The root-knot nematode (Meloidogyne incognita) is a highly specialized and polyphagous plant-pathogenic nematode. Its second-stage juveniles can penetrate plant root apical meristems via stylets. Thereafter, they migrate within the plant and establish parasitic relationships with vascular tissues, leading to the formation of giant cells and production of galls (Fuller et al., 2008). Moreover, the cytoskeletal system of giant cells undergoes rearrangement during its altered development (Jammes et al., 2005). The expression levels of five ADF genes (i.e., AtADF2–AtADF6) were higher in the galls of infected roots compared with those of an uninfected control. Of these genes, AtADF2 exhibited a nearly three-fold increase in expression 2–3 weeks following nematode infection, and its expression was concentrated in giant cells. Moreover, AtADF2 knockdown in Arabidopsis increased the bundling of actin filaments, resulting in delayed giant cell development and decreased nematode reproduction. Thus, these findings imply that AtADF2 positively regulates plant resistance to root-knot nematodes (Clément et al., 2009). Similarly, in cucumber five of eight CcADF genes demonstrated increased expression in nematode-induced galls, suggesting that CcADF genes may facilitate nematode feeding on cucumber roots (Liu et al., 2016).

Aphids (Hemiptera: Aphididae), a diverse family of ~250 different species, are pests who feed on plants and affect plant growth and productivity via removing nutrients from sieve elements, altering source–sink relationships, and spreading viral diseases (Goggin, 2007). Arabidopsis atadf3 mutants were more susceptible to green peach aphids (GPAs; Myzus persicae Sülzer) infestation compared with wild type plants. GPAs fed faster and for a longer duration on atadf3 mutants, and their populations could therefore reproduce more quickly. Introducing AtADF3 into atadf3 mutant plants rescued the resistance to GPAs, indicating that AtADF3 has a critical role in limiting GPAs infestation (Mondal et al., 2018). By monitoring aphid feeding behavior, the authors found that the AtADF3 expression hinders with the ability of GPAs to find and feed from sieve elements. PAD4 (phytoalexin-deficient 4) is an important regulatory factor in Arabidopsis defense against peach aphids, negatively regulating aphid feeding and fecundity (Louis et al., 2010). Further research confirmed that PAD4 is a critical downstream player of the AtADF3-dependent defense mechanism (Mondal et al., 2018).

The corn borer is a major maize pest in many regions of the world, where it severely affects its yield by feeding on organs such as leaves, stems, and male and female inflorescences (Meihls et al., 2012). A recent genome-wide association analysis revealed that ZmADF4 is significantly associated with resistance to the Mediterranean corn borer (Sesamia nonagrioides) in the stem (Samayoa et al., 2015). The biological function of ZmADF4 in maize resistance to corn borer is worth further exploration.

Powdery mildew is a obligate biotrophic fungal pathogen that seriously threatening over 10,000 plant species, including crops, vegetables, trees, and ornamental plants (Hirose et al., 2005; Hückelhoven and Panstruga, 2011). ADF genes from different plants have been found to play different roles in regulating powdery mildew resistance. In Arabidopsis, the four Group I AtADF genes have been found to play a negative regulatory role regarding resistance to Golovinomyces orontii (G.orontii), with AtADF4 exhibiting the most significant effect. In atadf4 mutants and atadf1-4 quadruple knockdown plants, researchers identified an accumulation of hydrogen peroxide and cell-specific death at the sites of G.orontii infection. In addition, they also found an increase in the abundance of microfilaments, and the plants show enhanced resistance against powdery mildew (Inada et al., 2016). RPW8.2 (resistance to powdery mildew 8.2) is an atypical mildew resistance protein found in Arabidopsis (Xiao et al., 2001). Overexpression of AtADF6 (belongs to group IV) inhibited its localization to the membrane surrounding the powdery mildew fungal haustorium, which is required for inducing resistance against powdery mildew. Thus, this evidence indicates that AtADF6 may play a negative role toward powdery mildew resistance (Wang et al., 2009b). In contrast, the overexpression of AtADF5 (belongs to group III) had no effect on RPW8.2 localization, implying the existence of functional diversification among ADF members in plant response to powdery mildew (Wang et al., 2009b). It is reasonable to presume that the functional diversification between AtADF5 and AtADF6 may resulted from their difference in biochemical activity (Nan et al., 2017) or expression profile (Dong et al., 2001). Ectopic expression of HvADF3 in barley (Hordeum vulgare) epidermal cells was found to disrupt the integrity of the actin cytoskeleton in cells. This in turn enhanced fungal entry and lead to increased susceptibility to the barley pathogen Blumeria graminis f. sp. hordei (Bgh) (Miklis et al., 2007). Moreover, transient overexpression of AtADF1, AtADF5, AtADF6, AtADF7, and AtADF12 was found to significantly increase the entry rate of Bgh in barley, while the overexpression of AtADF2, AtADF3, AtADF4 and AtADF9 had no significant effect.

