Auxin is a key plant hormone that in some way determines all processes of the growth and morphogenesis of plants. In the course of evolution, plants have formed a large number of systems that regulate the concentration of auxin in the organs and tissues at different stages of their development. The investigation of mechanisms of the auxin regulation, function, and the interaction with other regulatory cellular systems allows for understanding many aspects of the
plant growth and development. As a main plant hormone, auxin determines completely different processes, such as gravitropism, phototropism, the formation of organs, tissues, and the root and stem architecture and the development and structure of the conduction system. The root and stem formation in plants is strictly coordinated with each other and requires an intensive exchange of signals between various organs and tissues, as well as the integration of the internal morphogenetic programs with a variety of environmental signals. For example, the development of leaves occurs in concert with the simultaneous development of the lateral roots, which provide the necessary amount of water and mineral substances. Both of these processes are modulated by specific light and soil moisture conditions. In both growing leaves and root tips, the intensive synthesis of auxin and its metabolism occur in this case and, first of all, its accumulation for the future in the form of inactive conjugates in some parts of the plant and its release from the previously formed depot in the other parts. Auxin is then moved to all parts of the plant by the passive or active transport and forms complex auxin gradients in different regions of emerging organs. The effect of auxin on plant growth and development depends primarily on its amount and distribution in organs and tissues. The amount of the hormone is regulated by the intensity of its biosynthesis, conjugation, hydrolysis of the conjugates, and metabolism. The distribution of auxin in organs and tissues of the plant is also of great importance; it is primarily due to the system of the active or polar transport of auxin. Finally, the susceptibility of tissues and cells to the auxin signals plays a significant role for the auxin effects on the plant phenotype. The susceptibility depends on the presence of specific auxin receptors and different components of the signal transmission chain. Thus, the regulation of auxin is implemented on at least four functional levels: the auxin biosynthesis in tissues; its metabolism, especially the formation and
hydrolysis of conjugates that have no auxin activity but play the role of the auxin depot in plant, the active or passive auxin transport, and the receipt and processing of auxin signaling by nuclear protein receptors of auxin.
The tryptophan_independent pathway of auxin biosynthesis functions constantly during the entire vegetation period, which provides the basic level of the hormone concentration, and is nearly unregulated by internal and external factors. This is the most evolutionarily ancient way, since it is present in lower terrestrial plants and mosses. The tryptophan dependent pathways of the auxin synthesis are only switched on at full power mainly at critical moments in life of the plant when they require an increased amount of the hormone, i.e., during embryogenesis, seed germination, flower formation, recovery after various injuries, etc. Maintenance of homeostasis of the IAA concentrations is significantly more typical for biosynthetic pathways with main enzymatic chains in chloroplasts. First of all, these include gene products of the CYP family, which belong to the cytochrome P450 class. The overexpression of these genes leads primarily to the accumulation of the inactive IAA conjugates, but not free IAA, while the enhanced activity of cytoplasmic enzymes (in particular, products of the YUCCA and AMI1 genes) significantly increases the level of free IAA. The higher plants accumulate IAA in the form of inactive IAA conjugates, which can provide the necessary level of free IAA during their hydrolysis at critical
moments of plant development, such as seed germination. Moreover, the conjugates maintain auxin homeostasis in plant tissues. The conjugates are transported from a particular part of the plant, thus providing the necessary concentration of IAA in different organs. Monocotyledonous plants often reserve IAA as conjugates with sugar, whereas dicotyledonous plants form preferably amido conjugates with amino acids and peptides. To date, some genes involved in the formation and hydrolysis of the IAA conjugates have been cloned and characterized. In particular, these are the A. thaliana UGT84B1 gene and its ZmIAGlu ortholog in maize. These genes encode glycosyltransferase, which provides the conjugation of IAA with sugars. The reverse reaction of hydrolysis of these conjugates is catalyzed by IAA Glc hydrolase.
