Auxin and Environmental Factors Regulate Root Gravitropism

Abstract

Roots are important for plant anchoring, water and nutrient absorption, and other physiological processes. Gravity is a primary determinant of the spatial distribution of plant roots in the soil. Therefore, in-depth understanding of the molecular mechanisms and biochemical networks of root responses to gravity has both theoretical and practical significance in guiding the genetic improvement of plants. Gravitropism, the process through which plants sense the direction of gravity and respond by making the roots grow downward and the stem grow upward, has been widely studied in roots. The perception of gravity and the gravitational growth of roots-key steps in root growth and developmentare regulated by auxin and other factors. Here, we review the latest progress in the regulation of root gravitropism by hormone signals and environmental factors from a molecular perspective, and look forward to the direction of future research on root gravitropism.

Keywords: Root, Hormone, Gravity, Auxin, Environmental

Introduction

Plants sense external environmental signals, such as light, water, temperature, and gravity, and adjust their growth accordingly to adapt to a changing environment [1-3]. Among these signals, gravity is important for regulating plant growth, development, and morphogenesis. In gravitropism, plants relocate their growth after sensing a gravity stimulus to maintain an optimal angle between the organs and the direction of gravity [4]. For seeds sown in soil, regardless of the orientation of the embryo, the radicle and germ exhibit fixed growth directions: the root grows downward (consistent with the direction of gravity) in a process termed positive gravitropism; the bud (stem) grows upward (in the opposite direction of gravity), which is termed negative gravitropism. Furthermore, the underground stems of some plants (such as reeds) grow in a horizontal direction, in a process called diagravitropism. Gravitational movement occurs only in growing areas of the plant. For example, when lodging occurs in rice and wheat plants, the upper part of the plant can stand upright again. Root gravitropism is especially important insofar as the roots of newly germinated seedlings must immediately enter the soil (i.e., grow in the direction of gravity) to ensure access to water and nutrients [5]. Here, to enable researchers to better understand root gravitropism research, we present a systematic review of research fields related to root gravitropism.

The Four Stages of the Root Gravity Response

In the root, gravity is sensed in the columella cells (CCs) of the root cap; however, the response to gravity occurs in the root elongation zone. Therefore, there are obvious physical zones between the area in the root that senses gravity and the area that responds to gravity. The root gravity response can be divided into four stages: the perception of gravity (the conversion of physical forces into biochemical signals), the transduction of gravity signals (the output of biochemical signals), the asymmetric distribution of auxin, and the bending of roots during growth [6].

Gravity Perception

The root cap is the main site where plant roots detect gravity. Early experiments found that removing the root cap leads to the loss of root geotropism [7]. Further experiments using techniques such as laser excision of root cap 83 cells, gene knockout, and heavy ion microbeam irradiation showed that CCs throughout the root cap are gravity-sensing sites [8,9]. In Arabidopsis thaliana (Arabidopsis), the central S2 zone of CCs is most 86 important for the geotropism of roots [8]. The nucleus is located at 87 the top of each CC, the endoplasmic reticulum forms a cup-shaped structure at the bottom of the cell, and the bottom of the cytoplasm is occupied by numerous amyloplasts [10].

At present, it is generally accepted that the mechanism by which plants sense gravity can be described by the starch– statolithhypothesis. According to this proposal, when the direction of gravity changes, starch grains in the cell settle along the direction of gravity, leading to changes in various signaling molecules. These changes affect the transport of auxin, causing it to be distributed asymmetrically in the direction of gravity stimulation, which results in the bending of roots during growth [11,12]. Mutation of the gene PGM causes loss of function of glucophosphomutase, the key enzyme in starch synthesis, and the loss of amyloplasts. As a result, Arabidopsis pgm mutant cannot sense the gravity signal and are therefore deficient in gravitropism [13]. Although the pgm-1 mutant is less sensitive to gravity than the wild type, however, the roots could still bend after prolonged gravity stimulation treatment (the bending angle became smaller). This suggests that plants might have a variety of gravity-sensing mechanisms. When the osmotic pressure of the external environment changes, the root bends to varying degrees, but sedimentation of the starch grains is not affected. This has led to the proposal that the protoplast itself can sense changes in gravity (the protoplast hypothesis). According to this hypothesis, protoplasts can sense gravity via changes in their own mechanical buoyancy. Under gravity stimulation, protoplasts settle to the cell wall or extracellular matrix. The resulting pressure difference between the plasma membrane and extracellular matrix activates gravity receptors and transmits gravity signals [14]. Therefore, the mechanisms described by the starch-statolith and the protoplast hypothesis are coordinated to enable gravity sensing. Additional hypotheses to describe root gravitropism include the bifurcation theory and the cytoskeleton theory. The amyloplast theory states that in the roots of higher plants, balanced cells can sense gravity through interactions of both bound and unbound amyloplasts with the cytoskeleton. When amyloplasts are bound to the cytoskeleton through cell membrane-related receptors or proteins, sedimentation of the amyloplasts under gravity stimulation alters cytoskeletal tension, the receptors on the cell membrane are activated, and downstream signals are also triggered to respond to gravity stimulation. When not bound to the cytoskeleton, the amyloplasts can move freely with changes in the direction of gravity and eventually settle onto the endoplasmic reticulum on the lower side of the cell. This triggers downstream signal transduction and gravity sensing [15]. The bifurcation theory, meanwhile, proposes that the nucleus and cytoskeleton are particularly critical in gravity sensing and signal transduction, and a gravity-sensitive window switches in cells to sense gravity signals [16].

Transduction of Gravity Signals

After amyloplasts are deposited in the cell, the cytoskeleton might rearrange itself, and the stimulation might be transferred to the endoplasmic reticulum or cell membrane to induce the opening of ion channels. Ca2+, inositol 5-trisphosphate (InsP3), pH, and other signaling molecules are used as second messengers to participate in the gravity response [17,18]. Indeed, exogenous calcium channel inhibitors and Ca2+-ATPase activity inhibitors can block the gravity response, and the use of calcium analogs can enhance the gravity responses of hypocotyls and roots [19-23]. These findings indicate that changes in Ca2+ concentration are activated in response to gravity stimulation, thereby regulating the plant response to gravity.

