CYTOKININ OXIDASE/DEHYDROGENASE3 (CKX3) Plants Results in a More Than Additive Increase of Root and Shoot GrowthEctopic expression of BRI1 under the control of its own promoter leads to an increased response to brassinosteroids, resulting in longer petioles and larger leaves (Wang et al, 2001)
Detailed observation of leaf parameters showed that this was mainly attributed to an increase in leaf length that was comparable to that of Col-03BRI1 (Supplemental Fig. S2, B and D), suggesting that ectopic expression of BRI1 overcomes the effects of P10-CKX3 in the leaf length direction
Rosette growth catches up with wild-type plants after bolting when plants are grown in soil (Werner et al, 2010), and expression of P10-CKX3 has no significant influence on the leaf phenotypes obtained by simultaneous ectopic expression of BRI1, GROWTH-REGULATING FACTOR5 (GRF5), GA20ox1, or ARABIDOPSIS VACUOLAR PYROPHOSPHATASE1 (AVP1) after bolting
Summary
Introduction of pBRIBRI1 into P10-CKX3 Plants Results in a More Than Additive Increase of Root and Shoot GrowthEctopic expression of BRI1 under the control of its own promoter leads to an increased response to brassinosteroids, resulting in longer petioles and larger leaves (Wang et al, 2001). When the plants were grown under in vitro conditions for 21 d, enhanced BRI1 expression did not result in a significant increase in either rosette fresh weight (Fig. 2A) or blade area for most leaves (Fig. 2B) but mainly in increased blade length (Supplemental Fig. S2, A and C). Detailed observation of leaf parameters showed that this was mainly attributed to an increase in leaf length that was comparable to that of Col-03BRI1 (Supplemental Fig. S2, B and D), suggesting that ectopic expression of BRI1 overcomes the effects of P10-CKX3 in the leaf length direction. Relative transgene expression levels in shoots of BRI13P10-CKX3 plants were similar to those of the heterozygous parents (Supplemental Fig. S3A), showing that the observed synergism in leaf blade area is not a consequence of an altered expression level of one transgene due to the presence of the other
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