The passive transport does not have its own regulation and fully depends on phloem flow rates. The active, or polar, transport is energy dependent and is performed by three types of protein transporters encoding by the AUX1, PIN, and PGP genes. The protein products of the AUX1 gene have strong polar localization on the cytoplasmic membrane of the upper part of the protophloem cells and actively deliver auxin from the extracellular space into cells, whereas proteins PIN1 and PIN3 are located on the opposite side of the cells and remove auxin to the outside. This localization of these proteins ensures the strong directionality of the active transport of auxin. Proteins of the PGP family do not have the strong polarity of their location on the membrane but like the PIN family proteins, transport auxin from the cell into the extracellular space. In addition, they move auxin through intracellular membranes. Apparently, the PGP proteins are the local regulators of the auxin concentrations in cells and between neighboring protophloem cells. The loss of the AUX1 gene function in plants leads to the appearance of the phenotypes similar to those of auxin resistant plants, including insensitivity to endogenous auxin, impaired growth and branching, and root agravitropism.
Auxin is known to regulate gene expression through degradation of the AUX/IAA proteins, which is a transcriptional repressors, which leads to gene activation. This degradation is dependent on the ubiquitin ligase Skp1-Cullin-F-box (SCF)TIR1 protein complex, where the associated F-box protein TIR1 confers target specificity . In the presence of auxin, the F-box protein TIR1 binds to Aux/IAA, resulting in the ubiquitination and degradation of the Aux/IAA . Auxin enhances TIR1–substrate interactions by acting as a “molecular glue”. F-box protein TIR1 is a true auxin receptor, mediating transcriptional responses to auxin in plants. Each TIR receptor targets specific Aux/IAA proteins for degradation, thus switching on transcription of a multitude of genes, including auxin response factors (ARFs). ARFs regulate a multitude of critical steps in plant development by converting local auxin into gene expression responses. ARFs bind to conserved DNA sequences (TGTCTC) called auxin-response elements (AuxREs) in the promoter regions of primary/early auxin response genes .ARFs can act as either transcriptional activators or repressors depending on the nature of their middle region (MR) domain . The ARFs with a Q-rich MR function as activators, whereas other ARFs with a P/S/T-rich MR function as transcriptional repressors. Thus, AUX/IAA proteins, which repress the activity of ARFs, function as negative or positive regulators of gene transcription. This indicates that the auxin-induced AUX/IAA–SCFTIR1 interaction does not depend on phosphorylation or hydroxylation of AUX/IAA proteins. Auxin stimulates the transcription of a set of genes called primary auxin responsive genes. The known primary genes include three gene families called the AUX/IAA, GH3 and SAUR (small auxin-up RNA) families. In Arabidopsis, the characterization of the auxin resistant mutants axr1 and tir1 led to the discovery that the ubiquitin–proteasome pathway is involved in auxin response . Ubiquitin-mediated protein degradation has emerged as a vital process that regulates the growth and development of eukaryotic organisms. Proteins that are destined to be destroyed are tagged with a polyubiquitin chain by a cascade of reactions involving three enzymes, known as the ubiquitin activating enzyme (E1), ubiquitin conjugating enzyme (E2) and ubiquitin protein ligase (E3). Ubiquitinated proteins are recognized and degraded by the 26S proteasome, a multiprotein complex comprising a 20S core unit and two 19S regulatory particles.
The auxin The auxin signaling pathway. (a) In low auxin conditions, the activity of AUXIN RESPONSE FACTOR (ARF) transcription factors is inhibited by interactions with Auxin/INDOLE-3-ACETIC ACID (Aux/IAAs) repressor proteins and the co-repressor TOPLESS (TPL). (b) Auxin acts a molecular glue between TRANSPORT INHIBITOR RESPONSE1/AUXIN SIGNALING F-BOX PROTEINS (TIR1/AFBs) and Aux/IAAs, leading to ubiquitination and proteasome-mediated degradation of the Aux/IAAs. This relieves repression on the ARFs, allowing auxin-induced transcription to proceed.
