Introduction

As an important organ by which plants maintain their normal growth and development, plant root mainly plays role in fixing plants, as well as absorbing and transporting water and nutrients from the soil (Abdi et al. 2019; Hodge et al. 2009; Martinez-Arias et al. 2020). Roots usually acquire vital resources from the soil and can directly feel changes in various environmental conditions in the soil and subsequently respond in real time (Armstrong et al. 1991; Chapman et al. 2020). Whether the root system grows well it can directly affects the growth status of the aboveground parts of the plant, that is, the yield of crops, but the problem of food yield still perplexes many countries throughout the world.

Phytohormones primarily refer to chemical substances that can regulate the growth and development of plants. These phytohormones are synthesized in plants and act as molecular signals at very low concentrations. Although their contents in plants are low, they play an indispensable regulatory role in the process of plant growth and development. Cytokinins (CKs), as one of the classic plant hormones, can promote cell division, accelerate the growth of collaterals, delay ageing, and regulate the transportation of nutrients (Collier et al. 2000; Jones and Schreiber 1997; Kurepa et al. 2018; Wang et al. 2020a, b; Werner et al. 2003; Zhang et al. 2005). At present, the effects of CKs on plant root growth and development, as well as the response of plants to abiotic stress, have been reported (Gao et al. 2014), which is primarily due to the important regulatory role played by the CKs signal transduction pathway and its components on plant roots. CKs have a fine regulation mechanism on roots and play a unique role in the growth and development of plant roots under abiotic stress. Additionally, CKs can combine with plant hormones, such as auxin (IAA) and abscisic acid (ABA), to regulate root growth and development (Albacete et al. 2008; Nguyen et al. 2018), but to date, there is no systematic summary.

With the progress of science and technology, researchers have made great progress in the area of plant hormone regulation of root growth and development and the abiotic stress response. This paper briefly describes the basic situation of Arabidopsis roots and CKs and summarizes the progress in the field of CKs regulation of the plant root response to abiotic stress. Based on the work in recent years, this article summarizes the regulatory mechanisms of CKs and its combination with other plant hormones on Arabidopsis root growth and development, as well as the abiotic stress response to provide certain information for researchers, which is conducive to performing follow-up research.

Basic characteristics of the Arabidopsis root

The root systems of plants can be divided into taproot and fibrous root systems according to their basic configuration (Fig. 1a). The axial root of the taproot system clear and well developed, and the direction is vertical downward. However, the fibrous root system has no obvious taproot and lateral roots (LRs), and the shape of the whole root system is flocculent. Generally, the root systems of dicotyledons are mostly taproot, while the roots of monocots are usually fibrous. The following is mainly elaborated by using the root system of Arabidopsis thaliana, a dicotyledonous model plant in plant biology, to briefly introduce the structure of plant root systems and the process of LRs development.

Fig. 1
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Overview of the Arabidopsis thaliana root

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Basic structure of the Arabidopsis root

The root of Arabidopsis is a typical taproot system, composed of the axial and LRs, which could be clearly distinguished in appearance. According to the cell morphology and structure of the root, it can be divided into the root cap, meristem, elongation, and maturation zone. The root cap and meristem zone together constitute the root apex, and cells in the elongation zone finally cannot elongate longitudinally to become mature cells, which are usually accompanied by root hair (Fig. 1b) (Dello Ioio et al. 2007). The cells at the junction of the root cap and meristem do not divide; thus, it is called quiescent center (QC). Various types of stem cells surround the QC, which can maintain root growth through asymmetric division. Generally, the QC can generate small scale signals to control the stem cells around it. The signal area covered by the QC is usually called the stem cell niche. The reason why stem cells can produce offspring cells is that they exceed the range of stem cell niches before they begin to differentiate (Fig. 1c) (Azpeitia et al. 2010, 2013). The Arabidopsis transcription factor SHORT-ROOT (SHR) can affect cell division by regulating the microenvironment of active oxygen in the root tip (Li et al. 2018). CKs are primarily produced in the root tip, and CK mutants ahk2/3/4 and arr1/10/12 will disrupt root tip QC specialization, as well as stem cell maintenance and differentiation, and reduce the de novo rate of root growth, which shows that in the absence of CKs, the process of the de novo guidance and initiation of roots can occur, while the specialization of QC in the root apical meristem and the differentiation of stem cells in the distal stem cannot proceed normally (Sun et al. 2016; Yokoyama et al. 2007; Zhang et al. 2013). Taken together, these findings indicate that the function of CKs is region-specific.