Not only do the concentrations of Ca2+ in cells change under gravity stimulation, but the abundance of the second messenger inositol 1,4,5-trisphosphate (InsP3) also increases [24]. Exogenous application of the amino acid steroid U73122, which inhibits the formation of InsP3 in roots, attenuated the gravitational response of roots [25]. The lack of enzymes to degrade InsP3 in the 5pt13 mutant caused InsP3 accumulation, resulting in an enhanced response to gravity. Phosphatidylinositol Monophosphate-5-Kinase (PIP5K) is the synthetic precursor of InsP3. In pip5k roots, the gravity response is delayed, and the ability of auxin transport carriers to flow between vesicles and the plasma membrane is weakened. These observations suggest that InsP3 regulates the locations of transport carriers by regulating vesicle transport, thereby participating in the regulation of the root gravity response [26-28]. Under stimulation by gravity, the pH of the proximal side of the root decreases, and the corresponding pH value of the distal side increases. This asymmetric change in pH mainly depends on the activities of Ca2+ channels [29]. These findings indicate that a variety of signaling molecules function in gravity signal transduction.

Asymmetric Distribution of Auxin and Curved Growth of Roots

Perception of gravity occurs in CCs, but response to gravity is completed in the root elongation zone. The Cholodny–Went hypothesis explains this phenomenon through the proposal that during gravity sensing, sedimentation of starch granules leads to the transmission of the gravity signal to the response area of gravity-the elongation zone-via auxin. Following gravity stimulation, auxin is asymmetrically distributed in the root, resulting in the asymmetric growth of cells on both sides of the root that leads to root curving during growth [30]. Analysis using the auxin response reporter gene constructs proDR5: GFP and proDR5: GUS revealed that, under gravity stimulation, the distribution of the signal in the distal and proximal sides of the root was asymmetric, indicating that auxin was distributed in an asymmetric manner [31]. Therefore, auxin is important in a plants reaction to gravity. Indeed, mutants in auxin biosynthesis, transport, and signal transduction can exhibit abnormal gravity responses [32-34].

Effects of Auxin Transport on the Gravity Response

The auxin transport pathway is important in regulating the gravity response [35]. The asymmetric distribution of auxin transport carriers leads to an asymmetric distribution of auxin. This, in turn, leads to the asymmetric growth of cells on both sides of the root, resulting in root bending [36,37].

AUX1, the first influx carrier identified for auxin, is crucial in the response of plants to gravity [38]. Auxin transport is abolished in the roots of the Arabidopsis aux1 mutant, and the response to gravity in its roots is weakened [39]. The gravitational response of roots depends on the transport of auxin from the lateral root cap to the epidermis of the elongation zone, a process mediated by AUX1 in the lateral root cap and epidermal cells [40,41]. AXR4, located in the endoplasmic reticulum, regulates the AUX1 cycle in cells. The accumulation of AUX1 in the endoplasmic reticulum and the loss of its asymmetric distribution in the epidermal cells of the root means that the axr4 mutant shows a weakened gravity response in the root. This suggests that the auxin transport pathway is associated with the gravity signal transduction pathway to regulate the gravity response of root [42-44].

Auxin is exported primarily by members of the PIN-FORMED (PIN) and ABCB/MDR/PCP protein families. The PIN protein family comprises eight members, PIN1 to PIN8. PIN2, PIN3, and PIN7 are directly involved in regulating root gravity response [45]. The asymmetric distribution of these three proteins on the cell membrane results in asymmetric distribution of auxin, which leads to the asymmetric growth of cells and the curved growth of roots [46]. Because of the altered location of PIN2 in the pin2 mutant, auxin cannot be asymmetrically distributed in the root, resulting in the loss of a gravity response in roots [47]. Under gravity stimulation, PIN2 is degraded by the ubiquitin-proteasome system, resulting in the asymmetric distribution of PIN2 protein on the two sides of the root. This, in turn, affects the distribution of auxin [48,49]. In pin3 mutants, the gravitational responses of the roots and hypocotyls of are weakened [50]. Under gravity stimulation, PIN3 protein localizes to the bottoms of CCs along the direction of gravity stimulation, resulting in an asymmetric distribution of auxin on the near and distal sides of the root [50-52]. The function of PIN7 is similar to that of PIN3. A change in the polar localization of PIN7 in cells affects the redistribution of auxin and helps regulate the gravity response [48]. The gravity response is weakened in the roots of the pin7 mutant, and the gravity response of the pin3pin7 double mutant is severely impaired, indicating that PIN3 and PIN7 are functionally redundant in regulating the root gravity response [50,48]. Members of the ABCB/MDR/PCP protein family are also important auxin export carriers and are widely involved in the response of plants to gravity. In the pgp4 mutant, the rate of auxin transport from the root to the base is significantly reduced, and the response of the root to gravity is also weakened. The root gravity responses of pgp1 and pgp19 single mutants and pgp1pgp19 double mutants are enhanced, likely because of decreased auxin polar transport, resulting in enhanced lateral auxin transport. This, in turn, increases the asymmetric distribution of auxin on both sides of the root, thereby enhancing the root gravity response [53,54].

Post-transcriptional modifications, such as phosphorylation and dephosphorylation, regulate the polar localization of PIN proteins and participate in the root gravity response. PID phosphokinase phosphorylates the hydrophilic rings of PIN proteins and is important in their polar localization. The abnormal response to gravity of pid1 roots may be caused by altered PIN localization, which affects the polar transport of auxin [55-57]. Specifically, PID might regulate the gravitational response of roots by affecting the polar localization of PIN2 on the plasma membrane [57]. Altering the phosphorylation site of PIN3 affected the gravity response mediated by this protein [58,59]. Members of the D6PK protein kinase family also regulate the gravity response of roots by altering the phosphorylation state of PIN proteins [60,61].

Effects of Auxin Signaling on the Gravity Response

Under gravity stimulation, the auxin signal transduction pathway specifically induces the differential expression of auxin response genes in the distal and proximal lateral cells of roots. This promotes the asymmetric distribution of auxin and leads to curved root growth.

