The Role of MYC2 Transcription Factors in Plant Secondary Metabolism and Stress Response Mechanisms

Abstract

Jasmonates (JAs) are essential signaling molecules that orchestrate plant responses to abiotic and biotic stresses and regulate growth and developmental processes. MYC2, a core transcription factor in JA signaling, plays a central role in mediating these processes through transcriptional regulation. However, the broader regulatory functions of MYC2, particularly in secondary metabolism and stress signaling pathways, are still not fully understood. This review broadens that perspective by detailing the signaling mechanisms and primary functions of MYC2 transcription factors. It specifically emphasizes their roles in regulating the biosynthesis of secondary metabolites such as alkaloids, terpenes, and flavonoids, and in modulating plant responses to environmental stresses. The review further explores how MYC2 interacts with other transcription factors and hormonal pathways to fine-tune defense mechanisms and secondary metabolite production. Finally, it discusses the potential of MYC2 transcription factors to enhance plant metabolic productivity in agriculture, considering both their applications and limitations in managing secondary metabolite synthesis.

1. Introduction

MYC2 proteins are key transcription factors within the jasmonic acid (JA) signaling pathway and belong to the basic helix-loop-helix (bHLH) superfamily, with wide distribution across land plants [1]. The MYC2 protein contains a conserved bHLH domain at its C-terminus, primarily responsible for DNA binding. It targets critical cis-regulatory elements, including the G-box sequence (CACGTG), E-box motifs (CANNTG), and the GCG-box. Adjacent sequences rich in A/T nucleotides are also crucial, as they significantly enhance the DNA-binding affinity of these transcription factors [2,3]. MYC2 proteins can form homodimers, heterodimers, or even homotetramers through their bHLH and aspartate kinase, chorismate mutase, and tyrA (ACT) domains, enabling both activation and repression of downstream genes [4,5,6].

Furthermore, the N-terminal bHLH-MYC_N domain of MYC2 includes two critical regions: the jasmonate ZIM-domain (JAZ) interaction domain (JID) and the transcription activation domain (TAD). The JID is integral for the binding of MYC2 to JAZ proteins, which are repressors in the JA pathway, while the TAD facilitates the interaction with Mediator complex subunit 25 (MED25). A mutation from Asp128 (D128) to D128N markedly diminishes the interaction between MYC2 and JAZ proteins, although it preserves a robust interaction with MED25 [7]. Additionally, phosphorylation at this site by casein kinase II (CK2) increases the stability of MYC2 and its responsiveness to JA, while alanine substitutions at these phosphorylation sites attenuate the ability to activate JA-dependent signaling [8,9].

JAZ proteins mediate their function through three conserved domains: NT, ZIM, and Jas. The NT domain interacts with DELLA proteins, which are growth repressors in the gibberellin (GA) signaling pathway. The ZIM domain enables dimerization and engages with novel interactor of JAZ (NINJA), while the Jas domain interacts with MYC2 and coronatine insensitive 1 (COI1), facilitating JAZ degradation in response to JA signaling. Some JAZ proteins contain an EAR motif that allows direct interaction with the co-repressor TOPLESS (TPL), while others require NINJA to mediate TPL binding [10].

In the MYC2-JA signaling pathway (Figure 1), MED25 interacts with MYC2, COI1, JAZ, and repressors such as NINJA and TPL [11,12]. Under repressive conditions, JAZ proteins inhibit MYC2. Upon activation by JA-Ile, COI1 mediates JAZ degradation, releasing MYC2 to activate transcription [13]. MED25 enhances MYC2 function by recruiting histone acetyltransferase 1 (HAC1) and RNA polymerase II, promoting histone acetylation (H3K9) and transcription of JA-responsive genes [14]. Additionally, members of the Gro/Tup1 family genes LUG/LUH further stimulate the transcription of MYC2-regulated genes by interacting with MED25 and HAC1 [15]. Moreover, the CUL3-based E3 ubiquitin ligase (CUL3BPM) establishes a negative feedback loop that finely tunes MYC2 expression levels during JA responses, ensuring balanced signaling [16].

In summary, MYC2 transcription factors play a central role in regulating the JA signaling pathway, influencing crucial developmental processes, including plant growth, flowering, and responses to biotic stresses. These factors are also integral in mediating light signaling, hormonal interactions, and broader developmental aspects of plants [17,18,19]. Furthermore, MYC2 proteins play a crucial role in the biosynthesis of secondary metabolites, crucial for plant defense and survival [20,21]. Given their significant roles, this review focuses on the contributions of MYC2 transcription factors to secondary metabolite biosynthesis, with a specific emphasis on elucidating the mechanisms through which these factors enhance plant stress resistance.

2. The Role of MYC2 in the Synthesis of Plant Secondary Metabolites

2.1. MYC2 Transcription Factors in Alkaloid Biosynthesis

Alkaloids are nitrogen-containing secondary metabolites extensively found in plants and known for their significant biological activity. MYC2 transcription factors regulate the expression of key enzymes and genes involved in alkaloid biosynthesis, such as methyltransferases and phosphoribosyltransferases, which play central roles in nicotine production in Nicotiana tabacum (Figure 2).

In tobacco, NtMYC2 functions as a core activator of nicotine biosynthesis. NtMYC2 binds to G-box elements within the proximal promoter regions of nicotine biosynthesis genes such as putrescine N-methyltransferase 2 (PMT2) and quinolinic acid phosphoribosyltransferase 2 (QPT2). It also synergistically upregulates the expression of ERF189, activating PMT2 and QPT2 [22]. Isoforms of NtMYC2, including NtMYC2a, NtMYC2b, and NtMYC2c, enhance nicotine synthesis by activating NtPMT1a, a gene encoding a rate-limiting enzyme in the pathway. The activity of NtMYC2 is also influenced by its interactions with JAZ repressor proteins [23]. CRISPR-Cas9 knockout of NtMYC2a in tobacco has shown a dramatic decrease in the expression of nicotine biosynthetic and transport genes, resulting in about an 80% reduction in nicotine levels in the leaves [24]. These findings confirm the essential role of NtMYC2a in regulating alkaloid biosynthesis and highlight its potential as a key target for metabolic engineering. Additional studies reveal that NtMYC2a-knockout tobacco plants display narrower leaves and increased accumulation of nor-nicotine and methylamine [25], suggesting a broader impact of NtMYC2a on secondary metabolic pathways.

