Despite multiple research have established physiological relation between salt stress and CK (Yang and Guo 2018), and several non-cytokinin sensing histidine kinases (AHK1, AHK5 and CKI1) have been attributed to osmosensing (Urao et al. 1999; Pham et al. 2012; Hofmann et al. 2020), the molecular basis of this process in plants is still unknown. Original research demonstrated the ability to directly sense NaCl for AHK1 expressed in yeast sln1Δ sho1Δ mutant. In such experimental conditions, AHK1-mediated downstream phosphorylation was depended on the NaCl concentration. However, the NaCl sensing mechanism was not identified (Urao et al. 1999). Currently, several salt-sensing receptors are known. For example, MOCA1 (At5g18480) responds to Na+, K+, or Li+ ions via the generation of Ca2+ waves (Jiang et al. 2019). Similarly, FERONIA (At3g51550) receptor-like kinase senses Na+ and responds in a Ca2+-dependent way. However, downstream signalling of this receptor was detected only after several hours after salt application, suggesting the involvement of other sensors and/or signalling pathways for prompt early salt responses (Feng et al. 2018). Therefore, in this section, we analyse relations between plants’ CK metabolism and salt stress (up- and down-regulation of CK signalling, biosynthesis, degradation and transport genes expression, levels of different CK forms and Na+ accumulation) (Table 2).
Table 2
Mutants affected by salt stress
Species | Organ/line | Mutant/Genetic modification | Relation to salt stress | CK levels | Expression of CK-related genes in WT under salt stress | Na+levels/localisation | |
Brassica rapa ssp. pekinensis | | | | | BrHK8, BrRR27 ↔ BrPHP1 ↓ BrHP1 and 5 ↑ BrRR8, BrRR19, and BrRR31↑ | | (Liu et al. 2014) |
Corchorus capsularis and Corchorus olitorius | | | | | Most Hps ↑ 4 RR-B and 4 RR-A, 5 CRE ↓ | | (Yang et al. 2017) |
Solanum lycopersicum cv. Alisa Craig | | | | | ARR-B5, ARR-B6, ARR-B8, ARR-B9, and ARR-B10 ↑ | | (Wang et al. 2020) |
Cicer arietinum | | | | | CarHK1, CarHP1, CarRR12 ↓ CarRR17 ↑ | | (Ahmad et al. 2020) |
Sorghum bicolor | | | | | Most SbHPs ↑ SbRR20, SbRR16, SbRR18, SbRR19, and SbPRR1 ↑ | | (Zameer et al. 2021) |
Sophora alopecuroides | | | | | SaRR-A1, A2, SaCKX 5, SaCRE1-9 ↑ SaRR-A3, SaIPT, SaCRE 10–15 ↓ | | (Zhu et al. 2021) |
Cynodon dactylon (genotypes) | C43 | | ST | tZR and DHZR ↑ iP, tZ, and DHZ ↓ | CYP735A ↑ | | (Yang et al. 2022) |
C198 | | SS | tZR and DHZR ↓ iP, tZ, and DHZ ↓ | CYP735A ↓ | |
Hordeum vulgare L. | Leaves | | | cZ- and iP-types ↑ tZ-types ↓ | | | (Torun et al. 2022) |
Roots | | | tZ- and iP-type ↔ cZ-types ↑ | | |
S. lycopersicum cv. MicroTom | | | | iPRMP, iPR, iP9G, tZ, tZOG, DHZR, cZRMP, cZR, cZ, cZOG, cZROG ↑ | 3 UGT85A1-like glucosyltransferase, 5 ZOG, CKX, RRs-A (8 and 9), RRs-B (1, 13), LOG‐like (1 and 8), PUP-like (3 and 5), HK3-like ↑ HPT-like (1 and 4), RR-A17, ZOG ↓ | | (Keshishian et al. 2018) |
Arabidopsis thaliana | Shoot | | | iPR, cZR ↑ DHZR, DHZ9G, cZROG, 2MeScZR ↓ | UGT73C1 ↑ UGT76C2, IPT9, CKX6 ↓ | | (Šimura et al. 2018) |
Root | | | iP, iPR, cZR, cZ9G, 2MeSiPR, 2MeScZR ↑ DHZ7G ↓ | LOG1-7, CKX6, UGT76C1 ↑ IPT3 and 5 ↓ | |