Development of the Arabidopsis LRs

The LRs of Arabidopsis usually originate from the pericycle cells near the xylem pole, which is different from the monocotyledonous where LRs generally derive from the pericycle cells near the phloem pole. However, only a few of the producing cells can eventually develop into LRs. LRs have an essential role in increasing the water absorption area of the root system and maintaining the overall structure of the root system. The development of LRs has been systematically divided into 8 detailed stages from initiation, germination, and elongation in recent years (Fig. 1d) (Malamy and Benfey 1997). In the initial stage, the two adjacent generating cells undergo unequal vertical division followed by radioactivity to expand the cell layer, gradually showing a conical shape. In the germination phase, the LR primordium further divides and expands, breaking through the endodermis, cortex, and epidermis in turn. Finally, the LR primordium enters the elongation stage and develops into mature LRs under the regulation of the newly formed LR meristem (Benkova and Bielach 2010). It has been reported that the two receptor-like protein kinases MUSTACHES (MUS) and MUSTACHES-LIKE (MUL) of Arabidopsis rich in leucine-rich repeat (LRR) sequences can regulate cell wall synthesis and remodelling to control the early development of Arabidopsis LR primordium (Xun et al. 2020). Usually, there are few atypical aspartic proteases (APs) in plants, but recently, it was found that overexpression of APs in Arabidopsis roots inhibited the development of LRs, suggesting that APs have a previously unknown biological role (Soares et al. 2019). The signal peptide treatment of CK oxidase 2 (CKX2) affects its enzymatic activity, thereby determining the degradation of CKs in natural Arabidopsis. The CK signal interferes with the growth of the upper LR flanks, thereby preventing downward bending (Waidmann et al. 2019). We can see that there are many ways to adjust the development of LRs in Arabidopsis, which involve different proteins and signalling molecules. However, there are still many unknown ways or patterns to be discovered.

Synthesis, signal transduction, and molecular mechanism in the regulation of Arabidopsis root growth by CKs

CKs are widely distributed in higher plants, and their content is in range of 1–1000 ng/g. Although the content is very low, its role is integral. CKs are usually distributed in the apex, stem tips, germinated seeds, and growing fruits of plants. The root tip is the primary place for CKs synthesis, which are primarily transported upward through the xylem (Aloni et al. 2005; Cedzich et al. 2008). The following principally introduces the synthesis of CKs, signal transduction pathway, and molecular mechanism in regulating the growth and development of the Arabidopsis root system.