A mutant of the auxin receptor TIR1 is less sensitive to auxin, and its roots show a weaker response to gravity than the wild type [62]. Mutations of the AUX/IAA gene family members AXR5/ IAA1, SHY2/IAA3, AXR2/IAA7, SLR1/IAA14, AXR/IAA17, IAA20, IAA30, and IAA31 led to the loss or weakening of the response to gravity in Arabidopsis roots and hypocotyls [63]. In addition, a gainof- function mutation of IAA28 led to a weakened gravity response in roots. By contrast, mutation of MSG2/IAA19 led to the loss of the gravity response of hypocotyls, but did not affect the root gravity response [64]. ARF gene mutations lead to abnormal auxin responses, which will affect the plant’s gravity response. Thus, the arf7-1arf1-19, arf10arf16, and msg2-1arf19-1 double mutants all show weakening of the root gravity response [65,66].

Plants can adjust the growth direction according to gravity and 243 ultimately control the structure of stems and roots. Gravitropism is a dynamic process. In this process, gravistimulation induces asymmetric distribution of plant hormone auxin, leading to asymmetric growth and organ bending, and then the auxin distribution returns to the original state before gravistimulation. During the reaction to gravity, the differential accumulation of auxin depends on the activity of polarly localized PIN-FORMED (PIN) auxin-efflux carriers. In particular, the time of this dynamic response is regulated by PIN2, and the MEMBRANE ASSOCIATED KINASE REGULATOR2 (MAKR2) controls the speed of the root to gravity response. During gravitropism, MAKR2 acts as a negative regulator of cell surface signaling mediated by the receptor-like kinase TRANSMEMBRANE KINASE1 (TMK1) and are required for PIN2 asymmetry. In addition, auxin itself can antagonize the inhibitory effect of MAKR2 on TMK1 signaling, thereby triggering the rapid dissociation of MAKR2 membrane in a TMK1 dependent manner. It was shown that the time of root to gravity reaction was coordinated by reversible inhibition of TMK1 signaling pathway on the cell surface [67].

Environmental Factors Regulating Root Gravitation

Effects of Salt Stress on Root Gravity Response

Salt stress alters the response to gravity by controlling the direction of growth of primary roots in Arabidopsis. Root structural morphology is highly plastic during plant growth to allow plants to adapt to various environmental stresses. The structures of roots growing in soil can be adjusted according to the environment, such as high salinity soil, dry soil, and nutrient-deficient soil. In particular, salt stress has strong effects on root structure in Arabidopsis, among other species. Although salt stress is known to inhibit root elongation and reduces root number, many questions remain about the mechanisms underlying the effects of salt stress on root growth. Various experiments suggested that salt stress inhibits root growth by weakening the gravity response. Salt stress causes a sharp decrease in the number of starch grains in root CCs of Arabidopsis. This change is related to strongly altered expression and localization of PIN2 mRNA. Moreover, several mutants of genes in the Salt Overly Sensitive (SOS) regulatory pathway show a significantly reduced gravity response, indicating that salt stress alters both geotropism in roots and the salt balance in plants. SOS pathway mutants also show reduced levels of starch grains in the CCs of root tips and reduced expression of PIN2. These findings suggest that under salt stress, the transcription of PIN2 in Arabidopsis decreases through a process involving SOS pathway proteins, thereby reducing the effects of salt stress on geotropism [68].

Effects of H2O2 on Gravitropism

When plants are stressed by drought [69], low temperature [70], salinity [71], pathogens [72], ozone [73], or heavy metals [74], hydrogen peroxide (H2O2) is rapidly produced in plant cells and released into the cytoplasm. H2O2 is a second messenger involved in many physiological and biochemical reactions in plants, such as stomatal movement induced by Abscisic Acid (ABA), auxin, and Cytokinin (CK) [75,76], root growth and lateral root development [77], and the lignification of cell walls [78]. By applying exogenous H2O2 to cucumber and mung bean [79] discovered that H2O2 interacts with auxin and promotes adventitious root growth. Following the application of different concentrations of exogenous H2O2, the primary roots of pea bent in a non-geotropic manner; the degree of bending increased with increasing concentrations of H2O2. Further analysis revealed an asymmetric distribution of auxin on the inside and outside of the curved primary root. The lateral cells of primary roots that were horizontally curved following H2O2 application were columnar and densely arranged, whereas the inner cells were loosely arranged in filaments and were significantly shorter than the lateral cells. These findings suggest that H2O2 functions downstream of the auxin-induced root geotropism response [77]. Arabidopsis AtCRK5 protein kinase is involved in the establishment of appropriate auxin gradients during many developmental processes. And the Atcrk5-1 mutant exhibited a delayed gravitation response through impaired PIN2 mediated apical auxin transport. This phenotype was associated with lower levels of superoxide anion (O2•-) and hydrogen peroxide (H2O2) compared with wild-type, but higher levels of Nitric Oxide (NO) in the mutant root tip. Paraquat (PQ), an oxidative stress inducer, triggers the formation of O2•- (and H2O2) and can rescue the gravity response of Atcrk5-1 roots. Direct application of H2O2 had the same effect. Under gravity simulation, auxin distribution in the mutant root tip was restored (at least partially restored) by PQ or H2O2 treatment. The redistribution of the PIN2 auxin efflux vector in the heavily simulated PQ treated mutant and untreated wild-type roots was similar. PQ treatment reduced endogenous NO levels in the root tip to normal levels. The mutant phenotype can be restored by directly manipulating endogenous NO levels using no scavengers (cPTIQ). AtCRK5 protein kinase plays an important role in the control of auxin-ROS-NO-PIN2-auxin regulatory loop [80].

Regulation of Gravitropism by Ubiquitin E3 Ligases

SGR9 (SHOOT GRAVITROPISM9) is a C3H2C3-type ring finger protein in Arabidopsis. This protein has ubiquitin E3 ligase activity and is located in amyloplasts, where it regulates amyloplast dynamics [81]. The sgr9 mutant shows a weakened gravity response toward the stem and a slight weakening in the roots. sgr9 amyloplasts do not sediment; instead, they exhibit enhanced mobility. Following site-directed mutagenesis of the conserved RING domain of SGR9, both SGR9W244A and SGR9C232A showed significant decreases in ubiquitin E3 ligase activity, suggesting that the ubiquitin E3 ligase activity of SGR9 in vitro depends on its RING finger domain [81]. Arabidopsis WAV3 is another RING-H2 ubiquitin E3 ligase.