A. thaliana | | | SS | cZ and cZR, CK N-glucosides ↑ tZ, CK O-glucosides ↓ | IPT9, CKX3, 5 and 6 ↑ ARR-B1 and 12, ARR-A8 and 9, IPT3,5,7, AHK 2 and 4 ↓ | | (Prerostova et al. 2017) |
Thellungiella salsuginea | | | ST | The basal level 2 times lower in comparison to Arabidopsis. cZ, cZR ↑ tZ, tZR, tZ phosphate, and CK O-glucosides ↓ | TsIPT3, TsHK3 and TsRR1 ↑ TsIPT9, TsHK2 ↓ | Salt stressed Thellungiella had the total Na+ content 2 times lower than Arabidopsis |
Trigonella foenumgraecum | | | | Roots tZ ↑ iP ↓ Leaf tZ ↓ iP ↔ | | Na+ level in the roots of NaCl-treated plants 6-fold higher than in roots of control plants | (Belmecheri-Cherifi et al. 2019) |
Oryza sativa spp. japonica ‘temperate’ | | | Variety Baldo– ST | | | Na+ was restricted to the roots and older leaves, with the exclusion of salt from the third leaf | (Formentin et al. 2018) |
| | Variety Vialone Nano – SS | | | Na+ was more uniformly partitioned between the roots and shoot, and salt accumulation was observed in all the leaves |
A. thaliana | | Ipt1,3,5,7 (T-DNA) | ST ↑ | iPRP, iPR, tZRP, tZR, tZ7NG, tZ9NG and tZOG, iP and tZ ↓ | | | (Nishiyama et al. 2012) |
A. thaliana | | Ahp2,3,5 (T-DNA) | ST ↑ | - | AHP 2, 3 and 5 ↓ | | (Nishiyama et al. 2013) |
O. sativa | | OsAHP1 and 2 (RNAi) | ST ↓ | | OsHP2 ↑ | | (Sun et al. 2014) |
Gossypium hirsutum | | GhLOG3 (VIGS silencing) | ST ↓ | | GhLOG3 ↑ | | (Wang et al. 2021) |
| GhLOG3 OE (35S) | ST ↑ | |
S. lycopersicum cv. Improved Pope | | | | IPA and t-ZR ↓ | CKX35 (37 kDa) protein found in chlorotic leaves after salt-stress and associated with decreased CK levels. CKX37 (37 kDa) protein found in green leaves associated with normal CK levels | | (Cueno et al. 2012) |
Medicago sativa | | MsCKX OE (35S) | ST ↑ | | MsCKX ↑ | OE Na+ ↓ | (Li et al. 2019) |
O. sativa | | Inflorescence meristem-specific OsCKX2 RNAi Knockdown | ST ↑ | | | | (Joshi et al. 2018) |
A. thaliana | | ahk5-1 (T-DNA) | ST ↑ | | | | (Pham et al. 2012) |
S. lycopersicum cv Ailsa Craig | | SlIPT3 OE (35S) in AtIPT3 Knockdown mutants | | iPRMP and iPRDP ↑ | AtIPT3 ↑ AtIPT2 and AtIPT5, SlIPT3 and SlIPT4 ↓ | | (Žižková et al. 2015) |
| SlIPT4 OE (35S) in AtIPT3 Knockdown mutants | |
| SlIPT3 OE (35S) | ST ↑ | total CK ↑ iP7G ↑ | CKX in WT and OE ↔ | SlIPT3 OE Na+ ↓ |
S. lycopersicum cv. UC82-B | | A. tumefaciens IPT gene root-specific transformation (HSP70::IPT) | ST ↑ | tZ, tZR, iP, iPR in WT and HSP70::IPT ↓ | | Mutant roots Na+ ↓ and leaves K+ ↓ | (Ghanem et al. 2011) |
A. thaliana | | AtIPT8 OE | ST ↓ | In OE iP, iP9G, tZ, ZR and ZRMP ↑ | | | (Wang et al. 2015) |
G. hirsutum | | A. tumefaciens IPT gene transformation (Ghcysp) promoter) | ST ↑ | Z, ZR, iP, iPA ↑ | | | (Liu et al. 2012) |
Nicotiana tabacum | | A. tumefaciens IPT gene transformation (stress-inducible rd29A promoter) | ST ↑ | | | | (Qiu et al. 2012) |
O. sativa | | osrr9/osrr10 (CRISPR/Cas9) | ST ↑ | | OsRR9 and OsRR10 ↑ | osrr9/osrr10 shoots and roots had ↓ Na+/K+ ratios | (Wang et al. 2019) |
O. sativa spp. japonica | | AtUGT76C2 OE (maize ubiquitin promoter) | ST ↑ | In OE tZ7NG and tZ9NG ↑ | | | (Li et al. 2020a) |