Synthesis of CKs

Currently, there are two known synthetic pathways of CKs, the de novo synthetic pathway is the primary one, and the tRNA decomposition pathway is the auxiliary one. Among them, the de novo synthetic pathway catalysed by isopentenyl transferase (IPT) is a conserved mechanism in evolution. The IPT protein encoded by the 7 AtIPT genes of Arabidopsis takes part in the de novo synthesis of CKs. Additionally, overexpressing the IPT gene can facilitate the synthesis of CKs (Galichet et al. 2008; Sakakibara and Takei 2002; Takei et al. 2004). The de novo synthetic pathway begins through the combination of dimethylallylpyrophosphate (DMAPP) and adenine ribonucleotide (AMP) or adenosine triphosphate (ATP)/adenine nucleoside diphosphate (ADP). Isopentenyl nucleoside is formed under the catalysis of cytochrome p450 monooxygenase (CYP735A), which is capable of converting isoamylene-type CK into zeatin nucleotide-type CK, but the CK at this stage does not have physiological activity and needs to be activated under the action of phosphoribosyl hydrolase (LOG) (Kakimoto 2003; Zurcher and Muller 2016). Nine LOG encoding gene families have been identified in Arabidopsis (Tokunaga et al. 2012). Another tRNA decomposition pathway is catalysed by tRNA-IPT, and the 3 ends of some tRNA anticodons are prenylated, which decomposes tRNA and finally produces cis-zeatin (cZT) (Mok and Mok 2001). Studies have demonstrated that the Arabidopsis AtIPT2/9 gene can encode these, and the cZT content is significantly reduced after mutation (Miyawaki et al. 2006; Xu et al. 2016). Currently, the research on the synthesis of CKs is relatively clear, and considerable progress has been made in the related research on the signal transduction pathway, but it is not very profound yet.

Signal transduction pathway of CKs

The signal transduction pathway of CKs is a multi-step phosphotransfer reaction (Fig. 2). CKs bind to the CHASE domain of CKs receptors (AHK2, AHK3 and AHK4/WOL1/CRE1), which are located on the plasma membrane, thereby activating their histidine kinase activity (Ishikawa et al. 2002; Pas et al. 2004). The phosphate group is transferred to the aspartate receptor domain connected to it, and it is subsequently transferred to the histidine of the histidine-phosphotransfer protein (AHP) and phosphorylated. The phosphorylated AHP protein enters the nucleus and transfers the phosphate groups to the type-A ARR and type-B ARR receptor domains in the nucleus. The phosphorylation of type-B ARR can activate the output domain and induce the transcription of the type-A ARR gene. The activated type-B ARR output domain is also able to affect downstream transcriptional regulation by affecting CKs. At the same time, type-A ARR can also be directly phosphorylated by the AHP protein to cause a CKs response (Haberer and Kieber 2002; Horak et al. 2008; Jeon and Kim 2013). For the research of the CKs signal transduction pathway, the core issue is the exploration of its new receptors. At present, it still only has several known receptors. In the future, we will focus on exploring and discovering new receptors. Going forward, the discovery of new receptors will inevitably become the focus and difficulty of foreign research.

Fig. 2
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Signal transduction pathway of Arabidopsis thaliana cytokinin

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Mechanism of CKs in regulating the root growth of Arabidopsis

Root growth is regulated by a variety of factors, including plant hormones, transcription factors, and small RNAs (miRNAs) (Chen et al. 2020; Hernandes et al. 2020; Lin and Sauter 2020; Wang et al. 2020a, b; Ye et al. 2020; Zhou et al. 2020a). To date, researchers have performed various studies in these areas. The root tip of Arabidopsis is one of the staple places where CKs are produced. The CK content in the plant is usually increased by externally applying CKs or genetically modified technology. Compared with the promotion effect on the above-ground part (Riefler et al. 2006), for the growth of the underground part, CKs mainly play a negative regulatory role, which can affect the growth and development process of the plant's axial root elongation and LR development. The following mainly summarizes the role of CKs in growth and development of the axial and LRs of Arabidopsis from different aspects.

