28.16: Signaling in Plants

Search Fundamentals of Biochemistry

Learning Goals 

(Learning goals written by Claude, Sonnet 4.6, Anthropic)

Auxin, Gibberellin, and Ubiquitin-Mediated Derepression

• Explain the auxin signaling mechanism as a ubiquitin-mediated derepression cascade: describe how auxin binding to the TIR1 subunit of the SCF E3 ubiquitin ligase (with inositol hexakisphosphate as a cofactor) enables recruitment and ubiquitination of AUX/IAA repressor proteins, their proteasomal degradation, and the consequent release of ARF transcription factors to activate auxin-responsive genes at their AuxRE promoter elements—and explain how the composition of the ARF middle region (proline/Ser/Thr-rich vs. Gln/Leu-rich) determines whether released ARFs activate or repress transcription.
• Draw an explicit mechanistic parallel between auxin and gibberellin signaling: explain how gibberellin binding to the nuclear GID1 receptor promotes interaction with and SCF-mediated ubiquitination of DELLA repressor proteins, leading to their proteasomal degradation and relief of growth repression—and describe the physiological consequences of GA deficiency illustrated by the ga1-3 dwarf mutant phenotype.

Two-Component-Related Signaling: Cytokinins, Ethylene, and Abscisic Acid

• Describe the cytokinin signaling pathway as a eukaryotic adaptation of the bacterial two-component phosphorelay: trace the sequential phosphotransfer events from cytokinin binding at the CHASE domain of the CRE1 histidine kinase → His autophosphorylation in the HK domain → Asp phosphorylation in the receiver-like domain → His phosphorylation on the HPt shuttle protein → final Asp phosphorylation on a type B response regulator that activates auxin-responsive gene transcription—and explain why type A response regulators provide negative feedback.
• Explain the ethylene signaling pathway as an inhibition-of-inhibition cascade: describe how in the absence of ethylene, the ETR1 receptor activates CTR1 kinase, which phosphorylates EIN2 to promote its proteolysis; and how ethylene binding inactivates ETR1, reducing CTR1 activity, stabilizing EIN2, and ultimately activating EIN3/EIL1 transcription factors for ethylene-responsive gene expression.
• Explain the ABA signaling pathway through a double-negative regulatory module: describe how in the absence of ABA, PP2C phosphatases maintain SnRK2 kinases in an inactive dephosphorylated state; how ABA binding to PYL receptors (inducing closure of the CL2 loop) creates a PP2C binding surface that occludes the PP2C active site; and how PP2C inhibition releases SnRK2 kinases for autophosphorylation and subsequent phosphorylation of ion channels (SLAC1, QUAC1, GORK) and transcription factors that drive stomatal closure and ABA-responsive gene expression.

Phytochrome-Mediated Light Signaling

• Describe the structural basis of phytochrome photosensing: explain how the linear tetrapyrrole chromophore phytochromobilin (PΦB) is covalently attached to a conserved Cys in the GAF domain, how red light absorption drives Z-to-E isomerization around the C15–C16 double bond to convert the inactive Pr form to the active Pfr form, how far-red light or thermal reversion regenerates Pr, and how the dimeric nature of phytochromes (producing Pr–Pr, Pfr–Pr, and Pfr–Pfr species) allows graded responses to light quality, quantity, and temperature.
• Trace how active Pfr phytochrome controls transcription factor networks to coordinate seedling development: explain how Pfr directly phosphorylates and promotes proteasomal degradation of PIF transcription factors (via LRB/EBF E3 ligases), how Pfr interacts with SPA to inhibit the COP1/SPA E3 ligase and thereby stabilize the pro-de-etiolation transcription factor HY5, and how in shade conditions (low R:FR ratio) reduced Pfr levels allow PIF accumulation that drives shade avoidance responses including stem elongation and early flowering.

Introduction

Plants are comprised of cells. Hence, they must engage in cell signaling within and between cells. It is beyond the scope of this book to give a detailed description of cell signaling in plants. Instead, we focus on five key classic plant hormones — auxins, cytokinins, ethylene, gibberellins, and abscisic acid — which are produced by leaves, flowers, shoots, roots, or fruit, and examine how they initiate signaling in plants. Finally, we would be remiss if we didn't include the profound signaling in plants initiated by light. Most of this section comes directly from several sources, with modifications and additions (mostly molecular models).

Auxins (3-indolebutyric acid derivatives) are regulators of growth and development and are found in actively growing parts of the plant (roots, shoots, leaves), but mostly in the stem cells. Auxins, for example, facilitate the bending of plants toward the light. They work in conjunction with other hormones, such as cytokinins. When auxins are higher in concentration than cytokinins, roots will form, while the opposite produces shoots. Auxins facilitate the elongation of cells, while cytokinins promote cell division, growth, and wound repair. Gibberellins are also plant growth regulators and facilitate cell elongation. They also help germinate seeds and elongate stems, aid fruit ripening, and promote flowering. Abscisic acid affects seed development and maturation and helps plants tolerate environmental or biotic stresses. It also inhibits growth and metabolism. Ethylene affects fruit ripening, organ abscission, and growth by restricting cell elongation.

We will focus on hormones, their protein receptors, and how the hormone-receptor complex initiates key cellular events.

Auxin

Much of this section derives from Kou et al. Appl. Sci. 2022, 12(3), 1360; https://doi.org/10.3390/app12031360. Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

Auxins, the first plant hormones discovered, regulate plant growth and development. The most common auxin is 3-indole acetic acid. Figure \(\PageIndex{1}\) shows the structures of naturally occurring auxins.

Figure \(\PageIndex{1}\): Structures of naturally occurring auxins.