In wheat, stripe rust caused by Puccinia striiformis f. sp. tritici is a widespread and devastating disease (Hovmøller, 2007). Different wheat ADF genes were found to exhibit varied response patterns against different physiological races of this pathogen, and may therefore play different roles in regulating stripe rust resistance. For example, the avirulent race CYR23 strongly induced the expression of TaADF4 and TaADF7 in wheat, while the virulent race CYR31 induced their expression to a lesser extent. Silencing TaADF4 and TaADF7 in wheat lines inoculated with CYR23 led to significant changes in microfilament structures, reduced accumulation of reactive oxygen species (ROS), and weakened hypersensitive reactions. Taken together, these effects indicate that TaADF4 and TaADF7 positively regulate wheat resistance against non-adapted races of stripe rust by modulating microfilament dynamics (Fu et al., 2014; Zhang et al., 2017). In contrast to the response patterns of these genes, TaADF3 showed elevated expression levels upon CYR31 induction but showed significantly decreased expression levels following CYR23 inoculation. Silencing TaADF3 enhanced wheat resistance to CYR31 while reducing both ROS accumulation and hypersensitive reactions (Tang et al., 2016).

Verticillium wilt is mainly caused by Verticillium dahliae or Verticillium alboatrum, two soil-borne vascular fungal pathogens that severely affect cotton production (Klosterman et al., 2009). After infection by Verticillium dahliae, the expression of GhADF6, a gene homologous to AtADF6 found in upland cotton (Gossypium hirsutum), was downregulated in root epidermal cells. Silencing of GhADF6 increased the abundance of microfilaments in root epidermal cells, and the plants showed enhanced resistance to Verticillium dahlia. Thus, GhADF6 likely plays a negative regulatory role with respect to cotton verticillium wilt resistance (Sun et al., 2021). Taken together, these findings highlight the complex regulatory roles of ADF genes regarding plant defense against fungal diseases.

Innate immunity is the first line of host defense against microbial invasion and is evolutionally conserved in all multicellular organisms, which is activated by pattern-recognition receptors (PRRs) that recognize microbe-associated molecular patterns (MAMPs) (Deng et al., 2020). MAMPs mediated microfilament rearrangement, as featured by increased abundance and remodeling of microfilament, plays an important role in plant innate immune signal transduction (Henty-Ridilla et al., 2014). Within minutes of treatment with a bacterial MAMP, elf26 (a conserved 26-amino acid peptide from bacterial elongation factor), a dose- and time-dependent increase in actin filament abundance was detected in epidermal cells throughout the Arabidopsis hypocotyl. However, actin architecture and dynamics in an atadf4 mutant fail to respond to elf26 treatment, suggested that AtADF4 plays a key role in modulating actin dynamics by participating in innate immune signal transduction caused by bacteria in plants (Henty-Ridilla et al., 2014).

Pseudomonas syringae pv tomato (Pst) is a hemibiotrophic bacterial pathogen. The Arabidopsis mutant atadf4 shows abnormal microfilament dynamics and increased susceptibility to that of Pst DC3000 expressing the incompatible effector AvrPphB, but not to strains expressing AvrRps2 or AvrB. Moreover, a transgenic experiment showed that AtADF4 is able to restore the resistance that is compromised in the atadf4 mutant, thereby indicating that AtADF4 is required for the resistance to Pst DC3000 expressing AvrPphB (Tian et al., 2009). RPS5 (resistance to Pseudomonas syringae 5) is a gene that encodes a resistance protein capable of recognizing AvrPphB and activating downstream defense signals in Arabidopsis (Chisholm et al., 2006). Subsequent studies have reported that the increased susceptibility of atadf4 mutant to Pst DC3000 expressing AvrPphB is associated with decreased RPS5 expression, suggesting that AtADF4 may regulate plant resistance to Pst DC3000 expressing AvrPphB via the coordinated regulation of microfilament dynamics and R-gene transcription (Porter et al., 2012).

