**1. Anatomy and development of root**

#### **1.1 Root system architecture**

In different plant species, root system architecture (RSA) has diverse morphologies. There are basically two types of RSA, the taproot system (or allorhizic system) in gymnosperms and dicotyledons, like *Arabidopsis thaliana* (*Arabidopsis*), tomato (*Solanum lycopersicum* L.), carrot (*Daucus carota*), and poplar (*Populus* spp.) and the fibrous root system (or homorhizic system) in monocotyledons such as maize (*Zea mays* L.), rice (*Oryza sativa* L.), onion (*Allium cepa*), garlic (*Allium sativum*), and tulip (*Tulipa* spp.) [1]. The taproot system consists of a single thick central primary root (PR) with thin or no lateral roots (LRs); the fibrous root system has small and short-lived primary and adventitious roots (ARs) derived from shoots, stems, or leaves [1].

### **1.2 Intrinsic developmental signals and environmental conditions modify root system architecture**

*Arabidopsis* as a eudicot has a taproot system, which consists of an embryonic radicle-derived PR and postembryonic-developed LRs and ARs. Root regeneration exists throughout the plants' lifetime; it is a distinctive feature of plants and contributes to their robustness in adverse conditions.

In *Arabidopsis*, LRs initiate from pairs of pericycle cells that possess developmental potential as plant stem cells. These pericycle cells are selected and directed to become LR founder cells and form LRs by both intrinsic and environmental signals [2–5]. The primary LR is initiated from the basal meristem of the PR, where root cap-derived auxin influences the amplitude of oscillatory gene expression in the basal meristem and the elongation zone of the root, which leads to the prepatterning of LR initiation sites [6, 7]. The pre-patterning process is marked by the expression of a series of genes, like *GATA23*, *MEMBRANE-ASSOCIATED KINASE REGULATOR 4* (*MAKR4*), and *IAA19* [7]. In the basal meristem and elongation zone, *DR5::Luciferase* expression was observed to rhythmically pulse with a period of ~6 h, which matched with the period of LR pre-branch site production [6]. It is recently reported that the source of auxin is provided by the cyclic programmed cell death of root cap cells [8, 9].

It is noteworthy that not all of the pre-branch sites emerge to be LRs [6]. The dormant pre-branch sites may present a selective mechanism for LR formation under certain growth conditions, such as water availability, nutrient levels, physical obstacles, or damage [5, 10–13]. It is interesting that many of the external signals converge on phytohormones to regulate root development. Among these phytohormones, auxin functions as a central mediator.

Mechanical forces are important regulators for plant morphogenesis. LRs always emerge from the convex side of PR bending, resulting in a left-right alternation of LRs. Bending caused by gravitropic curvature led to the initiation of LRs, where a subcellular relocalization of PIN1 was observed [11]. Release the pericycle cells from the restraints of adjacent endodermis by targeted single cell ablation of endodermal cells triggered the pericycle to reenter the cell cycle and induced auxindependent LR initiation [14]. Excision of the *Arabidopsis* PR leads to the promotion of LR formation, which is mediated by activated auxin biosynthesis and auxin transport [15].

## **2. Roles of phytohormones on root formation**

#### **2.1 Auxin**

The phytohormone auxin which plays fundamental roles in many aspects of plant growth and development is also a well-documented key regulator of LR development [16, 17]. The natural auxin, indole-3-acetic acid (IAA), is mainly synthesized in a two-step pathway from tryptophan. First, tryptophan is converted to indole-3-pyruvate (IPA) by the TAA1/SAV3 family of aminotransferases; IPA is then converted to IAA by the YUCCA (YUC) family of flavin monooxygenases [18–23]. Auxin biosynthesis has been shown to play an essential role on both programed and wound-induced LR and AR developments [15, 24, 25].

Polar auxin transport (PAT), mediated by auxin influx (AUX1 and LAXs) and efflux carriers (PINs and MDR/PGPs) [26–29], generates auxin gradients and maintains an auxin maximum to regulate LR formation and positioning [17, 30–33].