Mechanism of CKs affecting the growth of the axial roots of Arabidopsis

The effect of CKs on growth and development of Arabidopsis axial roots can mainly be affected by the regulators and receptor proteins in the CKs signalling pathway. Although the main physiological function of CKs is to affect cell division, regulation of taproot growth is not achieved by directly affecting the division of cells in the meristem area but by affecting the rate of differentiation. The meristematic region of taproot became smaller after exogenous application of CKs. But, if the key genes in CKs signaling pathway were mutated, the opposite phenotype appeared (Li et al. 2006). Additionally, after mutating all of the CK receptors, it was found that the rate of cell division in the root meristem decreased significantly (Higuchi et al. 2004; Nishimura et al. 2004). Nitrate can be used as one of the effective indicators to regulate the growth and development of plant roots. CKs sensing and biosynthesis mutants have shorter taproots than wild-type plants when nitrate is the only source of nitrogen, and the root tip tissue scientific analysis showed that the cell division and elongation of the double mutant of CK receptor ahk2/ahk4 were reduced compared with wild-type plants under sufficient nitrate conditions (Naulin et al. 2020). This indicates that nitrate can stimulate the growth of the primary roots by increasing the meristem activity and CKs signal. The balance between cell division and differentiation is mainly achieved by the SHORT HYOPCOTYL2 (SHY2) gene through the regulation of IAA and CKs signalling pathways. SHY2 protein is one of the key points for plant IAA, brassinosteroids, and CKs to regulate root meristem development (Li et al. 2020). Besides, according to Dello Ioio that an AHK3/ARR1, AHK3/ARR12 two component cytokinin signaling pathway mediates control of root-meristem size at the transition zone (Dello Ioio et al. 2007).

Except for these regulators and receptor proteins in the CKs signalling pathway can affect the axial root, the miRNAs can also have great impact on the axial root. The miRNAs are a type of endogenous non-coding RNA with regulatory functions that exist in eukaryotes, which can participate in multiple processes of plant growth and development, and their length is usually 22 nucleotide. The miR156 and SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) genes have opposite expression patterns during the primary root (PR) growth of Arabidopsis. Plants with a higher level of miR156 exhibit decreased size of the meristem, leading to the shortening of the main root. In contrast, plants with reduced levels of miR156 showed higher meristem activity. Meristem activity is regulated by SPL10 probably through the reduction of cytokinin responses, via the modulation of ARR1 expression (Barrera-Rojas et al. 2020).

Mechanism of CKs affecting the growth of Arabidopsis LRs

The growth and development of LRs are also regulated by many factors. CKs play an indispensable role in the distribution direction of LRs as well as the initiation and extension of LRs. IAA can promote the downward bending of the lower root organs, but the CKs signal, as a unique anti-gravity component of the LR, can promote the radial distribution of the root system. Among these factors, cytokinin oxidase 2 (CKX2) mainly determines the degradation of CK in natural Arabidopsis. The CK signal interferes with the growth of the upper root side and prevents downward bending (Waidmann et al. 2019). At present, most studies believe that CKs act as an antagonist of IAA during the formation of LRs, and CKs inhibit the initiation of LRs (Chang et al. 2013). The reason may be that CKs inhibit cyclin (CYC) and the cell cycle during the initiation of LRs. The expression level of protein-dependent kinase (CDK) can inhibit the extension of LRs by increasing the length of epidermal cells, and this pathway is regulated by IAA-dependent pathways. Mutations in CKs receptors AHK2 and AHK3 can increase the sensitivity of IAA to LR formation. With regards to the functional relevance of IAA-CKs interaction during LR formation, in addition to IAA-dependent pathways, LR formation may also be achieved by promoting CYC expression activity (Chang et al. 2013). Abscisic acid insensitivity factor 4 (ABI4) mediates the inhibition of ABA and CKs on the formation of LRs by reducing IAA transport, leading to the reduction of IAA in the roots and ultimately inhibiting the development of LRs (Shkolnik-Inbar and Bar-Zvi 2010). It can be seen that CKs can not only affect the initiation and extension of Arabidopsis LRs but also work together with other plant hormones. In this process, the factors in the CKs signalling pathway also play a vital role.