In Catharanthus roseus, CrMYC2 plays a significant role in early JA responses, controlling the expression of terpenoid indole alkaloid (TIA) biosynthetic genes. CrMYC2 specifically targets AP2 domain genes ORCA2, ORCA3, and ORCA4 by binding to G-box-like elements in their promoters, activating these genes and enhancing TIA production [26,27]. In addition to its activating role, CrMYC2 is also involved in a regulatory network that includes repressive factors. G-box binding factors 1 (CrGBF1) and CrGBF2 function as negative regulators of TIA biosynthesis by competing with CrMYC2 for binding to promoter regions of target genes, thereby limiting MYC2-mediated transcriptional activation [28]. Additionally, CrbHLH05, also known as repressor of MYC2 targets 1 (CrRMT1), does not dimerize with CrMYC2 but competes for binding to the T/G-box in the ORCA3 promoter, acting as a passive repressor that antagonizes the activity of CrMYC2 on its targets [29].

In tomato, SlMYC1 and SlMYC2 play significant roles in the production of phenylpropanoid–polyamine conjugates and steroidal glycoalkaloids (SGAs) through the JA signaling pathway. Double knockout of SlMYC1 and SlMYC2 in hairy roots results in dramatically reduced basal expression of SGA biosynthetic genes and significant decreases in α-tomatine and dehydrotomatine content [30]. In addition to their role in the transcriptional regulation of SGA biosynthesis, SlMYC2 also mediates JA–GA crosstalk by modulating the expression of GA catabolic genes, thereby influencing growth–defense trade-offs. JA deficiency suppresses SGA accumulation, whereas low GA levels or impaired GA signaling enhance SGA production [31]. In Lycoris aurea, LaMYC2 binds to E-box elements in the promoter of tyrosine decarboxylase (LaTYDC), a gene involved in lycorine biosynthesis, acting as a positive regulator of this Amaryllidaceae alkaloid [32].

2.2. MYC2 Transcription Factors in Terpenoid Biosynthesis

Terpenoids, a diverse class of compounds including monoterpenes, sesquiterpenes, diterpenes, triterpenes, and polyterpenes, play important roles as volatile and non-volatile secondary metabolites in plants. MYC2 transcription factors serve as pivotal regulators of terpenoid biosynthesis by modulating the expression of key biosynthetic genes.

Monoterpenes, such as pyrethrins derived from the flowers of Tanacetum cinerariifolium, are effective broad-spectrum insecticides that are safe for mammals and widely used in organic agriculture and household pest control [33]. The TcMYC2 gene in T. cinerariifolium responds to JA treatment, directly regulating pyrethrin synthesis by binding to the promoters of biosynthetic genes such as chysanthemyl diphosphate synthase (TcCDS), GDSL lipase-like acyltransferase (TcGLIP), and allene oxide cyclase (TcAOC) at E/G-box elements [34]. Another MYC2-like transcription factor, TcbHLH14, also targets TcCDS and TcAOC but not TcGLIP, resulting in only a moderate increase in pyrethrin levels [35].

Linalool is a major component of volatiles that plays a crucial role in plant fragrance and pollination. In Osmanthus fragrans, OfMYC2 promotes linalool biosynthesis by transcriptionally activating OfTPS2 in cooperation with OfMYB21, although this activation is suppressed by OfJAZ3 through its interaction with OfMYC2 [36]. In Chimonanthus praecox, the expression of CpMYC2 correlates with flowering stages, showing a significant increase from bud to full bloom. Overexpression of CpMYC2 in Arabidopsis and tobacco significantly enhances the production of volatile monoterpenes, especially linalool. CpMYC2 interacts with the JA signaling pathway to upregulate TPS gene expression in response to JA and GA treatments, increasing the release of aromatic compounds [37]. In Freesia hybrida, linalool synthesis is predominantly mediated by the monoterpene synthase TPS1. FhMYC2 acts as a negative regulator by restricting FhMYB21 binding to the FhTPS1 promoter, inhibiting linalool synthesis. In contrast, co-expression of AtMYC2 with AtMYB21 in Arabidopsis thaliana preferentially activates sesquiterpene genes while suppressing the monoterpene gene AtTPS14, favoring sesquiterpene over monoterpene emission, including linalool [38]. Additionally, FhMYB108 in F. hybrida enhances linalool synthesis by promoting FhTPS1 expression. Interestingly, interactions between FhMYC2 and FhMYB108 proteins in Arabidopsis inhibit AtTPS14 activation, revealing the complex molecular mechanisms by which MYC2 collaborates with MYB transcription factors to regulate volatile terpenoid biosynthesis [39].

Sesquiterpenes play significant roles in plant defense and pollinator attraction [40,41]. In sweet orange, CitMYC3, a MYC2 homolog, functions as a key regulator of valencene synthesis, activating the promoter and enhancing the expression of the sesquiterpene synthase gene CsTPS1 [42]. In A. thaliana, AtMYC2 controls sesquiterpene production by binding to the promoters of TPS21 and TPS11, thus stimulating the expression of these genes and facilitating the release of compounds such as (E)-β-caryophyllene, which is influenced by both GA and JA [43].