When wild-type Arabidopsis plants were grown on hard agarose medium in tilted plates, the roots showed curved growth owing to the simultaneous action of their own growth driving force, gravity, and resistance to the agar medium. In the wav3 mutant, the root bending angle was larger and showed a smaller half-wavelength distance [82]. The bending angle of wav3 in response to gravity was larger than that in response to light, indicating that the wav3 mutation enhances the root gravity response. WAV3 belongs to a small gene family that also includes EDA40, WAVH1, and WAVH2, and awav3wavh1wavh2 triple mutant showed a strongly altered gravitropic response [82].

Other Signals Influencing Gravitational Responses

Other signals affecting root gravitropism include Ph [83,84], calcium concentration [85,86], potassium ions [87,88], ABA [89], and Reactive Oxygen Species (ROS) [77,90]. When a plant is placed horizontally, the aggregation of auxin on the lower side of the root opens ion channels in the cell membrane. This leads to increased concentrations of intracellular Ca2+, which act as a second messenger to activate the H+/OH-pump in the membrane. OH- is then pumped out of the cell, resulting in an increase in pH at the cell surface on the lower side of the root [91]. The alkaline environment on this side of the root inhibits the extensibility of the cell wall, thereby inhibiting cellular expansion. The imbalance between the expansion of the cells at the top and their inhibited expansion at the bottom ultimately leads to bending of the root.

In contrast to auxin, ABA is a negative regulator of the gravity response. Treatment with 0.5μmol/L ABA significantly reduced gravitropism of the polar auxin transport carrier-related mutants auxl-7 and eirl-l, but the same concentration of ABA had no effect on wild-type plants [89]. The inhibitory effect on the gravity response of Naphthyl 1-Carbamylbenzoate (NPA), an inhibitor of polar auxin transport, was reduced in the ABA biosynthesis mutants abal-l, aba2-4, and aba3-2, suggesting that the role of ABA in inhibiting the gravity response involves the auxin pathway. However, the specific mechanism underlying the role of ABA in the gravity response requires further study. Studies in maize and soybean demonstrated that changes in the direction of gravity caused the accumulation of auxin at the bottom of the root, leading to the accumulation of ROS [77] and Nitric Oxide (NO) [90] in this region. The application of H2O2 on one side of the root caused the root tip to bend in the direction of the H2O2, and the application of sodium nitroprusside, the precursor of NO, to the bottom of the horizontally placed root also promoted the gravitational response of the root. NPA treatment inhibits the synthesis of ROS and NO mediated by auxin, but the exact role of auxin in these processes is unclear. The input and output vectors of auxin are important in the auxin transport pathway in Arabidopsis roots, but they are not the only carriers controlling auxin distribution. The KT/KUP/HAK polygene family is also inextricably related to auxin transport. TRH1 (TINY ROOT HAIR1), a member of this family, encodes a potassium ion transport carrier that affects the formation and growth of plant root hairs. There is a relationship between TRH1 activity and the polar transport of auxin, along with the redistribution of auxin, which directly affects the gravitropism of plant root tips. Therefore, TRH1 is essential for the occurrence and elongation of root hairs and the gravitational growth of roots in Arabidopsis. The transport of auxin from bud growth points to root tips and the efflux of 3H-labeled Indoleacetic Acid (IAA) were significantly inhibited in trh1 roots, whereas in yeast cells over expressing TRH1, the efflux of this radioisotope-labeled auxin was promoted. In trh1, the flow rate of auxin from the root tip columnar cells to the epidermis was significantly inhibited by isotope labeling, which affected the formation and gravitropism of root hairs. This inhibition was relieved by auxin treatment. The gravity sensing, growth, and occurrence and extension of root hairs in the trh1 restorer line trh1/TRH1 were similar to those of the wild type. TRH1-GUS was localized to the columnar cells of the root and the cells on both sides of the root cap, that is, the sensing site of gravity stimulation and the redistribution site of 379 auxin flow. When the K+ concentration in the medium decreased from 20 to 0.1 mmol/L, the gravitropism of the roots disappeared [87,88].