ABA is essential for normal plant growth and development. It plays a critical role in different abiotic stresses by regulating various downstream ABA-dependent stress responses. Plants maintain a balance between ABA levels constantly throughout the developmental processes at different tissues and organs, including under unfavorable environmental or physiological conditions. Abiotic stresses trigger ABA biosynthesis, which mediates stress adaptive responses by activating several specific signaling cascades and regulating different physiological and growth-related processes.
ABA which is reported in both primitive and higher organisms seems to have different biosynthesis pathways. In primitive organisms ABA biosynthesis is not well characterized, however, in plant-associated fungi, ABA is reported to be synthesized by the direct cytosolic pathway. In contrast, great progress has been made in identifying and characterizing the genes involved in ABA metabolism in land plants. ABA biosynthesis in plants follows the organelle-specific indirect pathway. The pathway involves the key precursor compound zeaxanthin, which is synthesized by the β-carotene pathway involving pyruvate. Further, zeaxanthin is converted to xanthoxin by the enzymatic reaction catalyzed by ZEP enzyme (zeaxanthin epoxide) and 9-cis-epoxy carotenoid dioxygenase (NCED) enzyme . Subsequently, xanthoxin is transferred from the plastid to cytosol and converted to its aldehyde intermediate and then to ABA by short-chain-dehydrogenase reductase (SDR/ABA2 in Arabidopsis) and abscisic aldehyde oxidase (AAO), respectively. Abiotic stresses and ABA treatment are reported to alter the transcript levels of key ABA biosynthesis genes, which in turn modulate the level of ABA in plants. It is known that ABA accumulates under specific conditions, such as abiotic stresses. Therefore, the endogenous concentration of biologically active ABA at the site of perception has to be regulated. Apart from biosynthesis, ABA catabolism and transport are the two key essential processes that control ABA-mediated stress regulation. Cytochrome P450 type enzymes (CYP707As) catalyze the deactivation reaction resulting in phaseic acid (PA) and dihydro phaseic acid (DPA) as the main ABA catabolites.
ABA metabolic pathway in higher plants: ZEP, zeaxanthin epoxidase; NCED, 9-cisepoxycarotenoid dioxygenase; SDR, short-chain dehydrogenase/reductase; AAO, aldehyde oxidase. CYP707A, abscisic acid-8′-hydroxylase (Leng et al., 2014).
The identification of ABA receptors in Arabidopsis and other plant species is one of the key findings in ABA signaling. The PYR/PYL/RCAR family of proteins are established as the most plausible ABA receptors. Expression profile study of these receptors revealed their role in ABA signaling as well as in the regulation of abiotic stresses. PYR/PYL/RCAR receptors in the presence of ABA form a complex and deactivate PP2C, which otherwise inactivates the SnRK2s, a central regulator of ABA signaling. ABFs belong to basic leucine zipper (bZIP) transcription factor family, which is one of the key regulators of ABA responses in plants. They interact with the cis-acting conserved regulatory element, ABREs (ABA-responsive elements) and in turn regulate transcription of several downstream ABA-responsive genes. The ABA-mediated phosphorylation of ABFs is necessary for their activation. The induction of AREB1/ABF2, AREB2/ABF4 and ABF3 by dehydration, high salinity and ABA treatment and enhanced drought tolerance by plants overexpressing these factors further validates the significance of these proteins and hence ABA in abiotic stress response. Other transcription factors from the MYC, MYB and NAC protein families are also known to function in an ABA-dependent manner. Overexpression of AtMYC2 and AtMYB2 transcription factors, besides exhibiting an ABA-hypersensitive response, also improved osmotic stress tolerance of transgenic plants . Likewise, transgenic plants overexpressing RD26 (a stress-inducible NAC transcription factor) showed high sensitivity to ABA and thus an upregulation of ABA- and stress-responsive genes.
Schematic representation of ABA signaling pathway