Artemisinin, a sesquiterpene lactone from Artemisia annua, is synthesized in glandular cells [44]. The JA-responsive transcription factor AaMYC2 enhances artemisinin production by binding to the promoters of key artemisinin biosynthesis genes CYP71AV1 and artemisinic aldehyde double bond reductase (DBR2). Transgenic plants overexpressing AaMYC2 show elevated levels of artemisinin [45]. Additionally, AaMYC2, together with AabHLH1, responds to both MeJA and abscisic acid (ABA), activating crucial genes involved in artemisinin synthesis and thus boosting its yield [46]. The MYC2 homolog AaMYC3 also plays a significant role by regulating glandular density and artemisinin biosynthesis. Overexpression of AaMYC3 results in increased glandular density and artemisinin content, while RNA interference targeting AaMYC3 decreases both. AaMYC3 activates the transcription of AaHD1, which is involved in gland development, and upregulates artemisinin biosynthetic genes, including CYP71AV1 and aldehyde dehydrogenase 1 (ALDH1). It further collaborates as a co-activator with AabHLH1 and AabHLH113 to enhance the transcription of key artemisinin biosynthesis genes, amorpha-4,11-diene synthase (ADS) and DBR2, thereby amplifying artemisinin production [47]. Contrarily, AabHLH2 and AabHLH3 act as transcriptional repressors by competing for the same cis-regulatory elements as AaMYC2, albeit lacking a conserved activation domain, which suggests they inhibit artemisinin synthesis [48].

Sesquiterpenes derived from agarwood are recognized for their antibacterial and antifungal properties and typically accumulate in response to elicitation or wounding signals. In Aquilaria sinensis, AsMYC2 is repressed by AsJAZ1 repressor protein, thereby preventing it from activating the expression of agarwood sesquiterpene synthase gene 1 (ASS1). Under normal conditions, ASS1 exhibits low expression levels, resulting in limited sesquiterpene production. However, in wounded A. sinensis, endogenous JA biosynthesis triggers the release of AsMYC2, which directly targets and activates ASS1, enhancing sesquiterpene biosynthesis [49]. In Gossypium hirsutum, the sesquiterpene aldehyde gossypol acts as a crucial antimicrobial metabolite for defense against pathogens and insect predation. GhMYC2 influences the gossypol biosynthetic pathway by regulating the activity of enzymes in the cytochrome P450 (CYP450) superfamily, specifically CYP71BE79, modulating gossypol production [50].

In Lavandula angustifolia, overexpression of MYC2 homolog LaMYC4 is associated with increased levels of sesquiterpenes, including caryophyllenes, in Arabidopsis and tobacco. This overexpression correlates with enhanced gene activity, which is crucial for terpenoid biosynthesis and increases the number and size of glandular trichomes [51]. In Curcuma wenyujin, CwMYC2, a key regulator within the JA signaling pathway, significantly upregulates genes associated with β-elemene biosynthesis, thereby enhancing β-elemene accumulation in the leaves [52]. In Oryza sativa, OsMYC2 promotes the biosynthesis of resistance compounds, including the monoterpene geraniol and the sesquiterpene caryophyllene [53].

Diterpenes are pivotal in plant defense mechanisms and valuable in human medicine. Paclitaxel, a crucial anticancer drug used to treat ovarian and breast cancers, is synthesized under strong induction by JA [54]. In Taxus species, transcription factors such as MYC2, MYC3, and MYC4 regulate essential genes in the paclitaxel biosynthetic pathway, including taxane synthase (TASY), 10-deacetylbaccatin III-10β-O-acetyltransferase (DBTNBT), and taxadiene 5-alpha hydroxylase (T5H). These transcription factors bind to the promoters of these genes, activating their transcription and thus promoting paclitaxel production [55]. In Taxus chinensis, the transcription factor TcMYC2a directly binds to the promoter of the TASY gene, enhancing TASY expression and potentially influencing paclitaxel biosynthesis by upregulating TcERF15 [56].

Tanshinones, representative diterpenoid compounds in Salvia miltiorrhiza, are synthesized through a pathway that involves key enzymes such as Hydroxycinnamate-CoA ligase 6 (SmHCT6) and P450 monooxygenase (SmCYP98A14), which are induced by JA. SmMYC2a activates the expression of both SmHCT6 and SmCYP98A14 by binding to the E-box elements in their promoters, while SmMYC2b specifically regulates SmCYP98A14 [57]. SmMYC2 collaborates with SmbHLHL37 to enhance the transcription of genes such as geranylgeranyl pyrophosphate synthase (SmGGPPS), thereby promoting tanshinone synthesis. It also dynamically interacts with SmMYB36 to regulate JA-mediated tanshinone accumulation [58,59]. Conversely, SmbHLH60 acts as a negative regulator, competing with SmMYC2 for control and repressing transcription of targets such as tyrosine aminotransferase 1 (SmTAT1) and dihydroflavonol 4-reductase (SmDFR), thus inhibiting phenolic acid and anthocyanin biosynthesis pathways in S. miltiorrhiza [60]. In Salvia sclarea, overexpression of A. thaliana AtMYC2 and AtWRKY40 in hairy roots activates the methylerythritol 4-phosphate pathway, enhancing diterpene content [61]. Heterologous expression of AtMYC2 in S. miltiorrhiza hairy roots significantly upregulates genes including 1-deoxy-D-xylulose 5-phosphate synthase (SmDXS2) and SmTAT, leading to enhanced accumulation of tanshinones and phenolic acids, with tanshinone yield reaching 14.06 mg/g dry weight—5.45 times that of the control—and phenolic acid yield at 95.9 mg/g dry weight, a 3.3-fold increase [62].

In Tripterygium wilfordii, TwMYC2a and TwMYC2b negatively regulate triptolide biosynthesis by inhibiting the expression of TwTPS27a and TwTPS27b in hairy roots [63]. In Ginkgo biloba, GbMYC2 binds directly to the G-box in the promoter of the levopimaradiene synthase (GbLPS) gene, activating genes associated with ginkgolide biosynthesis [64]. Furthermore, GbMYC2 enhances ginkgolide biosynthesis by activating the expression of the GbGGPPS gene through promoter binding. GbMYC2_4 selectively binds to the canonical G-box motif within the GbGGPPS promoter, while GbMYC2_5 preferentially interacts with an adjacent A/T-rich G-box-like motif, thereby synergistically activating GbGGPPS expression [65].