N. tabacum | | AtUGT85A5 OE (35S) | ST ↑ | | UGT85A5 ↑ | Na+/K+ ratios similar in OE and WT under normal conditions; Under stress Na+/K+ ratios ↑ 18-fold in WT, and in 12-fold in OE | (Sun et al. 2013) |
A. thaliana | | AtCKX3 OE (35S) | ST ↑ | In OE iP- and tZ-type CKs ↓ | IPT1 and IPT3; CKX2, CKX4, CKX5, and CKX7 ↓ CKX1, CKX3, and CKX6 ↑ | | (Nishiyama et al. 2011) |
| AtCKX4 OE (35S) |
| ipt1,3,5,7 (T-DNA) | |
A. thaliana | | crf2 | | N- and O- conjugated forms, all CK nucleotide RMP compounds ↓ iP, DHZ, and cZ↑ all base active forms ↔ | | Non-stressed crf2 had 30% ↓K+level; After salt stress Mn2+ ↑ ~2X and Na+ ↑ ~40X | (Keshishian et al. 2022) |
O. sativa | | pup7 (T-DNA) | ST ↓ | tZ and tZR ↔ iP and iPR ↑ | | | (Qi and Xiong 2013) |
A. thaliana | | ahp2,3,5 | ST ↑ | | | | (Abdelrahman et al. 2021) |
| arr1,10,12 (T-DNA) | | | |
A. thaliana | | arr1,12 (B)* | ST ↑ | | | Shoot Na+ ↓ | (Mason et al. 2010) |
| ahp2,3,4 | |
| ahp3,4 | |
| Arr5,6,8,9 (A) | ST ↓ | | | Leaves Na+ ↑ |
| Arr3,4,8,9 |
| Arr3,4,5,6 (T-DNA) |
Physcomitrella patens | | PpCKX1 OE (Pro) | ST ↑ | | | | (Hyoung et al. 2020) |
↔ unaffected; ↑ upregulated; ↓ downregulated |
VIGS - virus-induced gene silencing |
AtUGT76C2, AtUGT85A5 - cytokinin glycosyltransferases |
PUP-type cytokinin transporter gene (purine permease) |
* - B-type ARRs ST – salt-tolerant SS – salt-sensitive OE – overexpression CKs: cZ - cis-zeatin; cZR - cis‐zeatin riboside; cZRMP – cis‐zeatin riboside-5’-monophophate; cZOG - cis‐zeatin O-glycoside; cZROG – cis‐zeatin riboside, O-glucoside; tZ - trans‐zeatin; tZR - trans‐zeatin riboside; tZOG - trans‐zeatin O-glycoside; DHZ - dihydrozeatin; DHZR - dihydrozeatin riboside; iP – isopentenyladenine; iPRMP - isopentenyladenine 9-riboside-5’-monophosphate; iPR - isopentenyladenine riboside; iP9G - isopentenyladenine 9-glucoside |
The expression of histidine kinases in response to salt stress varies in the different plant species: while no influence on the expression of Brassica rapa BrHK8, Cicer arietinum CarHK1 was down-regulated and Solanum lycopersicum HK3-like was up-regulated (Table 2). Interestingly, in a salt-sensitive Arabidopsis thaliana two CK sensing HKs (AHK2 and AHK4) were down-regulated, while in salt-tolerant Thellungiella salsuginea TsHK2 was down-regulated, and TsHK3 – up-regulated (Prerostova et al. 2017). At the same time, analysis of the ahk5 mutant suggested that AHK5 positively regulates salt sensitivity (Pham et al. 2012).
Similarly, the expression of HP genes was mostly up-regulated by salt stress in Corchorus sp., and Sorghum bicolor, and BrHP1 and 5 in B. rapa, SlHP1 and 4 in S. lycopersicum and OsHP2 in Oryza sativa. At the same time, pseudo-HP1 from B. rapa, CarHP1 in Cicer arietinum and AHP2, 3 and 5 in A. thaliana were down-regulated by salt stress (Table 2). The effect of HPs mutations on salt-stress related phenotype is also apposite, while ahp2,3,5 triple mutant showed increased salt-stress tolerance (Nishiyama et al. 2013), ahp1/2 knockdown with RNAi in rice resulted in salt-stress hypersensitivity (Sun et al. 2014).