Figure \(\PageIndex{2}\) shows an interactive iCn3D model of auxin bound to its receptor, TIR1 ubiquitin ligase (2P1Q)

Figure \(\PageIndex{2}\): Auxin bound to its receptor TIR1 ubiquitin ligase (2P1Q) (Copyright; author via source). Click the image for a popup or use this external link: https://structure.ncbi.nlm.nih.gov/i...6xSkEZ3aJA4gT6

Auxin (IAA) is shown in spacefill CPK colors along with an unexpected binding cofactor, inositol hexakisphosphate (IHP), shown in spacefill CPK colors. The peptide shown in light brown sticks is part of the protein Auxin-responsive protein IAA7, a member of the AUX/IAA protein class. These short-lived transcriptional factors function as repressors of early auxin response genes at low auxin concentrations.

The magenta subunit, TIR1 (transport inhibitor response 1), is part of the larger TIR1 complex, the SCF(TIR1) E3 ubiquitin ligase, with only TIR1 shown. Its mere name suggests that it is involved in the ubiquitinylation of a key protein involved in auxin activity, which will be targeted for proteolysis. That protein is the repressor protein IAA7 (an AUX/IAA protein).

Auxin binds in a hydrophobic pocket, which accounts for the binding of the other largely hydrophobic auxins shown in Figure \(\PageIndex{1}\). Note, however, that Arg 401 forms a salt bridge (ion-ion interaction) with the carboxylate of 3-indole acetic acid.

Figure \(\PageIndex{3}\) shows an interactive iCn3D model of auxin bound in the hydrophobic pocket of its receptor, TIR1 ubiquitin ligase (2P1Q).

Figure \(\PageIndex{3}\): Auxin bound in the hydrophobic pocket of its receptor TIR1 ubiquitin ligase (2P1Q) (Copyright; author via source). Click the image for a popup or use this external link: https://structure.ncbi.nlm.nih.gov/i...TFkf6HdMYMiZT9

The green color represents nonpolar side chains. The brown side chains, Trp and Pro, are from the IAA7 peptide of the auxin-responsive protein. To reiterate, the protein IAA7 is a member of the AUX/IAA class of proteins, which represses auxin activity. The IAA7 peptide binds to the auxin in the binding pocket.

Now we can see how auxin regulates gene transcription. First, we must introduce another protein family, the auxin response factors (ARFs). These transcription factors bind to a key DNA sequence, the auxin response element (AuxRE), in the promoters of auxin-activated genes. Once bound, they can either activate or repress transcription from target genes. Auxin binds its receptor TIR1, enabling binding of an AUX/IAA repressor (like IAA7), and then the complex binds ARF. The TIR1 ubiquitin ligase activity of the complex ubiquitinates the bound AUX/IAA repressors (such as IAA7), targeting them for degradation and freeing ARF to regulate gene transcription in the nucleus.

ARFs are structurally similar, with most members containing three regions: DBD (DNA-binding domain), MR (middle region), and PB1 (Phox and Bem 1). Figure \(\PageIndex{4}\) shows a model of how auxin affects ARF transcriptional activity.

Figure \(\PageIndex{4}\): Model of ARF involvement in auxin response. DBD, DNA-binding domain; MR, middle region; PB1, Phox, and Bem 1. Kou et al. Appl. Sci. 2022, 12(3), 1360; https://doi.org/10.3390/app12031360. Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

The AUX/IAA repressor binds to the ARF transcription factor through its PB1 domains at low auxin concentrations. The PB1 (Phox and Bem1) domain is about 80 amino acids in length. It acts as a protein-binding module, enabling heterodimerization or homo-oligomerization with proteins that contain the PB1 domain.

The dimer of AUX/IAA and ARF recruits the co-repressor TPL (TOPLESS) to inhibit the ARF activity and the expression of auxin-responsive genes. When the auxin concentration increases, Aux/IAA binds to the SCF TIR1/AFB complex, is ubiquitinated, and then degraded by the 26S protease. The ARF transcription factors are released to activate the transcription of downstream genes. DBD, DNA-binding domain; MR, middle region; PB1, Phox, and Bem 1.

• At low concentrations of auxin, the AUX/IAA repressor binds to the ARF transcription factor and forms a dimer that recruits the co-repressor TPL (TOPLESS) to inhibit the ARF activity and the expression of auxin-responsive genes;
• When the concentration of auxin is increased, Aux/IAA binds to the auxin:SCF TIR1/AFB complex (remember that the auxin receptor is the TIR1 component of the complex) and is ubiquitinated by TIR1, which is also a ubiquitin ligase;
• The ubiquitinated AUX/IAA protein is degraded by proteolysis by the 26S protease, allowing the ARF to become an active transcription factor.

It appears that the MR domain of ARF determines whether it activates or inhibits transcription. It acts as an inhibitor when rich in proline, serine, and threonine. It acts as an activator when enriched in glutamine and leucine. Some reports show that Aux/IAA and ARFs can form not only dimers but also larger complexes (oligomers), suggesting that oligomerization of Aux/IAA proteins may be essential for inhibiting ARF proteins and that only sufficient amounts of Aux/IAA proteins can exert this inhibitory effect.

Figure \(\PageIndex{5}\) shows an interactive iCn3D model of the DNA binding domain of Arabidopsis thaliana auxin response factor 1 (ARF1) in complex with auxin response element-like sequence ER7 (4LDX)

Figure \(\PageIndex{5}\): DNA binding domain of Arabidopsis thaliana auxin response factor 1 (ARF1) in complex with promoter-like sequence ER7 (4LDX) (Copyright; author via source). Click the image for a popup or use this external link: https://structure.ncbi.nlm.nih.gov/i...9fNHEiFaVANAk6

The ARF protein must translocate to the nucleus to regulate gene transcription. ARF7 and ARF19 have been shown to form micron-sized aggregates in the cytoplasm. These have low responses to auxin. Aggregation occurs through interactions between ARF and PB1 domains and through intrinsically disordered regions. Mutation of a single lysine in the PB1 prevents aggregation and leads to morphological changes in the plant. This shows the importance of regulating transcription and protein translocation to the nucleus.

Figure \(\PageIndex{6}\) reviews the activation of ARFs and some of the genes affected by ARF.