Carotenoids, tetraterpenoid compounds responsible for the vibrant coloration in citrus fruits, are influenced by the red carotenoid β-citraurin, a key pigment in the peel of the Newhall orange. MeJA treatment significantly enhances β-citraurin production and coloration. This induction upregulates CsMYC2, which then activates the gene CsCCD4b in the β-citraurin biosynthetic pathway by binding to its promoter, thereby impacting fruit coloration. Additionally, CsMYC2 promotes the expression of CsMPK6, which interacts with CsMYC2, reducing its stability and DNA-binding activity, thus establishing a negative feedback loop that modulates JA signaling during fruit coloration [66]. In Chrysanthemum indicum var. aromaticum, MeJA induces the expression of CiMYC2, whose overexpression in tobacco leads to shorter plants, deeper leaf color, increased chlorophyll and carotenoid content, and the production of new terpenoid compounds [67].

In the rubber tree (Hevea brasiliensis), the small rubber particle protein (SRPP) plays a key role in natural rubber biosynthesis. HbMYC2b, highly expressed in the bark, binds to the HbSRPP promoter to activate its transcription, thus positively regulating HbSRPP expression [68]. In Taraxacum kok-saghyz, overexpression of TkMYC2 not only inhibits leaf development and promotes root growth but also enhances natural rubber production. TkSRPP and rubber elongation factor (TkREF) genes are upregulated in transgenic lines, suggesting that TkMYC2 regulates natural rubber synthesis by modulating TkSRPP/REF expression [69].

In Gynostemma pentaphyllum, GpMYC2 plays a critical role in activating the synthesis of gypenosides by binding to the promoters of genes involved in their biosynthesis, thereby confirming the significant role of the COI1/JAZ/MYC2 module in regulating responses induced by MeJA [70]. Taraxacum antungense, a traditional herb known for its antibacterial and antioxidant properties and rich triterpenoid content, also demonstrates the regulatory capabilities of MeJA. The overexpression of TaMYC2, induced by MeJA, significantly enhances triterpenoid accumulation, with the expression level of the squalene synthase gene (TaSS) elevated three to five times compared to control lines [71]. In Bupleurum chinense, BcMYC2 promotes the expression of key enzyme genes such as 3-Hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR), isopentenyl diphosphate isomerase (IPPI), farnesyl pyrophosphate synthase (FPS), and p-acetoxydihydrochalcone synthase (p-AS), which are essential for the saikosaponin biosynthetic pathway [72].

2.3. MYC2 Transcription Factors in Flavonoid Synthesis

Flavonoids, recognized for their potent antioxidant properties, are key plant secondary metabolites abundant in fruits, vegetables, and flowers. In A. thaliana, the MYC2 homolog MYC3 plays a leading role in enhancing plant resistance to insect herbivory. Mutants of MYC3 show significantly reduced levels of flavonoid compounds, potentially alleviating wound-induced growth suppression [73]. In apples, MdMYC2, responsive to JA, significantly upregulates genes associated with anthocyanin synthesis and effectively increases anthocyanin content [74]. The regulation of anthocyanins by MYC2 is complex, exhibiting both activation and repression. MYC2 can activate structural genes in the anthocyanin biosynthetic pathway. However, in the presence of the MYB/bHLH/WD40 complex, MYC2 binds to GL3, a bHLH component of the complex, thereby inhibiting its formation and repressing anthocyanin synthesis. This dual regulatory role is further influenced by interactions with negative regulators such as BBX21 and SPL9 [75]. In rice, OsMYC2 enhances the expression of naringenin 7-O-methyltransferase (OsNOMT), a key gene in sakuranetin production, by activating its promoter in response to JA treatment. This process is further amplified through interactions with OsMYC2 homologs, OsMYL1 and OsMYL2, collectively promoting JA-induced sakuranetin synthesis [76].

In Ipomoea batatas, MYC2 plays a crucial role in enhancing anthocyanin accumulation by binding to promoters of key biosynthetic genes such as IbCHI and IbDFR. Overexpression of IbMYC2 under saline and drought conditions enhances the expression of genes related to reactive oxygen species (ROS) scavenging and proline synthesis, increasing stress tolerance [77]. In Vitis vinifera, the biosynthesis of flavonols, anthocyanins, and proanthocyanidins (PAs) is distinctly regulated both temporally and spatially during berry development, co-regulated by MYC and MYB transcription factors. VvMYC2 interacts with MYB24 to co-regulate the synthesis of terpenes and flavonols under UV and high-intensity visible light stress in grape skin regions devoid of anthocyanins, enhancing fruit tolerance to light stress [78].

In Medicago sativa, the expression of MsMYC2 is significantly upregulated in the mtugt84a1 mutant, where the UDP-glycosyltransferase suppresses the JA signaling pathway through glycosylation and feedback regulation, impacting anthocyanin accumulation [79]. In Zea mays, overexpression of ZmMYC2 in Arabidopsis mutants restores JA sensitivity, leading to inhibited root growth and increased anthocyanin accumulation [80]. In Camellia sinensis, CsMYC2 positively regulates flavan-3-ol biosynthesis under JA signaling. However, splice variants CsJAZ1-1, CsJAZ1-2, and CsJAZ1-3 form complexes with CsMYC2, suppressing the expression of key flavan-3-ol biosynthetic genes [81].

2.4. Regulatory Role of MYC2 in VOCs, Essential Oils, and Other Compounds

In Litsea cubeba, an essential oil-rich plant valued for its high neral and geranial content (up to 80% of oil composition), LcMYC2 enhances essential oil biosynthesis by binding to the promoters of LcTPS42 and LcGPPS [82]. In A. thaliana, (E)-2-hexenal, a key compound involved in plant communication and pest resistance, induces the expression of WRKY46 and MYC2. Together, these factors activate respiratory burst oxidase homolog d (RBOHD) and flavonoid biosynthesis genes, thereby increasing total flavonoid accumulation and enhancing pest resistance [83].

In Chinese cabbage (Brassica rapa ssp. pekinensis), BrMYC2 overexpression significantly enhances the accumulation of glucosinolate (GS), compounds that confer strong resistance to bacterial soft rot [84]. In callus tissues of apple, MdMYC2 directly binds to the promoter of MdLOX5a, significantly enhancing the biosynthesis of volatile aldehydes and alcohols essential for flavor and aroma [85].

