Salt treatment inhibits SLs production by reducing the expression of genes related to SLs biosynthesis
To further explore the regulation mechanism of tillering process in response to salinity, the contents of strigolactones (SLs) in S. alterniflora roots were analyzed. In the present study, the contents of two derivatives of SLs including strigol and 5-deoxystrigol (5-DS) were measured using the HPLC-MS/MS. It was surprised that 5-DS was only detected in 0‰ salinity treatment and strigol was only detected in 15‰ salinity treatment, while none of them could be detected in 30‰ salinity treatment (Table 1). 5-DS is the precursor of strigol and both of them are the main natural compounds in plants (Ćavar et al. , 2015). Sanja et al. (2015) summarized that the structure of SLs in different plants or even in different varieties within species were different. Recently, the effects of abiotic stresses on SLs production in plants were widely investigated (Aroca et al. , 2013; Ruiz-Lozano et al. , 2016; Sharifi & Bidabadi, 2020). In lettuce, the SLs production assessed by germination bioassay was significantly reduced under salt stress (Aroca et al. , 2013). Similarly, the SLs production in tomato was obviously inhibited under drought stress, suggesting a negative relationship between stress and SLs production (Ruiz-Lozano et al. , 2016). Moreover, the application of GR24, a synthetic SL, reduced the tillering in rice, suggesting a negative relationship between SLs content and tiller number (Lin et al ., 2009; Tamiru et al ., 2014). Despite of different structure of SLs, the SLs content in 0‰ salinity treatment (5-DS: 0.1416 ± 0.0225 ng/g) was higher than that in 15‰ salinity treatment (strigol: 0.0632 ±0.0044 ng/g) and 30‰ salinity treatment (Table 1). This finding was consistent with previous studies and demonstrated that salt treatment would inhibit the production of SLs in S. alterniflora .
Furthermore, two genes involved in SLs biosynthesis (SaD10 ,SaD17 ) were amplified and sequenced (Supplementary Table S2, Supplementary Table S3). The expression pattern of SaD10 inS. alterniflora (Figure 5 a) was similar with Arabidopsis thaliana in which the highest expression levels of D10 was observed in root tissue (Auldridge et al. , 2006). For D17 , the highest levels of CCD7 /MAX3/HTD1/D17 was also found inArabidopsis root (Booker et al. , 2004). Moreover, Zouet al. (2006) used the binary vector (Pro HTD1:GUS) to examine the distribution ofHTD1 in different tissues and found that the intensity of GUS expression at node was higher than the other tissues. In the present study, the expression pattern of SaD17 in S. alterniflorawas basically consistent with previous studies that SaD17 was mainly expressed in node and root. Based on these results, the roots ofS. alterniflora were sampled for further SLs signaling related gene expression analysis.
As the key genes encoded enzymes involved in SLs biosynthesis,CCD7 /D17 and CCD8 /D10 were also found to be decreased under various abiotic stresses. For examples, drought stress profoundly reduced the SLs content and the transcriptional level ofSlCCD7 in tomato (Ruiz-Lozano et al. , 2016). Similarly, the SLs production and transcriptional levels of LjCCD7 andLjCCD8 were also decreased in lotus under osmotic stress (Liuet al. , 2015). In the present study, the transcriptional levels of SaD17 and SaD10 were decreased with the increase of salinity (Figure 6a, b), which were consistent with the measurement results of SLs content (Table 1). These results implied that soil salinity would reduce the SLs production by down-regulating the expression of SaD17 and SaD10 in S. alternifloraroots.