Plant architecture refers to the spatial distribution and morphological characteristics of the aboveground parts of a plant. A plant constantly adjusts its growth direction according to the direction of gravity, ultimately reaching a fixed angle with the gravity direction [92]. An abnormal gravity response of the aboveground parts of a plant will lead to a change in this fixed angle, which is important in shaping plant type [93]. Plant type is one of the most important factors determining crop yields. Therefore, it is important to study the mechanisms underlying the gravity responses of plants to facilitate crop improvement. Due to the increasingly serious pollution caused by industrial activities, burning fossil fuels, improper treatment of sewage sludge, excessive application of fertilizers and the use of fertilizers and pesticides in agriculture, heavy metal pollution such as nickel (Ni) in soil is a major environmental problem. Excessive Ni can cause leaf chlorosis and inhibit plant growth, but its potential growth inhibition mechanism is still unclear. Detailed analysis of root development of Arabidopsis in the presence of Ni showed that this heavy metal induced gravitation defects and locally inhibited root growth by inhibiting cell elongation without significantly damaging the integrity of stem cell niche. Analysis of auxin reactivity reports showed that excessive Ni inhibited the targeted distribution of auxin. PIN2 is very sensitive to Ni because the presence of this heavy metal rapidly reduces PIN2 levels in roots. Ni affects the expression of many genes associated with plant cell walls, and Ni induced transcriptional changes are largely independent of iron (Fe). Excessive Ni increases the accumulation of reactive oxygen species and interferes with the integrity and direction of microtubules [94]. In Arabidopsis gravitropism is affected by two antagonistically interacting proteins, AGD12 (ADP-RIBOSYLATION FACTOR GTPase-ACTIVATING PROTEIN) and EHB1 (ENHANCED BENDING 1). While AGD12 enhances gravitropic bending, EHB1 functions as a negative element [95]. The effect of light and dark on the interaction between tropism is not clear. It has been reported that compared with seedlings completely exposed to light, wild-type Arabidopsis seedlings have higher hydrotropism, their buds are illuminated, but their roots grow in the dark. Due to the reduction of amyloid content in root tip and the reduction of PGM1 (a key starch biosynthesis gene) gene expression, the gravitropism of root system is significantly reduced, which may help to enhance the gravitropism of root system under dark conditions. In addition, the pgm1-415 1 mutant showed greater hydrophilicity compared with the wild type. The amyloplast reaction and starch reduction of Arabidopsis roots occur under light and dark regulation, followed by a decrease in gravitropism and an increase in hydrotropism [79]. The gravitropism of root system is very important for soil anchoring and water and nutrient exploration. It affects the root structure, which is one of the factors affecting crop yield. The mechanism of gravitropism of primary roots has been widely studied, but the regulation mechanism of lateral root gravitropism is still unclear. Topoisomerase I a 423 (TOP1 a) inhibition of lateral root gravitation, contrary to previous reports [96]. TOP1a maintain the gravity of the primary root, revealing a dual function of TOP1a in root gravitropism regulation. The Target of Rapamycin (TOR) is inhibited in lateral root columnar cells, thus inhibiting the development of columnar cells, especially the biosynthesis of amyloplasts. The study revealed a new mechanism of lateral root adjustment to gravity, which may provide theoretical support for improving agricultural production [97]. The emerging role of asymmetric cytokinin signaling as a central anti-gravitropic signal. Differential cytokinin signaling is coopted in gravitropic lateral and hydrotropic primary roots to counterbalance gravitropic root growth [98]. Light and gravity are two key environmental factors controlling plant growth and structure. However, the molecular basis for the coordination of light and gravity signals in plants remains unclear. PHYTOCHROME INTERACTING FACTORS (PIFs) and ELONGATED HYPOCOTYL5 (HY5), can directly bind and activate the expression of LAZY4, a positive regulator of gravitropism in both shoots and roots in Arabidopsis. In the hypocotyl, light promotes the degradation of PIF to reduce the expression of LAZY4, thereby inhibiting the negative gravitropism of the hypocotyl. LAZY4 over expression can partially rescue the negative gravity phenotype of pifq in the dark without affecting the development of amyloplasts. The identification of PIF-LAZY4 regulatory module indicates that PIF protein has another role in gravitropism, and PIF positively regulates amyloid development to promote the negative gravitropism of hypocotyl. In the root, light promotes the accumulation of HY5 protein to activate the expression of LAZY4, thereby promoting the positive gravity in the root. By regulating the protein levels of PIFs and HY5, light exerts an opposite regulation on LAZY4 expression in shoots and roots to inhibit the negative gravitropism of Arabidopsis shoots and promote the positive gravitropism of roots [99]. Arabidopsis NITRATE TRANSPORTER 1/PEPTIDE TRANSPORTER FAMILY 7.3 (NPF7.3) protein functions as a transporter of Indole-3-Butyric Acid (IBA), a precursor of the major endogenous auxin Indole-3-Acetic Acid (IAA). When expressed in yeast, NPF7.3 mediated cellular IBA uptake. Loss-of-function npf7.3 mutants showed defective root gravitropism with reduced IBA levels and auxin responses [100].