3. Response of MYC2 to Stress

MYC2 plays a significant role in regulating the synthesis of plant secondary metabolites and acts as a key regulator in plant responses to abiotic stresses. It activates downstream stress-responsive genes, enhancing plant adaptation to cold, drought, salinity, and other adverse conditions. An overview of MYC2-mediated stress response pathways is illustrated in Figure 3.

3.1. The Role of MYC2 in Cold Stress

Plants enhance cold tolerance primarily through the activation of the C-repeat-binding factor (CBF) pathway. Under non-stress conditions, JAZ1/4 interacts with inducer of CBF expression 1 (ICE1) and ICE2 to suppress this pathway. Upon exposure to cold, JA biosynthesis is induced, leading to the degradation of JAZ1/4 proteins. This relieves repression of the ICE-CBF cascade and allows activation of downstream cold-responsive genes. MdMYC2 enhances frost resistance by directly regulating the expression of MdCBF1, thereby improving cold tolerance in apples [86]. In Manihot esculenta, MeMYC2.2 is significantly upregulated under cold stress. It activates MeCBF3 expression, and its overexpression in transgenic Arabidopsis plants leads to enhanced cold tolerance [87]. In peach fruits, exogenous MeJA treatment upregulates PpMYC2.2, activating the JA signaling pathway, reducing electrolyte leakage, and protecting cell membranes by regulating lipid metabolism. PpMYC2.2 also synergizes with the CBF pathway, particularly PpCBF3, to increase peach cold tolerance [88]. Similarly, in the winter wheat variety Dn1, which is capable of successfully overwintering at extremely low temperatures, expression of TaMYC2 is induced by cold stress and JA treatment. This activation triggers the ICE-CBF-COR cold resistance pathway, significantly enhancing cold tolerance. At freezing temperatures, cell lines overexpressing TaMYC2 exhibit reduced electrolyte leakage, lower malondialdehyde content, increased proline levels, and enhanced antioxidant defenses [89].

Beyond these examples, additional studies in other species have further highlighted the central role of MYC2 in cold stress responses. In cold-exposed Poncirus trifoliata, the betaine aldehyde dehydrogenase gene (PtrBADH-1) is activated by PtrMYC2, regulating the accumulation of cold-induced glycine betaine, which helps the plant to cope with low-temperature stress [90]. In Trifolium ambiguum, TaMYC2 is responsive to multiple stresses, including salt, alkali, cold, and drought, and is induced by plant hormones such as IAA, GA3, and MeJA. Cold and drought stresses specifically induce TaMYC2 expression, further enhancing the activity of antioxidant enzymes [91].

In tomato, SlMYC2 enhances cold tolerance by modulating polyamine biosynthesis and oxidative stress management. This gene upregulates arginine decarboxylase 1 (ADC1), leading to increased putrescine production, which decreases ROS levels and alleviates oxidative stress [92]. Treatment with MeJA boosts the activities of the ascorbate–glutathione (AsA-GSH) cycle enzymes and activates the SlICE-SlCBF-SlCOR signaling pathway, which is critical for cold damage mitigation. Silencing SlMYC2 curtails these protective effects, underscoring its crucial role in these pathways [93]. SlMYC2 also triggers the expression of glutathione S-transferase U24 (SlGSTU24) and β-Amylase 3 (SlBAM3), facilitating ROS reduction and starch degradation, respectively, thus improving cold tolerance [94,95]. Moreover, MYC2 enhances the expression of nine-cis-epoxycarotenoid dioxygenase 2 (NCED2), increasing ABA accumulation and strengthening plant tolerance to low-temperature stress [96]. Simultaneously, MYC2 activates SlERF.B8. This gene not only responds to cold stress and JA signals but also forms a positive feedback loop with MYC2, amplifying JA signaling and further enhancing the plant’s cold tolerance [97]. Additionally, SlMYC2 binds to G/E-box elements to activate genes like arginase (SlARG1 and SlARG2), arginine decarboxylase (SlADC), and ornithine decarboxylase (SlODC), boosting polyamine levels and further mitigating cold stress [98]. Furthermore, SlMYC2 is integral to the crosstalk between JA and melatonin (MT) pathways. Cold stress induces JA accumulation, which upregulates MYC2-activated genes involved in MT biosynthesis, including Serotonin N-acetyltransferase (SlSNAT) and acetyl-5-hydroxytryptamine O-methyltransferase (SlAMT). Increased MT accumulation not only potentiates cold tolerance but also promotes further JA biosynthesis, creating a positive feedback loop that amplifies cold responses [99].

3.2. The Role of MYC2 in Drought Tolerance

MYC2 transcription factors play a significant role in plant drought tolerance by modulating diverse drought-responsive pathways. In A. thaliana, AtMYC2 enhances drought resistance by directly binding to the promoter of the early responsive to dehydration 1 (ERD1) gene, a critical player in drought adaptation [100].

In tomato, SlMYC2 improves drought tolerance through multiple regulatory routes. It represses protein phosphatase 2C1 (SlPP2C1) and the cytokinin signaling component SlRR26, negatively affecting ROS accumulation and stomatal closure. Overexpression of SlRR26 reduces drought tolerance, whereas slrr26 mutants show enhanced resistance [101]. Moreover, SlMYC2 represses chalcone synthase 1 (SlCHS1), decreasing flavonol levels and increasing ROS in guard cells. This promotes stomatal closure by elevating JA and ABA levels [102].

Similar regulatory functions of MYC2 have been observed in other crops. In Brassica napus, silencing BnMYC2 disrupts stomatal closure under light and dark conditions, resulting in excessive water loss and decreased drought tolerance [103]. In sorghum, SbMYC2 is strongly induced by polyethylene glycol (PEG)-simulated drought and JA treatment. Its overexpression enhances drought resistance in Arabidopsis, rice, and sorghum by reducing ROS levels and maintaining higher chlorophyll content [104].

