Micrografting Provides Evidence for Systemic Regulation of Sulfur Metabolism between Shoot and Root

Abstract

The uptake of sulfate by roots and its reductive assimilation mainly in the leaves are not only essential for plant growth and development but also for defense responses against biotic and abiotic stresses. The latter functions result in stimulus-induced fluctuations of sulfur demand at the cellular level. However, the maintenance and acclimation of sulfur homeostasis at local and systemic levels is not fully understood. Previous research mostly focused on signaling in response to external sulfate supply to roots. Here we apply micrografting of Arabidopsis wildtype knock-down sir1-1 mutant plants that suffer from an internally lowered reductive sulfur assimilation and a concomitant slow growth phenotype. Homografts of wildtype and sir1-1 confirm the hallmarks of non-grafted sir1-1 mutants, displaying substantial induction of sulfate transporter genes in roots and sulfate accumulation in shoots. Heterografts of wildtype scions and sir1-1 rootstocks and vice versa, respectively, demonstrate a dominant role of the shoot over the root with respect to sulfur-related gene expression, sulfate accumulation and organic sulfur metabolites, including the regulatory compound O-acetylserine. The results provide evidence for demand-driven control of the shoot over the sulfate uptake system of roots under sulfur-sufficient conditions, allowing sulfur uptake and transport to the shoot for dynamic responses.

Keywords: O-acetylserine; grafting; organ communication; sulfate transporter; sulfite reductase; sulfur homeostasis.

Figure 1

Phenotypic analysis of micrografted plants. (a,b) Phenotype of grafted plants five weeks after germination; selected representatives are shown for the four different grafting combinations. (c,d) Fresh weight (FW) of roots and shoots. Data represent mean and standard error and are modified according to [11] (n > 15). Different letters indicate statistically significant differences; p < 0.05, ANOVA followed by Dunn´s test; FW = fresh weight.

Figure 2

Steady-state transcript levels of key genes in the sulfur-assimilation pathway in micrografted plants. Transcript levels were determined by qRT-PCR using TIP41-like (At4g34270) as the reference gene in the roots (a,c,e,f) and shoots (b,d) of grafted plants. The level in control roots and shoots was set to 1, and the other values are expressed as x-fold of control. Data are shown as means ± SE (n = 4). Different letters indicate individual groups identified by multiple pairwise comparisons with a Holm–Sidak, one-way ANOVA (p, 0.05) followed by the Student–Newman–Keuls posthoc test.

Figure 3

Steady-state levels of macronutrients in the roots and shoots of micrografted plants. Sulfate (a,b), phosphate (c,d) and nitrate (e,f) concentrations in roots (a,c,e) and shoots (b,d,f) of micrografted Arabidopsis plants. Data represent the means ± SE (n = 4). Different letters indicate individual groups identified by multiple pairwise comparisons with a Holm–Sidak, one-way ANOVA (p, 0.05) followed by the Student–Newman–Keuls posthoc test.

Figure 4

Steady-state levels of OAS and thiols in micrografted plants. Concentrations of OAS (a,b), Cys (c,d) and GSH (e,f) in roots (a,c,e) and shoots (b,d,f). Data represent the means ± SE (n = 4). Different letters indicate individual groups identified by multiple pairwise comparisons with a Holm–Sidak, one-way ANOVA (p, 0.05) followed by the Student–Newman–Keuls posthoc test.