References

1. Zhang Y, Ma Y, Zhao D, Tang Z, Zhang T, et al. Genetic regulation of lateral root development. Plant Signal Behav. 2022c; 1: 2081397.
2. Oh E, Seo PJ, Kim J. Signaling Peptides and Receptors Coordinating Plant Root Development. Trends Plant Sci. 2018; 23: 337-51.
3. Kazan K. Auxin and the integration of environmental signals into plant root development. Ann Bot. 2013; 112: 1655-1665.
4. Firn RD, Digby J. Solving the puzzle of gravitropism--has a lost piece been found? Planta. 1997; 203: S159-163.
5. Bailey PH, Currey JD, Fitter AH. The role of root system architecture and root hairs in promoting anchorage against uprooting forces in Allium cepa and root mutants of Arabidopsis thaliana. Journal of experimental botany. 2002; 53: 333-340.
6. Baldwin KL, Strohm AK, Masson PH. Gravity sensing and signal transduction in vascular plant primary roots. American journal of botany. 2013; 100: 126- 142.
7. Blancaflor EB, Masson PH. Plant gravitropism. Unraveling the ups and downs of a complex process. Plant physiology. 2003; 133: 1677-1690.
8. Blancaflor EB, Fasano JM, Gilroy S. Mapping the functional roles of cap cells in the response of Arabidopsis primary roots to gravity. Plant physiology. 1998; 116: 213-222.
9. Tsugeki R, Fedoroff NV. Genetic ablation of root cap cells in Arabidopsis. Proceedings of the National Academy of Sciences of the United States of America. 1999; 96: 12941-12946.
10. Hashiguchi Y, Tasaka M, Morita MT. Mechanism of higher plant gravity sensing. American journal of botany. 2013; 100: 91-100.
11. Ottenschläger I, Wolff P, Wolverton C, Bhalerao RP, Sandberg G, et al. Gravity-regulated differential auxin transport from columella to lateral root cap cells. Proceedings of the National Academy of Sciences of the United States of America. 2003; 100: 2987-2991.
12. Sack FD. Plastids and gravitropic sensing. Planta. 1997; 203: S63-68.
13. Wolverton C, Paya AM, Toska J. Root cap angle and gravitropic response rate are uncoupled in the Arabidopsis pgm-1 mutant. Physiologia plantarum. 2011; 141: 373-382.
14. Staves MP, Wayne R, Leopold AC. Cytochalasin D does not inhibit gravitropism in roots. American journal of botany. 1997; 84: 1530-1535.
15. Baluska F, Hasenstein KH. Root cytoskeleton: its role in perception of and response to gravity. Planta. 1997; 203: S69-78.
16. Mesland DA. Mechanisms of gravity effects on cells: are there gravitysensitive windows? Advances in space biology and medicine. 1992; 2: 211- 228.
17. Taufiq Ur R, Skupin A, Falcke M, Taylor CW. Clustering of InsP3 receptors by InsP3 retunes their regulation by InsP3 and Ca2+. Nature. 2009; 458: 655-659.
18. White PJ. The pathways of calcium movement to the xylem. Journal of experimental botany. 2001; 52: 891-899.
19. Hazak O, Mamon E, Lavy M, Sternberg H, Behera S, et al. A novel Ca2+- binding protein that can rapidly transduce auxin responses during root growth. PLoS biology. 2019; 17: e3000085.
20. Laohavisit A, Shang Z, Rubio L, Cuin TA, Véry AA, et al. Arabidopsis annexin1 mediates the radical-activated plasma membrane Ca²+- and K+- permeable conductance in root cells. Plant cell. 2012; 24: 1522-1533.
21. Masson PH. Root gravitropism. BioEssays : news and reviews in molecular, cellular and developmental biology. 1995; 17: 119-127.
22. Tanaka-Takada N, Kobayashi A, Takahashi H, Kamiya T, Kinoshita T, et al. Plasma Membrane-Associated Ca2+-Binding Protein PCaP1 is Involved in Root Hydrotropism of Arabidopsis thaliana. Plant Cell Physiol. 2019; 60: 1331-1341.
23. Yuan F, Yang H, Xue Y, Kong D, Ye R, et al. OSCA1 mediates osmoticstress- evoked Ca2+ increases vital for osmosensing in Arabidopsis. Nature. 2014; 514: 367-371.
24. Keizer J, Li YX, Stojilkovic S, Rinzel J. InsP3-induced Ca2+ excitability of the endoplasmic reticulum. Mol Biol Cell. 1995; 6: 945-951.
25. Garcia CRS, Alves E, Pereira PHS, Bartlett PJ, Thomas AP, et al. InsP3 Signaling in Apicomplexan Parasites. Curr Top Med Chem. 2017; 17: 2158- 2165.
26. Perera IY, Hung CY, Brady S, Muday GK, Boss WF. A universal role for inositol 1,4,5-trisphosphate-mediated signaling in plant gravitropism. Plant physiology. 2006; 140: 746-760.
27. Roderick HL, Bootman MD. Bi-directional signalling from the InsP3 receptor: regulation by calcium and accessory factors. Biochemical Society transactions. 2003; 31: 950-953.
28. Solovyova N, Fernyhough P, Glazner G, Verkhratsky A. Xestospongin C empties the ER calcium store but does not inhibit InsP3-induced Ca2+ release in cultured dorsal root ganglia neurones. Cell calcium. 2002; 32: 49-52.
29. Monshausen GB, Miller ND, Murphy AS, Gilroy S. Dynamics of auxindependent Ca2+ 635 and pH signaling in root growth revealed by integrating high-resolution imaging with automated computer 636 vision-based analysis. Plant J. 2011; 65: 309-318.
30. Perrin RM, Young LS, Murthy UMN, Harrison BR, Wang Y, et al. Gravity signal transduction in primary roots. Ann Bot. 2005; 96: 737-743.
31. Abel S. Auxin is surfacing. ACS Chem Biol. 2007; 2: 380-384.
32. Baster P, Robert S, Kleine-Vehn J, Vanneste S, Kania U, et al. SCF(TIR1/ AFB)-auxin signalling regulates PIN vacuolar trafficking and auxin fluxes during root gravitropism. EMBO J. 2013; 32: 260-274.
33. Palme K, Nagy F. A new gene for auxin synthesis. Cell. 2008; 133: 31-32.
34. Rakusová H, Gallego-Bartolomé J, Vanstraelen M, Robert HS, Alabadí D, et al. Polarization of PIN3-dependent auxin transport for hypocotyl gravitropic response in Arabidopsis thaliana. Plant J. 2011; 67: 817-826.
35. Rakusová H, Han H, Valošek P, Friml J. Genetic screen for factors mediating PIN polarization in gravistimulated Arabidopsis thaliana hypocotyls. Plant J. 2019; 98: 1048-1059.
36. Gallavotti A. The role of auxin in shaping shoot architecture. J Exp Bot. 2013; 2593-2608.
37. Leyser O. Regulation of shoot branching by auxin. Trends in plant science. 2003; 8: 541-545.
38. Marchant A, Kargul J, May ST, Muller P, Delbarre A, et al. AUX1 regulates root gravitropism in Arabidopsis by facilitating auxin uptake within root apical tissues. EMBO J. 1999; 18: 2066-2073.