In wheat, MYC2 co-regulates melatonin biosynthesis by directly interacting with the promoter of the N-acetylserotonin methyltransferase (ASMT) gene. This MYC2-ASMT module enhances drought tolerance by modulating melatonin levels in wheat leaves [105]. Additionally, the miR1119-MYC2 regulatory module has been identified in wheat roots, where it influences ABA hormone levels, water relations, and photosynthetic activity, enhancing drought tolerance through hormonal crosstalk [106]. In poplar, MYC2 regulates stomatal development by targeting the promoters of stomatal density-related genes, including epidermal patterning factor 2 (PpnEPF2), PpnEPFL4, and PpnEPFL9. Overexpression of PpnMYC2 in both poplar and Arabidopsis results in reduced stomatal density, improved water use efficiency, and enhanced drought resistance [107]. In barley (Hordeum vulgare), MYC2 directly binds to the JA response element in the promoter of Ribulose-1,5-bisphosphate carboxylase/oxygenase activase A (RcaA), enhancing photosynthetic efficiency under combined drought and salt stress. This regulatory module improves plant water status [108].

3.3. The Role of MYC2 in Water Stress

Environmental water stress, including rainfall, submersion, and osmotic stress, triggers short-term molecular responses and long-term developmental adjustments in plants. MYC2 transcription factors play a central role in regulating these responses by integrating hormonal and oxidative stress signaling. Simulated rainfall, through water spray stress, activates the JA signaling pathway. In this process, MYC2 interacts with bHLH19 and ERF109 to activate octadecanoid-responsive AP2/ERF-domain 47 (ORA47), promoting JA biosynthesis and forming a positive feedback loop that enhances JA accumulation [109]. In C. sinensis, MYC2 activates the transcription of JA biosynthesis-related genes and peroxidase (PER) genes by binding to their promoters, forming a positive feedback loop that enhances tea plant tolerance to osmotic stress [110]. Similarly, in sunflowers, MYC2 integrates JA and ABA signaling pathways, activating stress-responsive transcription factors such as dehydration 20 (RD20), RD22, RD26, ANAC19, and ANAC29. Concurrently, the JA and SA pathways jointly activate the WRKY70 transcription factor, enhancing plant tolerance to water stress [111]. In A. thaliana, MYC2 interacts with MYB30 to regulate the expression of antioxidant genes, including Vitamin C defective 1 (VTC1) and Glutathione synthetase 1 (GSH1). This interaction integrates light signals with reoxygenation stress responses, enhancing the plant’s antioxidant capacity and improving survival rates. Overexpression of VTC1 and GSH1 can completely rescue the hypersensitivity to submersion observed in myc2 mutants [112].

Although these studies differ in experimental systems, stress types, and target genes, they consistently support the notion that MYC2 acts as a central regulatory hub in plant water stress responses. Apparent differences in downstream mechanisms likely reflect species-specific signaling networks and stress-specific regulatory crosstalk.

3.4. Role of MYC2 in Salt Stress

The role of MYC2 transcription factors in response to salt stress is complex, exhibiting both positive and negative regulatory effects across different plant species. Within the JA signaling pathway, MYC2 genes generally increase plant sensitivity to salt. For instance, in Arabidopsis, JA suppresses the expression of the antioxidant enzyme Catalase 2 (CAT2) through MYC2, reducing seedling salt tolerance [113]. Additionally, MYC2 binds to the 5-UTR of the key rate-limiting enzyme delta1-pyrroline-5-carboxylate synthase (P5CS1) in proline biosynthesis, negatively regulating proline synthesis and further diminishing the plant salt tolerance [114]. In Curcuma wenyujin, the CwJAZ4/9 complex inhibits the JA-induced terpene synthesis pathway by interacting with CwMYC2, leading to reduced terpenoid accumulation but enhancing the salt tolerance of hairy roots, maintaining their growth under salt stress [115].

Conversely, MYC2 genes positively regulate salt stress tolerance in some plants, particularly through ABA-related pathways. In S. miltiorrhiza, overexpression of SmMYC2 boosts salt tolerance by increasing the activities of antioxidant enzymes (SOD, POD, and CAT) and proline content. This upregulation also involves the activation of flavonoid biosynthesis genes, further improving the plant’s antioxidant capacity and salt stress tolerance [116]. Similarly, in wheat, the small ubiquitin-like modifier (SUMO) protease gene (TaDSU) interacts with MYC2, reducing its sumoylation levels and enhancing its transcriptional activity. This interaction establishes a positive feedback loop, where MYC2 binds to the TaDSU promoter, boosting its expression and improving ion balance (higher K⁺/Na⁺ ratio) and salt tolerance [117]. In Caragana korshinskii, CkMYC2 regulates the expression of the pyrabactin resistance like 4 (CkPYL4) gene, promoting ABA accumulation in roots and thereby enhancing plant tolerance to both salt and drought stress [118]. Additionally, in rice, a MYC2-like transcription factor binds to the ABA-responsive element in the promoter of cytochrome P450 family 2 (OsCYP2), enhancing salt tolerance, partially restoring the salt tolerance of cyp2-RNAi rice, and increasing antioxidant enzyme activity [119].

3.5. Role of MYC2 in Heavy Metal Stress

Reports on the role of MYC2 transcription factors in heavy metal stress are limited. In A. thaliana, MYC2 suppresses the expression of heavy metal ATPase gene 2 (HMA2) and HMA4, altering cadmium (Cd) distribution and reducing tolerance. However, MYC2 degradation under Cd stress partially alleviates its repressive effects, aiding stress adaptation [120]. In wheat, TaMYC8 negatively regulates Cd-responsive ethylene signaling. TabHLH094, a Cd-induced bHLH transcription factor, inhibits TaMYC8 activity, reducing its binding to the TaERF6 promoter and limiting ethylene biosynthesis. Overexpression of TabHLH094 enhances wheat Cd tolerance by modulating TaMYC8 activity and suppressing ethylene production [121].