Figure \(\PageIndex{6}\): Auxin signal transduction pathways. Powers and Strader. Up in the air: Untethered Factors of Auxin Response (doi:10.12688/f1000research.7492.1). License CC BY 4.0

Auxin promotes the formation of the TIR1/AFB Auxin/Indole-3-acetic acid inducible (Aux/IAA) co-receptor to promote the ubiquitylation and subsequent degradation of the Aux/IAA repressor. Aux/IAA degradation relieves repression of auxin response factor (ARF) transcription factors, allowing for auxin-responsive gene expression. One of the transcript families upregulated by auxin is the SAUR family. The small auxin up RNA (SAUR) proteins encoded by these transcripts have been suggested to play roles in multiple processes, including interacting with and inhibiting members of the PP2C.D family of phosphatases, which regulate H⁺-ATPase activity. Further, indole-3-butyric acid response 5 (IBR5) and mitogen-activated protein kinase 12 (MPK12) have been implicated in regulating auxin-responsive gene transcription; this regulation is not through destabilization of the Aux/IAA repressors, suggesting a yet-to-be-discovered mechanism of regulating auxin-responsive gene expression. F

Cytokinins (CKs) and Ethylene (ET)

Much of this material derives from Bidon et al. Cells 2020, 9, 2526; doi:10.3390/cells9112526. Creative Commons Attribution (CC BY) license. (http://creativecommons.org/licenses/by/4.0/).

Cytokinins (CKs) and ethylene (ET) are among Earth's most ancient organic chemicals. The structures of a representative cytokinin (kinetin) and ethylene are shown in Figure \(\PageIndex{7}\).

Figure \(\PageIndex{7}\): Structure of kinetin (a cytokinin) and ethylene

Many organisms, including plants, algae, fungi, amoebae, and bacteria, use these substances as signaling molecules to regulate cellular processes. Because of their ancestral origin and ubiquity, CKs and ET are also considered ideal molecules for interkingdom communication. Their signal transduction pathways were first determined in plants and are related to the two-component systems of bacteria (which we explored in a previous section), using histidine kinases as primary sensors.

CKs share a common structure of N⁶-substituted adenine (see Figure \(\PageIndex{7}\)), with biological activities defined by the N⁶-substituents (isoprenoids or aromatic groups). They were originally described as the major hormones regulating cell division, but are also implicated in the control of morphogenesis, embryogenesis, and the inhibition of senescence. Conversely, ET is a simple gas, often referred to as the senescence hormone in plants, that stimulates the senescence of leaves and petals, as well as the ripening of fruits. CK and ET are also well known to orchestrate plant responses to many biotic and abiotic stresses.

Plant signaling pathways are related to the two-component systems typically described in prokaryotes. CKs and ET are perceived by two types of membrane-bound histidine kinase receptors, CRE1 and ETR1, as shown in Figure \(\PageIndex{8}\).

Figure \(\PageIndex{8}\): Perception and transduction of cytokinin (CK) and ethylene (ET) signals in the model plant Arabidopsis.

Panel A shows the cytokinin signaling pathway. CKs in Arabidopsis are primarily recognized by dimerized receptors such as the CRE1 receptor via the cyclase/histidine kinase-associated sensing extracellular (CHASE) domain. CRE1 then auto-phosphorylates (histidine kinase (HK) activity) and immediately transfers its phosphate group to the conserved histidine of a protein belonging to the histidine-containing phosphotransfer (HPt) family. This small protein then acts as a cytoplasm-to-nucleus shuttle and in turn phosphorylates a type B response regulator, which, when activated, positively regulates the transcription of response genes to the CK signal.

Panel (B) shows the ET signaling pathway. Ethylene molecules are detected by ethylene receptors (labeled ETR1) with ethylene binding to the three transmembrane helices (shown in sky blue). ET binding to the dimerized ETR1 receptor downregulates its activity. In the absence of ET, ETR1 activates the serine/threonine kinase CTR1. The CTR1 protein then phosphorylates the EIN2 protein located in the ER membrane, leading to EIN2 proteolysis. In the presence of ET, ETR1 activity is reduced, leading to reduced CTR1 activity; this reduces phosphorylation and accumulation of the EIN2 protein, thereby activating EIN3 and related transcription factors. EIN3 then positively regulates the transcription of ET signal response genes.

Panel (C) shows the domain structure of the Arabidopsis ET (ETR1) and CK (CRE1) receptors.

Mechanistically, the two pathways use fundamentally different families of downstream modules.

It is now known that bacteria also use CK and ET signaling, as described in Figure \(\PageIndex{9}\).

Figure 2.

Let's look in more detail at CRE1, cytokinin response 1, the main cytokinin receptor in plants. Different computational programs often show different domain structures. Figure \(\PageIndex{10}\) shows the domain structure determined by Pfam.

Figure \(\PageIndex{10}\): Domain structure of cytokinin response 1, the main cytokinin receptor in plants.

Uniprot describes this domain structure, color-coded as in Figure \(\PageIndex{11}\)

• 131-149: transmembrane
• 200-382: Green Chase (Cyclases/Histidine kinases Associated Sensory Extracellular)
• 420-443 transmembrane
• 472-537: Red His Kinase A Phosphoaccepter domain
• 584-760 Blue HK kinase, DNAgyrase, HSP-like ATPase
• 786-920: Yellow Reg REsp 1
• 946-1071: Yellow Reg Regulator receiver domain

Figure \(\PageIndex{11}\) shows an interactive iCn3D model of Histidine kinase 4 - Cytokinin receptor 1 (CRE) from Arabidopsis thaliana (AlphaFold model - Q9C5U0). The coloring matches the Pfam domains in Figure \(\PageIndex{10}\).

Figure \(\PageIndex{11}\): Histidine kinase 4 - Cytokinin receptor 1 (CRE) from Arabidopsis thaliana (AlphaFold model) - Pfam Domain organization (Q9C5U0) (Copyright; author via source). Click the image for a popup or use this external link: https://structure.ncbi.nlm.nih.gov/i...jq7BTpshpS3d46

Figure \(\PageIndex{12}\) shows an interactive iCn3D model of Histidine kinase 4 - Cytokinin receptor 1 (CRE) from Arabidopsis thaliana (AlphaFold model) - Domain organization (Q9C5U0) that clearly shows the extracellular and intracellular domains.