Effect of CKs on plant roots under abiotic stress

As the external environmental conditions change, the phytohormone content in plants often changes accordingly. In previous studies, it was found that ABA plays an important role in regulating the adaptation of plants to adversity; thus, it is also called the adversity hormone (Ma et al. 2018; Wu et al. 2020; Yu et al. 2020). In recent years, it has been discovered that CKs also play a unique role in plant resistance to adversity (Bielach et al. 2017; Cortleven et al. 2019; Ryu and Cho 2015). CKs can affect the growth and development of plant roots under drought stress, cold stress, salt stress, and various chemical element stresses. The influence of CKs on the plants growth and development under abiotic stress in recent years is summarized in Table 1. The related literature on root growth and development can provide a systematic understanding of the role of CKs in roots under different abiotic stresses. On the whole, CKs mainly affects the growth of plant roots under abiotic stress through its interaction with other phytohormones.

Table 1 Function of cytokinins in growth and development of plant root under abiotic stress
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Based on the research results in the table, it can be seen that there has been significant progress in CKs regulation of root growth and development under abiotic stress. While different types of stress are involved, the research objects are not limited to Arabidopsis but also in maize and rice. Corresponding research work has also been performed in other crops. At present, especially under cold stress and osmotic stress, the research on the effect of cytokinin on root system is more comprehensive. Cytokinin-responsive factors CRF2 and CRF3 play a significant role in regulating the initiation of Arabidopsis LRs at low temperatures, which mainly include two types of regulatory pathways, namely, the two-component signal transduction pathway (TCS) dependent on CK to induce the expression of CRF2, and it does not rely on the TCS pathway to induce CRF3 upregulation, thereby inhibiting or promoting the growth of LRs (Jeon et al. 2016). Additionally, CKs bind to receptors AHK2 and AHK3, and the expression of type-A ARR at this time has a negative regulatory effect on cold stress. At the same time, CKs binds to receptors to inhibit the response of ABA to cold stress (Fig. 3) (Jeon and Kim 2013; Jeon et al. 2010; Nishimura et al. 2004; Xia et al. 2009). Under osmotic stress, this response inhibits the synthesis of CKs, thereby inhibiting the growth and development of the root system. At the same time, osmotic stress promotes the high expression of PIN2 protein and inhibits the synthesis of IAA. The size of the root meristem increases with the decrease of the IAA concentration, thereby affecting the normal growth of the root system (Fig. 3) (Hussain et al. 2015; Lakehal et al. 2019; Rowe et al. 2016). It can be seen that CKs are indispensable in regulating of root growth and development under abiotic stress.

Fig. 3
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Pathway of CKs regulating root growth under cold and osmotic stress

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Conclusion and prospects

In recent years, researchers have conducted a series of in-depth studies on plant roots using molecular biology and genetics. These researchers have realized that plant roots play a critical role in plant growth and development, as well as the response of the plant to abiotic stress. Much outstanding progress has also been made in the field, especially in the related research represented by the dicotyledonous model plant Arabidopsis, which has notable reference significance for subsequent research on other plants.

Recently, many advances have been made in understanding the role of plant hormone abiotic stress and other factors in the root growth and development of Arabidopsis, but many problems still need to be further understood and explored as follows: (1) Although it has been reported that various phytohormones regulate growth and development of Arabidopsis roots through the mutual interaction, are there other factors that work together in the interaction between hormones? The dominating factors and the roles they play need to be further explored. (2) When plant hormones regulate the growth and development of Arabidopsis roots, they usually act through the signal transduction pathway. New signalling pathways and receptors based on pathways are need to be explored along with enriching the regulatory pathways and signal networks of phytohormones on Arabidopsis roots. (3) Future research needs to fully integrate the growth and development of the above-ground and underground parts of plants to systematically explore the mutual regulation mechanisms and signal transduction pathways between them.

In brief, further strengthening the research on the regulation mechanism of plant hormones and root growth and development and the response to abiotic stress can motivate our in-depth understanding of the molecular regulation mechanism and signal network of plant hormone regulation of plant root growth and development. These results may help achieve greater research progress and then provide a solid technology and information basis for crop breeding and cultivation, as well as mechanized intensive planting in the future.