39. Swarup R, Friml J, Marchant A, Ljung K, Sandberg G, et al. Localization of the auxin permease AUX1 suggests two functionally distinct hormone transport pathways operate in the Arabidopsis root apex. Genes & development. 2001; 15: 2648-2653.
40. Schoenaers S, Balcerowicz D, Breen G, Hill K, Zdanio M, et al. The Auxin- Regulated CrRLK1L Kinase ERULUS Controls Cell Wall Composition during Root Hair Tip Growth. Current biology. 2018; 28: 722-732.e726.
41. Swarup R, Kramer EM, Perry P, Knox K, Leyser HM, et al. Root gravitropism requires lateral root cap and epidermal cells for transport and response to a mobile auxin signal. Nature cell biology. 2005; 7: 1057-1065.
42. Dharmasiri S, Swarup R, Mockaitis K, Dharmasiri N, Singh SK, et al. AXR4 is required for localization of the auxin influx facilitator AUX1. Science. 2006; 312: 1218-1220.
43. Hobbie LJ. Auxin and cell polarity: the emergence of AXR4. Trends in plant science. 2006; 11: 517-518.
44. Navarro L, Dunoyer P, Jay F, Arnold B, Dharmasiri N, et al. A plant miRNA contributes to antibacterial resistance by repressing auxin signaling. Science. 2006; 312: 436-439.
45. Ganguly A, Park M, Kesawat MS, Cho HT. Functional Analysis of the Hydrophilic Loop in Intracellular Trafficking of Arabidopsis PIN-FORMED Proteins. Plant Cell. 2014; 26: 1570-1585.
46. Wisniewska J, Xu J, Seifertová D, Brewer PB, Ruzicka K, et al. Polar PIN localization directs auxin flow in plants. Science. 2006; 312: 883.
47. Abas L, Benjamins R, Malenica N, Paciorek T, Wisniewska J, et al. Intracellular trafficking and proteolysis of the Arabidopsis auxin-efflux facilitator PIN2 are involved in root gravitropism. Nat Cell Biol. 2006; 8: 249- 256.
48. Kleine-Vehn J, Ding Z, Jones AR, Tasaka M, Morita MT, et al. Gravityinduced PIN transcytosis for polarization of auxin fluxes in gravity-sensing root cells. Proc Natl Acad Sci U S A. 2010; 107: 22344-22349.
49. Kleine-Vehn J, Langowski L, Wisniewska J, Dhonukshe P, Brewer PB, et al. Cellular and molecular requirements for polar PIN targeting and transcytosis in plants. Mol Plant. 2008; 1: 1056-1066.
50. Friml J, Palme K. Polar auxin transport--old questions and new concepts? Plant Mol Biol. 2002; 49: 273-284.
51. Rakusová H, Gallego-Bartolomé J, Vanstraelen M, Robert HS, Alabadí D, et al. Polarization of PIN3-dependent auxin transport for hypocotyl gravitropic response in Arabidopsis thaliana. Plant J. 2011; 817-826.
52. Rakusová H, Han H, Valošek P, Friml J. Genetic screen for factors mediating PIN polarization in gravistimulated Arabidopsis thaliana hypocotyls. Plant J. 2019; 1048-1059.
53. Bouchard R, Bailly A, Blakeslee JJ, Oehring SC, Vincenzetti V, et al. Immunophilin-like TWISTED DWARF1 modulates auxin efflux activities of Arabidopsis P-glycoproteins. J Biol Chem. 2006; 281: 30603- 30612.
54. Cecchetti V, Brunetti P, Napoli N, Fattorini L, Altamura MM, et al. ABCB1 and ABCB19 auxin transporters have synergistic effects on early and late Arabidopsis anther development. J Integr Plant Biol. 2015; 57: 1089-1098.
55. Friml J, Yang X, Michniewicz M, Weijers D, Quint A, et al. A PINOIDdependent binary switch in apical-basal PIN polar targeting directs auxin efflux. Science. 2004; 306: 862-865.
56. He Y, Yan L, Ge C, Yao XF, Han X, et al. PINOID Is Required for Formation of the Stigma and Style in Rice. Plant physiology. 2019; 180: 926-936.
57. Sukumar P, Edwards KS, Rahman A, Delong A, Muday GK. PINOID kinase regulates root gravitropism through modulation of PIN2-dependent basipetal auxin transport in Arabidopsis. Plant physiology. 2009; 150: 722-735.
58. Ganguly A, Cho HT. The phosphorylation code is implicated in cell typespecific trafficking of PIN-FORMEDs. Plant Signal Behav. 2012; 7: 1215- 1218.
59. Ganguly A, Lee SH, Cho HT. Functional identification of the phosphorylation sites of Arabidopsis PIN-FORMED3 for its subcellular localization and biological role. Plant J. 2012; 71: 810-823.
60. Barbosa IC, Zourelidou M, Willige BC, Weller B, Schwechheimer C. D6 PROTEIN KINASE activates auxin transport-dependent growth and PINFORMED phosphorylation at the plasma membrane. Dev Cell. 2014; 29: 674-685.
61. Willige BC, Ahlers S, Zourelidou M, Barbosa IC, Demarsy E, et al. D6PK AGCVIII kinases are required for auxin transport and phototropic hypocotyl bending in Arabidopsis. Plant cell. 2013; 25: 1674-1688.
62. Han H, Rakusová H, Verstraeten I, Zhang Y, Friml J. SCF (TIR1) (/AFB) Auxin Signaling for Bending Termination during Shoot Gravitropism. Plant Physiol. 2020; 183: 37-40.
63. Berleth T, Krogan NT, Scarpella E. Auxin signals--turning genes on and turning cells around. Curr Opin Plant Biol. 2004; 7: 553-563.
64. Tatematsu K, Kumagai S, Muto H, Sato A, Watahiki MK, et al. MASSUGU2 encodes Aux/IAA19, an auxin-regulated protein that functions together with the transcriptional activator NPH4/ARF7 to regulate differential growth responses of hypocotyl and formation of lateral roots in Arabidopsis thaliana. Plant Cell. 2004; 16: 379-393.
65. Muto H, Nagao I, Demura T, Fukuda H, Kinjo M, et al. Fluorescence cross-correlation analyses of the molecular interaction between an Aux/ IAA protein, MSG2/IAA19, and protein-protein interaction domains of auxin response factors of arabidopsis expressed in HeLa cells. Plant Cell Physiol. 2006; 47: 1095-1101.
66. Okushima Y, Overvoorde PJ, Arima K, Alonso JM, Chan A, et al. Functional genomic analysis of the AUXIN RESPONSE FACTOR gene family members in Arabidopsis thaliana: unique and overlapping functions of ARF7 and ARF19. Plant cell. 2005; 17: 444-463.
67. Marquès-Bueno MM, Armengot L, Noack LC, Bareille J, Rodriguez L, et al. Auxin-Regulated Reversible Inhibition of TMK1 Signaling by MAKR2 Modulates the Dynamics of Root Gravitropism. Curr Biol. 2021; 31: 228-37. e10.
68. Sun F, Zhang W, Hu H, Li B, Wang Y, et al. Salt modulates gravity signaling pathway to regulate growth direction of primary roots in Arabidopsis. Plant Physiol. 2008; 146: 178-188.