Rhizobia is a class of gram-negative soil bacteria that includes Rhizobium, Bradyrhizobium, Sinorhizobium, and Azorhizobium. These bacteria can form symbiotic nitrogen-fixing nodules with leguminous plants and increase nitrogen fixation in arable fields by as much as 30% (Mus et al., 2016). PvADFE is one of the nine ADF genes found in common bean, primarily expressed in roots and nodules inoculated with Rhizobium tropici (Ortega-Ortega et al., 2020). In addition, PvADFE silencing increases the number and size of nodules and enhances nitrogen fixation activity. Conversely, the overexpression of this gene resulted in the opposite phenotype. In addition, the expression levels of two genes related to nodulation development and signaling, NIN and ENOD2, were significantly decreased in the roots of plants overexpressing PvADFE, thereby indicating that PvADFE plays a negative regulatory role in rhizobial infection and nodulation of common bean (Ortega-Ortega et al., 2020).

Cold stress, including chilling (cold temperatures of above 0°C) and freezing stress (below 0°C), causes plant growth to slow down, stagnate, and retrogress, thereby reducing the yield (Zhang et al., 2019). Aside from membrane rigidification, ROS accumulation, protein destabilization, and metabolic disequilibrium, cold stress has also been reported to disrupt the microfilament of plant cells and interfere with all cellular processes (Fan et al., 2015; Liu et al., 2018). The expression profiles of AtADF genes in response to temperature stress had been reported to diversely among different groups. The expression levels of AtADF genes of Group I, III, and IV have been found to be significantly induced by cold or heat stress, with two group III ADF genes (i.e., AtADF5 and AtADF9) responding the most strongly (Fan et al., 2015; Fan et al., 2016). In Arabidopsis plants exposed to cold stress, the survival rate of atadf5 mutants significantly decreased relative to the wild type. Moreover, mutant plants showed disordered actin cytoskeleton in root epidermal cells, suggesting that AtADF5 plays an important role in mediating cold stress tolerance in Arabidopsis (Zhang et al., 2021). In the face of cold stress, plants depend on C-repeat binding factors (CBFs) as their key molecular switches (Liu et al., 2018). CBFs can activate the expression of AtADF5 by binding to CRT/DRE elements in its promoter, and AtADF5 can in turn regulate dynamic changes in the actin cytoskeleton to modulate the cold response of Arabidopsis plants (Zhang et al., 2021). In freezing-tolerant wheat cultivars, the ADF gene Wcor719 can be specifically induced by exposure to low temperatures, while the expression levels of this gene do not significantly change in freezing-sensitive cultivars. Moreover, its expression level is also insensitive to high temperature, salt stress, mechanical damage, and abscisic acid (ABA) (Danyluk et al., 1996; Ouellet et al., 2001). In contrast, wheat TaADF4 is induced by heat stress, but its expression levels are significantly decreased in response to low temperature or salt stress (Zhang et al., 2017). A genome-wide analysis showed that 25 TaADF genes exist in the genome of the “Chinese Spring” wheat cultivar, and that cold stress can affect the expression levels of seven TaADF genes, six of which (i.e., TaADFs 13, 16, 17, 18, 21, and 22) are upregulated (Xu et al., 2021). In Arabidopsis, the heterologous expression of TaADF16, the most highly expressed and upregulated ADF gene in response to cold stress, can enhance plant cold stress resistance by accelerating ROS scavenging and by altering osmotic regulation in cells (Xu et al., 2021). Moreover, the expression levels of seven cold stress-responsive genes were found to be significantly higher in a TaADF16-overexpressing line than in the wild type regardless of whether the transgenic Arabidopsis plants were exposed to cold conditions. This indicates that the overexpression of TaADF16 can generally induce the expression of cold-related genes (Xu et al., 2021). Antarctic hairgrass is the only monocotyledonous flowering plant in Antarctica and its genome contains eight ADF genes. Cold stress can induce the expression of five DaADF genes, with DaADF3 showing the most significant cold stress response (Byun et al., 2021). In rice, plants that overexpress DaADF3 exhibit improved cold stress resistance, as measured via survival rate, leaf chlorophyll content, and electrolyte leakage along with changes in microfilament organization in the root tips (Byun et al., 2021).