3.6. Role of MYC2 in Biotic Stress

MYC2 transcription factors play significant roles in enhancing plant biotic stress. In tomatoes, the Solyc08g005050, which belongs to the MYC2 subfamily, interacts synergistically with the HD-ZIP IV transcription factor (Wo) to promote trichome development and terpene biosynthesis, thereby enhancing resistance to spider mites [122]. Similarly, in chrysanthemums, CmMYC2 regulates the development of T-shaped and glandular trichomes and the accumulation of terpenoid compounds. By directly activating CmMYBML1 and forming a feedback inhibition loop with it, while CmMYC2 promotes the initial activation of CmMYBML1, the overexpression of CmMYBML1 subsequently binds and consumes CmMYC2, preventing sustained activation of CmMYBML1. These mechanisms significantly enhance the plant’s resistance to herbivorous larvae [123]. The MYC2-like transcription factor pigment gland formation (GoPGF) activates the expression of jasmonate-associated VQ motif-like protein (JAVL) in cotton, which establishes a negative feedback loop by inhibiting the transcription of GoPGF, thereby balancing GoPGF and JAVL expression. Furthermore, JAVL regulates JA levels by inhibiting the expression of JA synthesis-related genes through its interaction with GoPGF. An increase in the GoPGF to JAVL expression ratio leads to enlarged pigment glands and increased accumulation of JA and defense compounds, thereby enhancing cotton’s resistance to insects and pathogens [124]. In cotton, the MYC2-like transcription factor GhMYC1374 is significantly induced under aphid attack. Studies indicate that GhMYC1374 enhances cotton’s resistance to aphids by activating the biosynthesis of flavonoid compounds and free gossypol. The overexpression of GhMYC1374 significantly increases cotton’s resistance to aphids, while silencing GhMYC1374 via VIGS technology reduces its resistance [125].

In Arabidopsis, MYC2 induces the expression of the cell wall acetylation gene trichome birefringence-like 37 (AtTBL37), enhancing cell wall acetylation, thereby strengthening resistance to herbivores [126]. In B. napus, MeJA-induced BnMYC2 positively regulates the expression of the anti-insect gene vegetative storage protein 2 (VSP2), enhancing the plant defense against insect stress [127]. In tomatoes, SlMYC2 regulates the MeJA-induced gene (SlJIG) by directly binding to its promoter, leading to the activation of TPS genes and enhanced resistance to insect and microbial stress [128].

In rice, MYC2 triggers transcriptional cascades by regulating secondary transcription factors like bHLH6, amplifying JA responses across tissues. Additionally, MYC2 establishes a feedback mechanism by modulating the expression of JA repressors and catabolic genes, including NAC transcription factors (NAC1, NAC3, and NAC4), which attenuate JA responses and reduce defense capabilities against insect herbivores [129]. In tomato, MYC2 activates the E3 ubiquitin ligase (PUB22), facilitating JAZ protein degradation via the 26S proteasome pathway. This mechanism strengthens defense against Helicoverpa armigera. The MYC2-PUB22-JAZ4 module also regulates JA-mediated responses, such as resistance to Botrytis cinerea, inhibition of root elongation, and anthocyanin accumulation [130].

In C. sinensis, CsMYC2.2 directly binds to the promoter of CsGSTU45, activating its expression and reducing resistance to Colletotrichum camelliae. Silencing CsMYC2.2 significantly enhances resistance to tea cake disease [131]. In maize, ZmMYC7 activates ZmERF147 and defense genes like pathogenesis-related protein 1 (ZmPR1), ZmPR2, and ZmPR3, bolstering resistance to Fusarium graminearum and Setosphaeria turcica [132]. Furthermore, ZmMYC2a and ZmMYC2b enhance the synthesis of defense metabolites such as benzoxazinoids and volatile terpenes in response to JA signaling or simulated herbivory. Double mutant studies reveal increased insect susceptibility and reduced production of these metabolites [133].

3.7. MYC2 Mediators of Hormonal Interactions and Stress Adaptation in Plants

MYC2 transcription factors are central players in JA signaling, integrating multiple hormonal pathways. In the JA-ABA pathway, MYC2 interacts with ABA-insensitive 5 (ABI5) and directly activates the ABA2 promoter through the MKK3-MPK6-MYC2 module, enhancing ABA biosynthesis and modulating ABA-mediated responses such as drought responses and seed germination inhibition [134,135].

In JA-SA crosstalk, MYC2 plays dual regulatory roles. It interacts with SA-induced NPR1, which inhibits JA-responsive genes by forming complexes with MYC2 and blocking its interaction with MED25 [136]. Simultaneously, MYC2 promotes SA biosynthesis and signaling, enhancing defenses like PAMP-triggered immunity (PTI) and effector-triggered immunity (ETI) [137]. This JA-SA division of labor is evident in plant defense against chewing and piercing-sucking insects, where JA enhances resistance to caterpillars while inhibiting whiteflies [138].

In JA-GA crosstalk, MYC2 regulates SGA biosynthesis and GA catabolism by activating GA metabolic genes such as GA2ox3 and GA2ox7, which inactivate GA and redistribute resources under stress conditions, including herbivore attacks like those from Nilaparvata lugens. Additionally, the DELLA interacts with MYC2, disrupting JA regulation of SGA metabolism and affecting MYC2 activity [31,139].

This multi-hormonal cross-regulatory network not only optimizes plant adaptability to environmental conditions but also enhances survival strategies under complex adversities [140,141].

4. Applications and Prospects of MYC2 Transcription Factors

4.1. MYC2 as a Central Regulatory Hub Gene Across and Within Species

MYC2 transcription factors are known to regulate the expression of more than 1300 genes and play a pivotal role as core regulators in the JA signaling pathway [109]. Their broad involvement in various biological processes, including the regulation of secondary metabolism, hormonal crosstalk, and plant responses to abiotic and biotic stresses, has led to the widely accepted view that MYC2 functions as a central transcriptional hub. However, this inference is often drawn from scattered observations across multiple plant species. To determine whether MYC2 indeed acts as a central regulator, assessing its roles within individual species is important.

Evidence from several models and crop plants supports the multifaceted regulatory role of MYC2 within individual species. Beyond its well-established functions in plant growth and development [19], AtMYC2 in A. thaliana regulates flavonoid biosynthesis, drought and salt tolerance, ABA signaling, and SA-JA antagonism by interacting with a range of downstream targets, including ERD1, ABA2, HMA2, GL3, and NPR1 [73,100,114,134]. Similarly, in tomato, SlMYC2 controls steroidal glycoalkaloid biosynthesis, cold and drought resistance, melatonin and polyamine metabolism, and volatile production activating genes such as SlGSTU24, SlBAM3, SlADC1, SlERF.B8, and SlLOX5a [30,92,93,99]. These cases illustrate that MYC2 acts as an integrative regulatory factor capable of synchronizing multiple environmental signals and developmental programs within a single plant species.