Figure \(\PageIndex{12}\): Histidine kinase 4 - Cytokinin receptor 1 (CRE) from Arabidopsis thaliana (AlphaFold model) - Domain organization (Q9C5U0) (Copyright; author via source). Click the image for a popup or use this external link: https://structure.ncbi.nlm.nih.gov/i...2Q9WsRkZk75ZZ8

The N-terminal methionine is in cyan spacefill and the C-terminal Ser is in spacefill. Two transmembrane helices are shown in dark gray spacefill (125-145) and light gray spacefill (430-450). These connect the extracellular domain (cyan, 146-429) and the two cytoplasmic domains (magenta 1-124, which is mostly disordered in the model, and 451-1080). This model does not reflect the protein's relative disposition in the actual structure, but it clearly shows the extracellular and cytoplasmic domains. The CHASE domain is the extracellular domain (cyan, 146-429).

Here are the steps involved in cytokinin signaling through its receptor (shown in Figure \(\PageIndex{8}\) :

• the cytokinin binds to the CHASE domain
• the receptor autophosphorylates a His in the HK domain
• a phosphotransfer from the pHis to an Asp in the Yellow Reg (Regulator) Receiver domain
• a phosphotransfer from pAsp to the His in the histidine-containing phosphotransfer protein (HPt)
• a final transfer from pHis to an Asp in a response regulator (RR)

The MAPK cascade is activated in the cytokinin signaling pathway. Phosphorylated pRR can also regulate target gene transcription. Type-A RRs are negative regulators of cytokinin signaling. It also acts with phytochromes (discussed at the end of this section) to regulate red light signaling. Cytokinin receptors can bind synthetic chemicals that act as defoliants and herbicides.

Intermolecular interactions in the cytokinin signaling pathway leading to transcriptional effects are illustrated in Figure \(\PageIndex{13}\).

Figure \(\PageIndex{13}\): Intermolecular interactions in the cytokinin signaling pathway. CK, cytokinin; Ade, adenine; R, ribose; P, phosphate; D, conserved aspartate; H, conserved histidine; DI, dimerization interface domain of the sensor module; PAS and PAS-like are subdomains of the CHASE domain of the sensor module; TM1 and TM2, transmembrane domains; HisKA (DHpD), histidine kinase A domain (dimerization and histidine phosphotransfer domain); HATPase (CAD), adenosine triphosphatase domain (catalytic and ATP-binding domain); REC-like, receiver-like domain; REC, receiver domain; HPt, histidine-containing phosphotransfer protein (phosphotransmitter); RR-B, type B response regulator (transcription factor). Protein–protein interactions (PPI) are indicated by a red dotted line. Arkhipov et al. Int. J. Mol. Sci. 2019, 20(9), 2096; https://doi.org/10.3390/ijms20092096Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).

Gibberellin

Much of this material derives from Hedden, P., Sponsel, V. A Century of Gibberellin Research. J Plant Growth Regul 34, 740–760 (2015). https://doi.org/10.1007/s00344-015-9546-1. https://doi.org/10.1007/s00344-015-9546-1. Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/)

Gibberellins control growth and development pathways in plants and fungi. They act in plants by removing growth limitation by promoting the degradation of the growth-inhibiting DELLA proteins, which contain the Asp-Glu-Leu-Leu-Ala (DELLA) motif. The name gibberellin derives from the fungus Gibberella fujikuroi. There are many types of gibberellins, all of which are diterpenoids. The structures of the main bioactive GAs in plants, GA₁ and GA₄, are shown below in Figure \(\PageIndex{14}\).

Figure \(\PageIndex{14}\): Structures of gibberellin A1 and A4

Gibberellin initiates signaling by binding to the nuclear gibberellin receptor. One such receptor is the Gibberellin Insensitive Dwarf1 (GID1). When bound, it triggers the proteolysis of another protein bound to it, called a DELLA protein (an example is GAI), a transcriptional regulator that inhibits growth. Gibberellins' control of transcriptional activity is reminiscent of that of auxins.  RGA (repressor of ga1-3) and GAI (gibberellin insensitive) are negative regulators of gibberellin (GA).

Figure \(\PageIndex{15}\) shows an interactive iCn3D model of the gibberellin(GA3)- active gibberellin receptor GID1L1 bound to the DELLA domain of GAI (2ZSH).

Figure \(\PageIndex{15}\): gibberellin(GA3)- Active gibberellin receptor GID1L1 bound to the DELLA domain of GAI (2ZSH). (Copyright; author via source). Click the image for a popup or use this external link: https://structure.ncbi.nlm.nih.gov/i...jVpzjnbUoQMhz5

The gibberellin receptor is gray and associated with the membrane's inner leaflet (blue). The plant hormone gibberellin A3 is bound to the receptor. Amino acid side chains in the receptor involved in interactions with gibberellin A3 are shown as sticks, colored in CPK. The DELLA domain of GAI is shown in cyan. The three amino acid motifs within it (DELLA, cyan spacefill; and VHYNP and LExLE, both magenta spacefill) are also shown. As with the auxin receptor, GID11A binds gibberellin in a deep pocket, which is covered by an N-terminal helix of the receptor. That helix recognizes and binds to the DELLA sequence of the DELLA transcription regulator protein.

Figure \(\PageIndex{16}\) shows the effects of a mutation that leads to deficiencies in gibberellin 1-3 (right-hand side).

Figure \(\PageIndex{16}\): Physiological action of GA as illustrated by a comparison of the Landsberg erecta Arabidopsis plant with a GA-deficient mutant (ga1-3). In the absence of a GA, response stem elongation, leaf enlargement, floral development, seed set, and fruit development do not occur.

The mechanism by which GAs promote growth is shown in Figure \(\PageIndex{17}\).

Figure \(\PageIndex{17}\): Representation of GA perception and signal transduction.

