Ectopic phytocystatin expression leads to enhanced drought stress tolerance in soybean (Glycine max) and Arabidopsis thaliana through effects on strigolactone pathways and can also result in improved seed traits

Abstract

SummaryEctopic cystatin expression has long been used in plant pest management, but the cysteine protease, targets of these inhibitors, might also have important functions in the control of plant lifespan and stress tolerance that remain poorly characterized. We therefore characterized the effects of expression of the rice cystatin, oryzacystatin-I (OCI), on the growth, development and stress tolerance of crop (soybean) and model (Arabidopsis thaliana) plants. Ectopic OCI expression in soybean enhanced shoot branching and leaf chlorophyll accumulation at later stages of vegetative development and enhanced seed protein contents and decreased the abundance of mRNAs encoding strigolactone synthesis enzymes. The OCI-expressing A. thaliana showed a slow-growth phenotype, with increased leaf numbers and enhanced shoot branching at flowering. The OCI-dependent inhibition of cysteine proteases enhanced drought tolerance in soybean and A. thaliana, photosynthetic CO 2 assimilation being much less sensitive to droughtinduced inhibition in the OCI-expressing soybean lines. Ectopic OCI expression or treatment with the cysteine protease inhibitor E64 increased lateral root densities in A. thaliana. E64 treatment also increased lateral root densities in the max2-1 mutants that are defective in strigolactone signalling, but not in the max3-9 mutants that are defective in strigolactone synthesis. Taken together, these data provide evidence that OCI-inhibited cysteine proteases participate in the control of growth and stress tolerance through effects on strigolactones. We conclude that cysteine proteases are important targets for manipulation of plant growth, development and stress tolerance, and also seed quality traits.

Ectopic phytocystatin expression increases nodule numbers and influences the responses of soybean (Glycine max) to nitrogen deficiency

“…The variation in the levels of expression between different independent transgenic lines is consistent with the known features of 35S-driven transgene expression in A. thaliana, in which a bimodal expression pattern has been reported consisting of 20% high-level expressers and 80% lowlevel expressers (De Bolle et al, 2003). We compared the effects of ectopic OCI expression in the responses of soybeans to LN using these transgenic lines (Quain et al, 2014). Our earlier studies had indicated that OCI-1 line had about half the levels of the OCI protein in their leaves than the OCI-2 line (Quain et al, 2014).…”

“…Our aim was to explore whether the levels of transcripts encoding these proteins was influenced by ectopic OCI expression in soybean plants that had been grown for four weeks with either a high (5 mM) nitrate (HN) supply or in the absence of added nitrate (LN). We selected two independent transgenic OCI expressing soybean lines (OCI-1 and OCI-2) that had different levels of OCI transcripts in the leaves (Quain et al, 2014). The variation in the levels of expression between different independent transgenic lines is consistent with the known features of 35S-driven transgene expression in A. thaliana, in which a bimodal expression pattern has been reported consisting of 20% high-level expressers and 80% lowlevel expressers (De Bolle et al, 2003).…”

“…Soybean (Glycine max cultivar Williams) transformation was performed as described previously (Quain et al, 2014). Two independent transformed soybean lines (OCI-1 and OCI-2) that had been produced by selfing the primary transformants twice were used in this study.…”

“…The coordinated expression of transcripts encoding cysteine protease and cystatin interacting partners has been reported in senescent spinach leaves (Tajima et al, 2011). Ectopic OCI expression not only altered the growth and development of tobacco (Van der Vyver et al, 2003;Prins et al, 2008), soybean and Arabidopsis thaliana plants (Quain et al, 2014) but also enhanced tolerance to abiotic stresses, such as low temperatures and drought (Prins et al, 2008;Quain et al, 2014).…”

“…We compared the effects of ectopic OCI expression in the responses of soybeans to LN using these transgenic lines (Quain et al, 2014). Our earlier studies had indicated that OCI-1 line had about half the levels of the OCI protein in their leaves than the OCI-2 line (Quain et al, 2014). The data presented here show that growth under LN condi-tions induced changes in the levels of transcripts encoding cysteine proteases, VPEs and cystatins, and that the LN-dependent responses of the soybean plants are modified in the OCI-1 and OCI-2 lines.…”