RR genes were mostly up-regulated by salt stress in several species (B. rapa, S. lycopersicum, S. bicolor, T. salsuginea and O. sativa), and down-regulated in others (Corchorus sp. and A. thaliana) and mixed changes (both up- and down-regulation) in Cicer arietinum and Sophora alopecuroides) (Table 2). The relation to the salt stress could be clearly visible on the available A. thaliana RR mutants: type A mutants (arr5,6,8,9, arr3,4,8,9 and arr3,4,5,6) had increased salt stress sensitivity, while type B mutants (arr1,10,12, arr1,12 and osrr9/osrr10) had increased salt stress tolerance (Mason et al. 2010; Wang et al. 2019; Abdelrahman et al. 2021).
IPT genes were down-regulated by salt stress in S. alopecuroides and A. thaliana shoots and roots. However, in other experiments on A. thaliana and T. salsuginea IPT genes were both up- and down-regulated (Table 2). Interestingly, both types of genetic modifications (ipt1,3,5,7 T-DNA mutants and overexpressing (OE) lines of IPT3, IPT4 and IPT from A. tumefaciens) resulted in increased salt-stress tolerance. However, AtIPT8 OE plants demonstrated increased sensitivity to salt stress (Wang et al. 2015).
LOG genes have shown a rather unified response to salt stress – in all known plant species they were up-regulated (Table 2). These results were confirmed also on Gossypium hirsutum IPT3 gene (both VIGS silencing and 35S OE) mutants, where IPT3 OE lines were associated with increased salt-stress tolerance while silencing of this gene – with salt-stress hypersensitivity (Wang et al. 2021).
Mostly, CKX genes were up-regulated by salt stress (Table 2). Interestingly, both CKX OE (in Medicago sativa, A. thaliana and Physcomitrella patens) and knockdown (in O. sativa) resulted in increased salt-stress tolerance (Nishiyama et al. 2011; Joshi et al. 2018; Hyoung et al. 2020).
Despite the presence of diverse forms of CKs and their dynamic regulation and conversion in different organs, their levels are clearly correlating with corresponding biosynthesis (mainly IPTs, LOG and CYP735A), degradation (CKX) and transformation (N- and O--glucosyltransferases). In general, tolerance to abiotic stresses (including salt stress) is associated with reduced total CK content. Specifically, reduced levels of iP and tZ CK-types and increased cZ-types were related to salt tolerance. However, some deviations from this pattern have been found with respect to the plant age, type of explant used for measurements, salt-stress severity and exposure (Table 2).
The exposure of plants to salt (NaCl) stress leads to a significant increase in plant Na+ content. However, salt-tolerant plants had lower total Na+ content compared to salt-sensitive (Sun et al. 2013) (Prerostova et al. 2017). Furthermore, salt-sensitive plants have the uni-formal distribution of Na+ in different organs, while salt-tolerant plants tend to restrict Na+ into roots and elder leaves, with new leaves preserving photosynthetic function (Formentin et al. 2018) (Belmecheri-Cherifi et al. 2019). Interestingly, salt-tolerant transgenic plant lines with decreased (CKX OE) and increased (IPT OE) CK levels, or interrupted CK signalling (hpt and B-type arr) demonstrated reduced levels of the shoot and root Na+, thus, were less affected by toxic Na+ overload (Table 2).
Under salt stress, the alterations in the amount and forms of endogenous cytokinins vary greatly in different plant species and different organs of the same plant. Similarly, the expression of genes, responsible for CK signalling and metabolism, has no one universal pattern. Only plant LOG genes demonstrate stable up-regulation upon salt treatment in all studied cases. Therefore, at the moment there is no one-size-fits-all strategy that could be suggested to improve plant tolerance to salt stress via manipulation of CK signalling or metabolism genes. Apparently, every plant species should be evaluated independently and results from one species could not be extrapolated to another. At the same time, we have to admit that most analysed papers were descriptive by nature, when a certain parameter was evaluated and compared (CK form, gene or protein level) under normal and stress conditions. However, with no attempt to elucidate the causative role and molecular mechanism of the stressor effects on the studied system. Nonetheless, we strongly believe that further intensification and expansion of plant salinity and stress response research will significantly clarify and uniform the role of CK signalling and metabolism as well as other phytohormones in these mechanisms