The binding of bioactive GA induces a conformational change in the GID1 receptor, thereby promoting its interaction with DELLA proteins. Recruitment of an F-box protein initiates ubiquitination of DELLA by an SCF E3 ubiquitin ligase, targeting the DELLA for proteasomal degradation. Loss of DELLA relieves growth repression and suppresses other DELLA-mediated responses.

Abscisic acid

Much of this material derives from Hewage et al. (2020). Advanced Science. https://doi.org/10.1002/advs.202001265. This is an open-access article under the terms of the Creative Commons Attribution License.

The phytohormone abscisic acid (ABA) is the best-known stress signaling molecule in plants. As such, it will be crucial for plants to adapt to climate change. Its structure is shown in Figure \(\PageIndex{18}\).

Figure \(\PageIndex{18}\): Structure of abscisic (ABA)

ABA protects land plants from biotic and abiotic stresses. ABA receptor proteins (PYLs) contain a conserved pyrabactin resistance/pyrabactin resistance-like/regulatory domain (PYR/PYL/RCAR) that binds ABA and triggers signaling events.

ABA has significant roles throughout a plant's life cycle. From the single-celled zygotic stage to the mature multicellular plant, plant development involves ABA. ABA allows germination only under optimum conditions and inhibits growth under stress conditions. The adult plant and the seedling experience biotic and abiotic stressors that vary in severity and persistence. ABA enables the plant to survive by inducing short- and long-term stress responses, including rapid, reversible stomatal closure, long-term growth inhibition, dormancy, senescence, and abscission. ABA is therefore both a developmental and a stress-signaling molecule with diverse roles, as shown in Figure \(\PageIndex{19}\).

Figure \(\PageIndex{19}\): Summary of ABA involvement in development and stress tolerance.  Hewage et al. ibid.

ABA signaling in drought

Let's look at a specific example of ABA signaling under drought stress, a stressor that will intensify as the world's climate changes due to the combustion of fossil fuels.

Insufficient soil water levels can result in an imbalance of water between the cells and the outer environment. Changes in cellular electrolyte content affect metabolism, leading to osmotic imbalance or stress. The osmotic stress thus leads to the accumulation of ABA in cells and triggers ABA signaling. The cellular pool of ABA is dramatically increased during drought. Biosynthesis, catabolism, conjugation, and transportation of ABA are coordinated to increase ABA levels. ABA rapidly regulates plant water levels by controlling stomata. Stomata, microscopic pores controlled by two highly differentiated epidermal cells (guard cells), primarily regulate gas exchange between air and plants. Open stomata allow CO₂ to diffuse into the leaf mesophyll, reaching the sites of photosynthesis. They also allow water vapor to exit the plant interior to the atmosphere.

By allowing transpirational water loss, stomata help cool the plant and regulate internal water levels. Thus, stomata are essential regulators that connect the plant interior to the outside environment. Increases in osmotic pressure in guard cells lead to water uptake and cell expansion; as the cells expand, the pore opens due to differential thickening of the guard cell walls. Stomatal movements are regulated by numerous environmental signals, including light, plant growth regulators, pathogens, drought, cold, and nutrient status. Stomatal movement is the quickest response to ABA signaling. Therefore, core ABA signaling is essential for guard cell function. The involvement of ABA signaling events in guard cell function is summarized in Figure \(\PageIndex{20}\).

Figure \(\PageIndex{20}\): Simplified representation of ABA-mediated core signaling events in guard cells. Hewage et al. ibid

ABA is involved in seed maturation, dormancy, and germination. During seed development, ABA contributes to the accumulation of stored reserves and desiccation tolerance, maintains seed dormancy, and regulates the expression of late embryogenesis abundance (LEA) genes.

Without ABA, (left) ABA receptors (PYLs) are in ligand-free form. H⁺ATPase pumps H⁺ ions outside of the plasma membrane. The SnRK2 protein kinases and the S-type anion channel SLAC1 are kept dephosphorylated by PP2Cs. The dephosphorylation state of SLAC1 prevents the nonspecific activation of S-type anion channels.

In the presence of ABA (right), PYLs bind to and inhibit PP2Cs. ABA inhibits H⁺ATPase activity, blocking pumping of H⁺ to the outside. The Ca²⁺-independent protein kinases (SnRK2s) are released from PP2C inhibition and activated by auto-phosphorylation. Ca2+-permeable cation (ICa) channels are released from PP2C-mediated inhibition, causing an increase in ABA-responsive Ca²⁺ in the cytosol and activating CPKs. The activated SnRK2s and CPKs phosphorylate SLAC1. The SnRK2.6/OST1 protein kinase phosphorylates and activates the R-type anion channel ALMT12/QUAC1. K⁺ ions are effluxed via the voltage-dependent outward K+ (K+out) channel GORK, decreasing guard cell turgor and leading to stomatal closure. PYLs: ABA receptors; ABA: abscisic acid; PP2C: protein phosphatase 2C proteins; OST1: open stomata 1/SnRK 2.6 protein kinase; Ca/CPK: Ca^2+/calcium dependent protein kinases; I_Ca2+: plasma membrane nonselective cation channel permeable to Ca²⁺ SLAC1: slow anion channel-associated 1 (SLAC1); QUAC: aluminum-activated malate transporter 12/quickly activating anion channel 1 (ALMT12/QUAC1); GORK: guard cell outward rectifying K⁺ channel (GORK); KAT1: K⁺ activated 1 potassium ion channel; A−: anions; K⁺: potassium ions.

The ABA receptor core complex

ABA signaling has three main phases: ABA synthesis/metabolism, long-distance transport, and ABA binding to its receptor. Downstream signaling ensues through transcriptional activators/repressors and plasma membrane-located channel proteins. Figure \(\PageIndex{21}\) shows the core ABA signaling pathway's main components.

Figure \(\PageIndex{21}\): Core ABA signaling pathway components.  Hewage et al. ibid.