69. Ramachandra Reddy A, Chaitanya KV, Vivekanandan M. Drought-induced responses of photosynthesis and antioxidant metabolism in higher plants. J Plant Physiol. 2004; 161: 1189-1202.
70. Zhou T, Gui L, Liu M, Li W, Hu P, et al. Transcriptomic responses to low temperature stress in the Nile tilapia, Oreochromis niloticus. Fish Shellfish Immunol. 2019; 84: 1145-1156.
71. Shalata A, Mittova V, Volokita M, Guy M, Tal M. Response of the cultivated tomato and its wild salt-tolerant relative Lycopersicon pennellii to saltdependent oxidative stress: The root antioxidative system. Physiol Plant. 2001; 112: 487-494.
72. Mur LAJ, Kenton P, Draper JH. In planta measurements of oxidative bursts elicited by avirulent and virulent bacterial pathogens suggests that H2O2 is insufficient to elicit cell death in tobacco. Plant Cell & Environment. 2010; 28: 548-561.
73. Evans NH, McAinsh MR, Hetherington AM, Knight MR. ROS perception in Arabidopsis thaliana: the ozone-induced calcium response. Plant J. 2005; 41: 615-626.
74. Romero-Puertas MC, Rodrguez-Serrano M, Corpas FJ, Gomez MD, Sandalio LM. Cadmium-induced subcellular accumulation of O2• and H2O2 in pea leaves. Plant Cell & Environment. 2010; 27.
75. Pei ZM, Murata Y, Benning G, Thomine S, Klüsener B, et al. Calcium channels activated by hydrogen peroxide mediate abscisic acid signalling in guard cells. Nature. 2000; 406: 731-734.
76. Zhang X, Zhang L, Dong F, Gao J, Galbraith DW, et al. Hydrogen peroxide is involved in abscisic acid-induced stomatal closure in Vicia faba. Plant physiology. 2001; 126: 1438-1448.
77. Joo JH, Bae YS, Lee JS. Role of auxin-induced reactive oxygen species in root gravitropism. Plant physiology. 2001; 126: 1055-1060.
78. Potikha TS, Collins CC, Johnson DI, Delmer DP, Levine A. The involvement of hydrogen peroxide in the differentiation of secondary walls in cotton fibers. Plant Physiol. 1999; 119: 849-858.
79. Li Y, Yuan W, Li L, Miao R, Dai H et al. Light-Dark Modulates Root Hydrotropism Associated with Gravitropism by Involving Amyloplast Response in Arabidopsis. Cell Rep. 2020; 32: 108198.
80. Cséplo Á, Zsigmond L, Andrási N, Imran Baba A, Labhane NM, et al, The AtCRK5 Protein Kinase Is Required to Maintain the ROS NO Balance Affecting the PIN2-Mediated Root Gravitropic Response in Arabidopsis. Int J Mol Sci. 2021; 751: 22.
81. Nakamura M, Toyota M, Tasaka M, Morita MT. An Arabidopsis E3 ligase, SHOOT GRAVITROPISM9, modulates the interaction between statoliths and F-actin in gravity sensing. Plant Cell. 2011; 23: 1830-1848.
82. Sakai T, Mochizuki S, Haga K, Uehara Y, Suzuki A, et al. The wavy growth 3 E3 ligase family controls the gravitropic response in Arabidopsis roots. Plant J. 2012; 70: 303-314.
83. Fasano JM, Swanson SJ, Blancaflor EB, Dowd PE, Kao TH, et al. Changes in root cap pH are required for the gravity response of the Arabidopsis root. Plant cell. 2001; 13: 907-921.
84. Scott AC, Allen NS. Changes in cytosolic pH within Arabidopsis root columella cells play a key role in the early signaling pathway for root gravitropism. Plant physiology. 1999; 121: 1291-1298.
85. Lee JS, Mulkey TJ, Evans ML. Inhibition of polar calcium movement and gravitropism in roots treated with auxin-transport inhibitors. Planta. 1984; 160: 536-543.
86. Urbina DC, Silva H, Meisel LA. The Ca2+ pump inhibitor, thapsigargin, inhibits root gravitropism in Arabidopsis thaliana. Biol Res. 2006; 39: 289- 296.
87. Desbrosses G, Josefsson C, Rigas S, Hatzopoulos P, Dolan L. AKT1 and TRH1 are required during root hair elongation in Arabidopsis. J Exp Bot. 2003; 54: 781-788.
88. Vicente-Agullo F, Rigas S, Desbrosses G, Dolan L, Hatzopoulos P, et al. Potassium carrier TRH1 is required for auxin transport in Arabidopsis roots. Plant J. 2004; 40: 523-535.
89. Han W, Rong H, Zhang H, Wang MH. Abscisic acid is a negative regulator of root gravitropism in Arabidopsis thaliana. Biochem Biophys Res Commun. 2009; 378: 695-700.
90. Hu X, Neill SJ, Tang Z, Cai W. Nitric oxide mediates gravitropic bending in soybean roots. Plant physiology. 2005; 137: 663-670.
91. Monshausen GB, Miller ND, Murphy AS, Gilroy S. Dynamics of auxindependent Ca2+ and pH signaling in root growth revealed by integrating high-resolution imaging with automated computer vision-based analysis. Plant J. 2011; 65: 309-318.
92. Digby J, Firn RD. The gravitropic set-point angle (GSA): the identification of an important developmentally controlled variable governing plant architecture. Plant, cell & environment. 1995; 18: 1434-1440.
93. Firn RD, Wagstaff C, Digby J. The use of mutants to probe models of gravitropism. Journal of experimental botany. 2000; 51: 1323-1340.
94. Leškovï A, Zvarï KM, Araya T, Giehl RFH. Nickel Toxicity Targets Cell Wall-Related Processes and PIN2-Mediated Auxin Transport to Inhibit Root Elongation and Gravitropic Responses in Arabidopsis. Plant Cell Physiol. 2020; 61: 519-535.
95. Rath M, Dümmer M, Galland P, Forreiter C. A gravitropic stimulus alters the distribution of EHB1, a negative effector of root gravitropism in Arabidopsis. Plant Direct.2020; 4: e00215.
96. Zhang H, Guo L, Li Y, Zhao D, Liu L, et al. TOP1a fine-tunes TOR-PLT2 to maintain root tip homeostasis in response to sugars. Nat Plants. 2022a; 8: 792-801.
97. Zhang H, Tang Z, Zhang Y, Liu L, Zhao D, et al. TOP1a suppresses lateral root gravitropism in Arabidopsis. Plant Signal Behav. 2022b; 17: 2098646.
98. Waidmann S, Kleine-Vehn J. Asymmetric cytokinin signaling opposes gravitropism in roots. J Integr Plant Biol. 2020; 62: 882-886.
99. Yang P, Wen Q, Yu R, Han X, Deng XW, Chen H. Light modulates the gravitropic responses through organ-specific PIFs and HY5 regulation of LAZY4 expression in Arabidopsis. Proc Natl Acad Sci U S A. 2020; 117: 18840-18848.
100. Watanabe S, Takahashi N, Kanno Y, Suzuki H, Aoi Y, et al. The Arabidopsis NRT1/PTR FAMILY protein NPF7.3/NRT1.5 is an indole-3-butyric acid transporter involved in root gravitropism. Proc Natl Acad Sci US A. 2020; 117: 31500-31509.