Comparative studies across plant species show that MYC2 transcription factors are highly conserved in their domain and JA-responsiveness, particularly in their ability to bind G-box/E-box cis-elements. However, their downstream regulatory targets are often species-specific. For instance, MYC2 regulates nicotine biosynthesis in N. tabacum [23], artemisinin in A. annua [45], taxol in Taxus [56], ginkgolides in G. biloba [65], and pyrethrins in T. cinerariifolium [34]. These compounds are biosynthesized in a species-dependent manner, indicating that MYC2 has been independently recruited to regulate diverse metabolic pathways across species.

Together, these findings support a model in which MYC2 transcription factors serve as evolutionarily conserved regulatory switches that control distinct specialized metabolic pathways in different plant species. This regulatory versatility makes MYC2 an attractive target for metabolic engineering. However, such broad regulatory roles also raise important concerns about pleiotropic effects when manipulating MYC2 function.

4.2. Balancing Growth and Stress Resistance: The Dilemma of MYC Transcription Factors in Plants

Although MYC2 plays a central role in coordinating JA-mediated defense and specialized metabolism, its activation is frequently associated with growth inhibition. This growth–defense trade-off poses a significant challenge for its direct application in metabolic engineering and the development of stress-resilient cultivars.

One of the underlying causes of this dilemma is the dual regulatory nature of MYC2. It functions as a molecular switch that activates secondary metabolite biosynthetic genes and induces transcriptional repressors involved in metabolic downregulation, thereby maintaining cellular homeostasis [142]. Upon accumulation of JA-Ile, JAZ proteins are ubiquitinated and degraded, releasing MYC2 to activate defense-related genes. Concurrently, JA signaling stabilizes BPM proteins, which serve as adaptors for Cullin3-based E3 ubiquitin ligases and promote the proteasomal degradation of MYC2 itself. This negative feedback mechanism ensures a tightly controlled transcriptional output and prevents overactivation of the JA response [16,143].

Beyond molecular feedback, MYC2 exhibits pronounced pleiotropy, impacting both defense responses and plant development. In rice, MYC2 upregulates GA catabolism genes, such as GA2ox3 and GA2ox7. This suppresses GA levels, prioritizing resources for defense at the expense of growth [139]. Similarly, JA treatments inhibit GA biosynthesis and activate GA catabolism via miR5998 and MYC2, further reducing endogenous GA levels to curtail growth [144]. In A. thaliana, jaz-deficient mutants that exhibit constitutively active MYC2 signaling display stunted growth, including reduced shoot and root size, smaller fruits, and lower seed yield. MYC2, together with MYC3 and MYC4, negatively regulates seed size, weight, and storage compound accumulation, while triple mutants produce larger, heavier seeds [145,146]. Furthermore, JA-related components like COI1, MED25, and MYC2 restrict seed size, with JA treatment reducing seed growth in a COI1-dependent manner [147]. MYC3/4 also restricts seed growth by impeding seed coat cell proliferation and elongation, collaborating with the KIX-PPD complex to regulate seed size [148].

These findings emphasize that direct overexpression or knockout of MYC2 may result in undesirable developmental trade-offs, such as reduced biomass or fertility. Therefore, biotechnological applications must move beyond simplistic manipulations of MYC2 expression and adopt more refined strategies that preserve stress resistance while minimizing growth penalties.

One promising direction is the use of tissue-specific or inducible promoters to restrict MYC2 expression to particular organs or stress conditions, thereby limiting its activity to scenarios where defense activation is most beneficial. In parallel, synthetic promoter engineering can be used to design MYC2-responsive regulatory elements that drive the expression of only selected downstream genes, enabling targeted activation of secondary metabolic pathways without interfering with core developmental processes. Moreover, recent advances in CRISPR/Cas9-based transcriptional control systems allow for precise activation or repression of individual MYC2 target genes, offering another layer of regulatory specificity. In addition, protein engineering approaches could be employed to modulate MYC2 interactions with co-regulators such as MED25, or to modify its degradation motifs, thereby altering its stability and transcriptional specificity. Finally, manipulation of MYC2-interacting repressors, such as specific JAZ or TPL family members, may allow for fine-tuned redirection of MYC2 activity toward beneficial regulatory branches, while minimizing unwanted pleiotropic effects.

In conclusion, while MYC2 represents a powerful regulatory node for enhancing secondary metabolism and stress tolerance, its complex feedback mechanisms and broad physiological roles necessitate precision engineering. A deeper understanding of MYC2 spatiotemporal activity and interaction networks will be essential to fully exploit its potential in developing resilient and productive crops.

5. Conclusions

Extensive research on MYC2 has significantly advanced our understanding of plant responses to environmental stresses and the regulation of secondary metabolism. MYC2 functions as a central regulatory hub, integrating multiple signaling pathways—especially those mediated by JA to modulate gene expression associated with plant adaptation. This review highlights the pivotal roles of MYC2 in promoting secondary metabolite biosynthesis and enhancing stress resilience, traits that are essential for sustainable agriculture. However, efforts to manipulate MYC2-mediated pathways face several challenges, including trade-offs between growth and defense, as well as the risk of inducing potentially deleterious gene expression. Addressing these limitations will require precise, spatiotemporal regulation rather than constitutive overexpression. Future research should focus on identifying MYC2 interaction partners within specific genetic and epigenetic networks, as well as elucidating its regulation of key metabolic pathways. The integration of genome editing technologies such as CRISPR-Cas9 with multi-omics approaches offers promising avenues for fine-tuning MYC2 function across diverse plant species. Leveraging advanced genetic, molecular, and biotechnological tools to optimize MYC2 activity could facilitate a more effective balance between plant growth and defense responses, ultimately contributing to enhanced crop productivity and resilience.