Without ABA (A above), SnRK2 kinases are dephosphorylated by protein phosphatase 2C (PP2Cs). In the presence of ABA (B above), PP2Cs are inhibited by the complexes PYLs-ABA. Thus, the SnRK2 kinases are released and initiate a cascade of downstream transcription factors, NADPH transporters, and ion channels. These phosphorylate the transcription factors, inducing ABA-responsive gene transcription. Additionally, ion channels act on guard cells to regulate transpiration.

ABA receptors (PYLs) bind ABA and PP2Cs and are phosphorylated by protein kinases. The ABA:PYL complex binds PP2Cs, inducing conformational changes in their active sites that inhibit phosphatase activity. This leads to the release of downstream protein kinases (SnRK2s) from PP2C-mediated inhibition. The SnRK2s undergo autophosphorylation, thereby activating a series of ion channels, NADPH oxidases, and transcription factors. This activates short-term and long-term ABA responses such as stomatal closure and upregulation of ABA-dependent gene expression. MAPKKKs (MAPK3s) also activate SnRK2.6 by phosphorylating a specific site during salinity stress.

ABA binding regulates a double-negative regulatory system in which the ABA receptors (PYLs) act as negative regulators, PP2Cs act as negative regulatory coreceptors, and SnRK2s act as negative regulators. In addition to the regulation by SnRK2 and PP2Cs, several post-translational modifications also regulate ABA signaling. Phosphorylation, dephosphorylation, ubiquitination, farnesylation, and sumoylation have been found to modulate ABA signaling by targeting core components (PYLs or PP2Cs) or downstream interacting proteins.

ABA Receptors (PYLs)

PYLs are soluble proteins, and among the 14 PYLs in Arabidopsis, 13 function as ABA receptors. All PYLs are known to share a dominant helix-grip structure. This characteristic motif consists of a seven-stranded antiparallel β-pleated sheet flanked by two α helices. The β-pleated sheets enfold a long carboxy-terminal α-helix of PYLs. The apo-PYLs contain a sufficiently large hydrophobic pocket of 543˚A between the C-terminal helix and β-sheet. The size of this pocket is estimated at 480 Å in the ABA-bound state. The 23-pocket residues are highly conserved and more hydrophobic than other parts of PYLs. The interactions of ABA and PYL2 are shown in Figure \(\PageIndex{22}\).

Figure \(\PageIndex{22}\): The binding mode of ABA and PYL2 (PDBID: 3KDI) in A) 3D and B) 2D (redrawn). In the 3D structure, the PYL2 cartoon is colored white (A). The important residues and ABA are shown as blue and yellow sticks, respectively. The H-bonds are marked with red dotted lines.

Figure \(\PageIndex{23}\) shows an interactive iCn3D model of Abscisic acid bound to the Abscisic Acid Receptor (PYL2) (pdbid: 3KDI)

Figure \(\PageIndex{23}\): Abscisic acid bound to the Abscisic Acid Receptor (PYL2) (pdbid: 3KDI) (Copyright; author via source). Click the image for a popup or use this external link: https://structure.ncbi.nlm.nih.gov/i...uToGsSPpnZfPeA

In the absence of ABA, the apo-PYL2 has a pocket surrounded by four surface loops. When ABA binds, one loop (CL2) closes onto the pocket, forming a PP2C binding site for the phosphatases ABI1 and ABI2. This blocks the phosphatase's active site.

Figure \(\PageIndex{24}\) shows an interactive iCn3D model of ABA-bound PYL1 and the Protein Phosphatase 2C ABI1 (pdbid 3kdj)

Figure \(\PageIndex{24}\): ABA-bound PYL1 and the Protein Phosphatase 2C ABI1 (pdbid 3kdj) (Copyright; author via source). Click the image for a popup or use this external link: https://structure.ncbi.nlm.nih.gov/i...tQgyC7NVXk1UVA

The PP2C phosphatase (ABI1) is gray, with its active site highlighted in green spacefill. The ABA receptor PYL1 is shown in cyan, and ABA is shown in sticks and colored CPK. The CL2 loop of the ABA-bound PYL1 receptor is shown in red spacefill. It projects into the PP2C active site, inhibiting its activity.

Light Signaling through Phytochromes

Much of this material derives from Liu, Y., Jafari, F. & Wang, H. Integration of light and hormone signaling pathways in the regulation of plant shade avoidance syndrome. aBIOTECH 2, 131–145 (2021). https://doi.org/10.1007/s42994-021-00038-1. Creative Commons Attribution 4.0 International License. http://creativecommons.org/licenses/by/4.0/.

Plants compete with other plants in regions of high plant density by sensing changes in light intensity and wavelength. Signaling leads to responses (stem elongation, reduced branching, early flowering, etc) called shade avoidance syndrome (SAS). A photosensory system initiates signaling that alters gene transcription. In the SAS in plants in a large canopy, the upper leaves use red and blue light for photosynthesis. Multiple photoreceptors are used. Some transcription factors are also sensitive to light. For example, PIF3, a transcription factor, binds to light-responsive genes only when it binds to another transcription factor called Pr. Pr is resident in the cytoplasm but moves to the nucleus after altering conformation on absorbing red light.

Legris, M., Ince, Y.Ç. & Fankhauser, C. Molecular mechanisms underlying phytochrome-controlled morphogenesis in plants. Nat Commun 10, 5219 (2019). https://doi.org/10.1038/s41467-019-13045-0. Creative Commons Attribution 4.0 International License. http://creativecommons.org/licenses/by/4.0/. do

Phytochromes exist in bacteria, cyanobacteria, fungi, algae, and land plants. We will focus mostly on phytochromes in Arabidopsis. In land plants, phytochromes are red and far-red light receptors that exist in two forms. They are synthesized in the inactive Pr state, which converts to the active Pfr conformation upon light absorption. Pfr is inactivated upon far-red (FR) light absorption or through thermal relaxation, which depends on temperature. Phytochromes act as dimers, producing three possible phytochrome species: Pr–Pr, Pfr–Pr, and Pfr–Pfr. Pr and Pfr have different absorption maxima, but due to overlapping spectra, both conformers are always present in the light, while only prolonged darkness returns all phytochrome to Pr. Given that phytochrome responses depend on the proportion of Pfr conformers, signaling is influenced by a combination of light quantity, color, and temperature. These features of phytochromes are summarized in Figure  \(\PageIndex{25}\).

