NCED OE rootstocks have reduced gene expression for ABA receptors and signalling components
Rootstock SlNCED1 overexpression (Figure 6a) was consistent with transgene expression level in own-rooted plants (Thompson et al.2007b; Martínez-Andújar et al. 2020b), implying that shoot-to-root signalling has little effect on constitutive (root-specific in grafted plants) SlNCED expression. Although bulk root ABA status did not increase in the adult plants (Figure 3a), previously ABA in root exudates from approximately 7 week old detopped plants (Thompson et al . 2007a), in root cultures (Thompsonet al . 2007b) and in bulk root tissue and xylem sap of younger ungrafted plants (Martínez-Andújar et al . 2020b) was elevated; in addition, in grafted plants the bulk root ABA was determined by the root genotype and was elevated in SP5 and SP12 (Thompson et al.2007b). Therefore, the lack of bulk root ABA accumulation in this study is consistent with increased export (Figure S1) and catabolism of ABA (Figure 3b).
Many genes were down- or up-regulated in NCED OE rootstocks compared to the WT grafts (Figure 5). Amongst those genes, 7 PYL ABA receptors and 3 WRKY factors were downregulated in NCED OE roots, suggesting decreased sensitivity to ABA, as in own-rooted plants grown in optimal conditions (Martínez-Andújar et al. 2020b). Several ABA PYR/PYL receptors are highly expressed in tomato roots compared to other tissues (González-Guzmán et al., 2014), allowing root system adaptation to low water potential including via modulation of osmoregulation and architectural changes (Sharp et al. 2004; Des Marais et al. 2012; Duan et al. 2013). For example,PYL8 plays an essential role in regulating root ABA sensitivity in Arabidopsis (Antoni et al. 2013), promoting lateral root growth by enhancing MYB77 -dependent transcription of auxin-responsive genes (Zhao et al. 2014). Loss-of-function of several pyr/pyl loci impaired ABA signalling, causing a robust ABA-insensitive root phenotype (Park et al. 2009; González-Guzmán et al.2014). Thus, downregulation of PYL s in NCED OE rootstocks may account for their limited root system development and sensitivity to saline stress.
Despite the proposed decreased ABA sensitivity, genes involved in ABA biosynthesis (FLC/AAO, Solyc07g066480) and signalling (AREB , Solyc04g078840; ATHB12, Solyc01g096320) were only slightly induced or not affected in either SP rootstock compared to WT. Furthermore, transcriptomic and RT-qPCR data of stress-related genes (TAS14 , Solyc02g084850; KIN2 , Solyc03g095510; LEA,Solyc03g116390 and some MYB s) indicate SP12 was more responsive than WT rootstocks to low saline stress, with an attenuated response in SP5. These different transcriptomic responses between SP rootstocks mimicked the responses of own-rooted plants grown under optimal conditions (Martínez-Andújar et al. 2020b), implying that each SP rootstock senses a different stress intensity that optimizes physiological responses according to both basal and NCED OE ABA levels.