Auxin signaling is known to be an integrator of endogenous and exogenous signals for root branching [17, 30, 34, 35]. It begins with the degradation of a class of AUXIN/INDOLE-3-ACETIC ACID (Aux/IAA) through TRANSPORT INHIBITOR RESPONSE 1 (TIR1) auxin receptor [36, 37], resulting in the activation of the AUXIN RESPONSE FACTOR (ARF) [38, 39]. ARF7 and ARF19 transcription factors further induce the expression of downstream target genes like *LATERAL* 

**23**

*Phytohormone-Mediated Homeostasis of Root System Architecture*

*ORGAN BOUNDARIES DOMAIN/ASYMMETRIC LEAVES2-LIKE* (*LBD/ASL*) family genes *LBD16/ASL18* and *LBD29/ASL16*, promoting LR initiation at the

Cytokinin is also a main player in root development. In higher plants, isopentenyladenine (iP), trans-zeatin (tZ), and dihydrozeatin (dZ) are the predominant cytokinins [44]. Cytokinin level and patterning in plant are controlled by a fine equilibrium between cytokinin synthesis and catabolism [44, 45]. Cytokinin biosynthesis is dependent on the activity of *ATP/ADP-isopentenyltransferase* (*IPT*) and *LONELY GUY* (*LOG*) gene family [46–48], and the metabolism is mainly through the *CYTOKININ OXIDASE/DEHYDROGENASE* (*CKX*) genes [44, 45]. Cytokinin can also be inactivated through conjugation to glucose [49]. The spatial and temporal distribution of cytokinin is in part due to the specific expression of cytokinin

In *Arabidopsis*, cytokinin signaling starts with the perception by the transmembrane cytokinin receptors ARABIDOPSIS HISTIDINE KINASE (AHK), AHK2, AHK3, and AHK4/WOL1/CRE1 [53–55], which target the histidine phosphotransfer protein AHPs to activate the type-A and type-B ARABIDOPSIS RESPONSE REGULATORS (ARRs) that negatively and positively regulate cytokinin signaling, respectively [55–61]. Two type-A ARRs, ARR7, and ARR15 were induced by both cytokine and auxin and are essential for embryonic root development [62].

Although some evidences showed that cytokinins act as both local and longdistance signals [51, 63–65], and some transporter proteins have been shown to be involved in cytokine transport [66–70], the molecular mechanisms of cytokinin

Postembryonic root development is regulated by the root apical meristem (RAM), where cytokinin is known to act antagonistically with auxin to control the balance of cell division in the division zone and cell differentiation in the transition zone, which is essential for the maintenance of the RAM and affects the growth and patterning of the root [64, 71]. Application of cytokinin reduces the number of meristem cells and the size of RAM and promotes cell differentiation in the transition zone; cytokinin biosynthesis and signaling mutants as well as *CKX* overexpression mutants have a larger RAM with more meristem cells [45, 64, 72]. Conversely, auxin treatment increases meristem size and promotes cell division in the proximal meristem, and auxin transporter *PIN* mutants display a smaller RAM [64, 73]. The cross-talk of cytokine and auxin in regulating RAM activity was shown to converge on the auxin-inducible *AUX/IAA* family gene *SHORT HYPOCOTYL 2/ INDOLE-3-ACETIC ACID 3* (*SHY2/IAA3*) in the transition zone, where cytokinin activates *SHY2* via the *AHK3/ARR1* two-component signaling pathway to suppress *PIN3* and *PIN7* expression and promote cell differentiation, while auxin suppressed SHY2 protein, leading to the activation of PINs and promotion of cell division [71]. Furthermore, Moubayidin et al. [74] revealed that in transition zone, SCR, a member of the GRAS family of transcription factors, directly represses the expression of *ARR1*, which controls auxin production via the *ASB1* gene and sustains stem cell activity, to simultaneously control stem cell division and differentiation and ensure coherent root growth. Cytokinin affects the expression of multiple *PINs* differentially in a tissue-specific manner to regulate auxin distribution [75, 76]. Auxin-cytokinin interactions lead the generation of distinct hormonal response

zones, thus controlling the development of root vascular tissue.

On contrary to auxin, which is a positive regulator of LR development, cytokinin

acts as a negative regulator of LR formation. Cytokinin suppresses LR initiation

*DOI: http://dx.doi.org/10.5772/intechopen.82866*

protoxylem-pole pericycle cells [40–43].

synthesis and catabolism genes [45, 47, 48, 50–52].

transport are still not well characterized.

**2.2 Cytokinin**

*ORGAN BOUNDARIES DOMAIN/ASYMMETRIC LEAVES2-LIKE* (*LBD/ASL*) family genes *LBD16/ASL18* and *LBD29/ASL16*, promoting LR initiation at the protoxylem-pole pericycle cells [40–43].