Figure \(\PageIndex{25}\): Control of phytochrome activity.

Panel A shows factors that control phytochrome activity. Phytochromes exist in two conformations, Pr and Pfr, the latter being the active form. They exist as dimers, so three species are present. Each monomer can be activated by red light (R) and inactivated by far-red light (FR) or by thermal reversion, a process that depends on temperature (T). At least in the case of phyB, Pfr in heterodimers reverts much faster than in homodimers, allowing phyB to perceive temperature both during the day and at night.

Panel B shows plant phytochrome absorption spectra of the Pr and Pfr conformations. In dark-adapted seedlings, phytochromes are in the Pr form. Due to overlapping Pr and Pfr absorption spectra, only 87% of Pfr is achieved upon a saturating R pulse.

Panel C shows action spectra for phyA and phyB in the control of hypocotyl elongation. Fluence rate (number of particles passing per unit time) response curves are measured at different wavelengths, and the fluence rate (total radiant power or energy from all directions passing through an infinitesimally small sphere at a given point, divided by the cross-sectional area of that sphere) yielding 40% inhibition relative to the dark control is determined. To specifically determine action spectra for phyA and phyB, the curve was performed with phyB-GFP/phyAphyB seedlings, and for phyA, using phyB-5 seedlings. Values are relative to the response obtained at the most efficient wavelength in each case.

Plant phytochrome structure

Plant phytochromes are dimeric, each monomer consisting of ~1150 amino acids. The chromophore, a linear tetrapyrrole named phytochromobilin (PΦB), whose structure is shown in Figure \(\PageIndex{26}\), is attached to the protein.

Figure \(\PageIndex{26}\): Structure of phytochromobilin (PΦB)

The domain structure of phytochromes is shown in Figure \(\PageIndex{27}\).

Figure \(\PageIndex{27}\): Structural domains of phytochromes and their role in perceiving environmental signals and downstream signaling. NTE N-terminal extension, PSM photosensory module, CTM C-terminal module.

The apoprotein can be divided into the N-terminal PSM, which consists of the N-terminal extension (NTE), for which structural information remains scarce, and three structurally related domains: Period/Arnt/SIM (PAS), cGMP phosphodiesterase/adenylyl cyclase/FhlA (GAF), and a phytochrome-specific domain (PHY) and a C-terminal module (CTM) comprising two PAS domains and a histidine kinase-related domain (HKRD). The chromophore is bound covalently to a conserved cysteine in the GAF domain, which has intrinsic chromophore lyase activity. Light perception triggers a Z to E isomerization around the C15–C16 double bond of PΦB, which leads to a cascade of structural modifications in the protein. Figure \(\PageIndex{28}\) shows an interactive iCn3D model of phytochrome (Deinococcus) Pfr form in the Photoactivated State (5C5K)

Figure \(\PageIndex{28}\): Phytochrome (Deinococcus) Pfr form in the Photoactivated State (5C5K) (Copyright; author via source). Click the image for a popup or use this external link: https://structure.ncbi.nlm.nih.gov/i...VzE7AqtgejbfeA

The protein is shown in its active dimeric state. One chain is shown in secondary structure colors, and the other is cyan. The chromophores (heme derivatives) are shown in spacefill in both subunits. Side chains surrounding the chromophore are shown in colored sticks in the cyan chain.

At first glance, the presence of the histidine kinase-related domain (HKRD) suggests that phytochromes transduce their signal through the C-terminal module (CTM). Although many bacterial and cyanobacterial phytochromes have a C-terminal histidine kinase domain and act as light-regulated histidine kinases, plant phytochromes are not histidine kinases, and their role as Ser/Thr kinases remains contentious. The photosensory module (PSM) fused to a nuclear localization signal and a dimerization sequence is sufficient to restore most phyB functions, pointing to key signaling functions of the PSM.

The major functions of the plant phytochrome CTM are dimerization, nuclear import, and localization to sub-nuclear structures known as photobodies. However, it was recently shown that the C-terminal region of phyB also engages in light-regulated interactions and regulates PIF activity. Moreover, the activity of the CTM is controlled by post-translational SUMOylation, which limits the ability of active phyB to interact with downstream signaling targets, thereby attenuating light responses. In addition, the CTM modulates active (Pfr) phytochrome levels with the HKRD inhibiting the Pr–Pfr photoconversion while the PAS–PAS promotes thermal reversion. Hence, while dividing plant phytochromes into PSM and CTM helps describe the molecule, both parts of the photoreceptor contribute to regulating active Pfr levels and downstream signaling activities.

Figure \(\PageIndex{29}\) shows a simplified mechanism for phytochrome control of transcription factors in different light environments.

Figure \(\PageIndex{29}\): Simplified mechanism for phytochrome control of transcription factors in different light environments.

Panel a shows the response below the soil surface during growth in partial or complete absence of light (called etiolated growth). For simplicity, we consider that phytochromes remain inactive (Pr) below the soil surface, accumulating transcription factors such as PIFs, EIN3, and ARFs, and subsequently inducing etiolation and auxin response genes. The COP1/SPA ubiquitin E3 ligase accumulates in the dark. It leads to proteasome-mediated degradation of HY5, a transcription factor that suppresses the expression of genes required for etiolation and induces expression of genes required for de-etiolation.

Pane b shows changes that occur when light intensity increases (de-etiolation). Light perception activates phytochromes (Pfr), which promote de-etiolation by directly inhibiting PIFs and EIN3, and indirectly inhibiting ARFs by stabilizing Aux/IAA proteins. The Pfr form of either phyA or phyB interacts with SPA proteins, inhibiting COP1/SPA. This stabilizes HY5, inducing de-etiolation-related gene expression and repressing etiolation genes.

Pane c shows de-etiolated plant in response to shade (reduced R/FR). Low R/FR in shade reduces the fraction of active phytochrome (Pfr/Ptot). PIFs accumulate and induce growth-promoting gene expression. In addition, PIFs induce a negative feedback loop exemplified by HFR1 expression. HFR1 (and other HLH proteins) bind to PIFs, forming non-DNA-binding heterodimers. COP1/SPA is also involved in this loop by leading HFR1 to proteasome-mediated degradation. Arrows indicate positive regulation, blunt-ended arrows indicate negative regulation, and dotted-lined arrows indicate nucleo-cytoplasmatic relocalization.

Figure \(\PageIndex{30}\) summarizes how phytochromes affect transcription.

Figure \(\PageIndex{30}\): Mechanisms of phytochrome-mediated regulation of transcription factors.

Panel a (top to bottom) shows the sequential steps by which Pfr inhibits PIFs. Top: PfrA interacts with PIF1 and PIF3, while PfrB interacts with PIF1–PIF8. Middle left: for PIF1, 3, and 4, phytochrome inhibits DNA binding. Middle right: Interaction with Pfr leads to the phosphorylation of PIFs. Many kinases have been found to phosphorylate PIFs (see text) with PPKs phosphorylating PIFs in response to light. Bottom: after light-induced phosphorylation, PIF3 is degraded by LRBs and EBFs, with phyB co-degradation occurring in the LRB-mediated process (left, center). Phosphorylated PIF7 interacts with 14-3-3 proteins and remains in the cytoplasm (right).

Panel b shows other mechanisms of transcriptional control by phytochromes. Left: PfrA and PfrB interact with SPA, inhibiting the COP1/SPA complex. Center: PfrB interacts with EIN3 to promote ERF-mediated EIN3 degradation. Right: PfrA and PfrB interact with Aux/IAA to prevent their degradation by SCF^TIR1/AFB.

Panel c shows Patterns of PIF abundance across developmental states and growth conditions. In etiolated seedlings, PIFs accumulate to high levels, promoting etiolated growth. Upon light exposure, PIFs are rapidly degraded in a phytochrome-dependent manner, with half-lives of ~5 min for PIF1 and PIF5, and ~10 min for PIF3 and PIF4 (left). In contrast, in light-grown seedlings, PIFs are under strong transcriptional control, allowing accumulation of the protein even in conditions when phytochrome activity is predicted to be high (right), SD (short days), LD (long days)

Summary

(Summary written by Claude, Sonnet 4.6, Anthropic)

Plant hormone signaling illustrates several recurring biochemical strategies—ubiquitin-mediated proteolysis, two-component phosphorelays, and double-negative regulatory cascades—repurposed and elaborated for the specific demands of sessile organisms that must integrate developmental cues with environmental stresses.

Auxin and gibberellin signaling operate through structurally analogous derepression mechanisms. In both cases, a hormone-bound intracellular receptor recruits a repressor protein (AUX/IAA for auxin, DELLA for gibberellin) to an SCF E3 ubiquitin ligase complex, targeting the repressor for proteasomal degradation. Auxin binding to TIR1—with inositol hexakisphosphate acting as an unexpected cofactor—enables the hydrophobic pocket to simultaneously accommodate the IAA carboxylate (anchored by Arg401) and recruit the AUX/IAA peptide, which is then ubiquitinated and degraded. This frees ARF transcription factors, whose activation versus repression activity is determined by the amino acid composition of their middle region, to bind auxin response elements (AuxREs) and regulate genes involved in cell elongation, lateral root formation, and tropisms. Gibberellin binding to the nuclear GID1 receptor induces a lid conformational change that recruits DELLA repressors for SCF-mediated ubiquitination, with loss of DELLA relieving growth repression across multiple developmental processes.

Cytokinin and ethylene signaling reveal that eukaryotic plants have retained and elaborated the prokaryotic two-component histidine kinase paradigm. Cytokinin is perceived by the dimerized CRE1 receptor through its extracellular CHASE domain, triggering a multi-step phosphorelay (His → Asp → His → Asp) through the receptor, the HPt shuttle protein, and finally type B response regulators that drive transcription. Ethylene operates as an inhibitor-of-inhibitor system: the ETR1 receptor maintains active CTR1 kinase in the absence of ethylene, which phosphorylates and destabilizes EIN2; ethylene binding inactivates ETR1, stabilizes EIN2, and activates the EIN3/EIL1 transcription factor cascade.

Abscisic acid (ABA) coordinates drought responses through an elegant double-negative regulatory module involving three core components: the PYL receptor family, PP2C phosphatases, and SnRK2 kinases. At basal ABA levels, PP2Cs constitutively dephosphorylate and inactivate SnRK2 kinases. ABA binding triggers closure of the CL2 loop in PYL receptors, creating a surface that inserts directly into the PP2C active site and blocks phosphatase activity. Released SnRK2 kinases autophosphorylate and then phosphorylate downstream effectors including the anion channels SLAC1 and QUAC1 and the K⁺ channel GORK to reduce guard cell turgor and close stomata, as well as transcription factors that drive longer-term ABA-responsive gene expression.

Plant photoreception through phytochromes exemplifies how protein conformational switches driven by photoisomerization can relay information across cellular compartments to control gene transcription networks. Phytochromes are dimeric photoreceptors whose linear tetrapyrrole chromophore (phytochromobilin) undergoes Z-to-E isomerization at the C15–C16 double bond upon red light absorption, converting the inactive Pr form to the active nuclear-localizing Pfr form. The dimeric architecture (Pr–Pr, Pfr–Pr, Pfr–Pfr) and temperature-dependent thermal reversion allow phytochromes to integrate light quality, quantity, and temperature into a graded output. Active Pfr directly phosphorylates PIF transcription factors for proteasomal degradation, inhibits the COP1/SPA E3 ubiquitin ligase to stabilize HY5, and stabilizes Aux/IAA proteins to modulate auxin responses—demonstrating extensive crosstalk between light and hormone signaling that coordinates germination, de-etiolation, shade avoidance, and flowering time throughout the plant life cycle.