Heat stress is commonly defined as the increase in temperature beyond a specific threshold level for a duration that is adequate to induce irreversible harm to the growth and development of plants (Bita and Gerats, 2013). The effects of heat stress on plants and cells are numerous. For example, high temperatures alter membrane fluidity and denaturate proteins which impair enzyme function (Malerba et al., 2010). Recently, ADF genes have also been found to be involved in plant tolerance to high temperatures. For example, AtADF1 expression was repressed by high temperatures, and atadf1 mutant seedlings exhibited greater actin filament stability and faster growth than the wild type (Wang L. et al., 2023). Conversely, AtADF1 overexpression showed the opposite phenotype. Further experiments revealed that AtADF1 transcription is regulated by AtMYB30, a key transcription factor involved in responses to various forms of abiotic stress, including heat (Liao et al., 2017). This finding indicates that AtADF1 is a target gene in the AtMYB30-mediated plant response to abiotic stress (Wang B. et al., 2023). Similarly, the authors found that BrADF1 from Chinese cabbage (Brassica rapa), a gene that is highly homologous to AtADF1, regulates F-actin dynamics and plant tolerance to heat stress in a manner similar to that of AtADF1 (Wang L. et al., 2023).

Salt stress refers to the adverse effect of excessive soluble salts in soil on plant growth and development, which has both osmotic and ionic or ion-toxicity effects on cells (Zhu, 2016). More than one third of the world’s irrigated lands are affected by salinization, a worldwide problem that threatens the growth and yield of crops (Zhao et al., 2020). In Arabidopsis, the expression levels of AtADF1 rapidly increase in response to salt stress, and the survival rate of the atadf1 mutant decreases significantly compared with the wild type under salt stress. Moreover, mutant plants exhibit cytoskeletal changes, including increased microfilament bundles in cells, while AtADF1-overexpressing plants exhibit opposite macroscopic and microscopic phenotypes. These results indicate that AtADF1 positively regulates plant salt tolerance by promoting actin depolymerization (Wang et al., 2021). Moreover, AtMYB73 is a negative regulatory factor for salt stress in Arabidopsis (Kim et al., 2013), and further experiments have revealed that AtMYB73 negatively regulates AtADF1 expression (Wang et al., 2021). Therefore, AtADF1 is likely an important player in the AtMYB73-mediated salt stress response pathway (Wang et al., 2021). Smooth cordgrass (Spartina alterniflora) is a perennial grass halophyte that has adapted to salt and drought conditions owing to specific alleles for genes involved in stress tolerance (Baisakh et al., 2008). The SaADF2 of smooth cordgrass is homologous to the OsADF2 of rice. Although the sequence similarity between their proteins exceeds 95%, six amino acid differences (i.e. the 6^th Ser, 19^th Asp, 25^th Leu, 118^th Gln, 132^nd Pro and 133^rd Thr in OsADF2 were substituted by Thr, Asn, His, His, Ser and Ser in SaADF2, respectively) may responsible for the substantial differences in their three-dimensional structures (Sengupta et al., 2019). Biochemical analysis revealed that SaADF2 displays greater actin-binding affinity and can depolymerize microfilaments more efficiently than OsADF2, which enhances cellular actin dynamics in cells. In rice, SaADF2 overexpression engenders greater drought and salt tolerance compared with that in the wild type and OsADF2-overexpression lines (Sengupta et al., 2019). A detailed biochemical investigation is required to determine the specific amino acid(s) that play a critical role in ADF’s biochemical activity. This knowledge could present an opportunity to utilize genome-editing technology for performing site-specific mutations, enabling the manipulation of ADF activity in crop breeding practices for enhancing stress tolerance.

Osmotic stress, often caused by drought and high salinity, occurs when soil contains excess soluble salt that prevents water absorption of plants (Yoshida et al., 2014). Microfilament cytoskeleton had been confirmed to participates, and plays a crucial role, in responses to osmotic stress in plants (Wang et al., 2010). In Arabidopsis seedlings subjected to osmotic stress, expression level of AtADF4 considerably increased. In addition, the survival rate of atadf4 mutants was higher than that of the wild type, while the survival rate of AtADF4-overexpressing lines decreased. Thus, these results indicate that AtADF4 plays a negative regulatory role in the plant response to osmotic stress (Yao et al., 2022). Further experiments demonstrated that a phosphopeptide-binding protein, 14-3-3κ, acts as an upstream regulator of AtADF4 to regulate the Arabidopsis response to osmotic stress (Yao et al., 2022). Root hairs are important organs for plants to absorb nutrients and water. Osmotic stress can induce AtADF7 expression, which in turn is responsible for inhibiting the expression of the actin-bundling protein VILLIN1 (VLN1) in root cells, thereby reducing microfilament bundles in root cells and promoting root hair growth. These findings indicate that the AtADF7–VLN1 pathway is essential for root hair formation under osmotic stress tolerance, and plays critical role in enhancing plant osmotic stress tolerance (Bi et al., 2022).

Drought is an adverse environmental stress that hampers normal growth, disrupts water relations, and decreases water-use efficiency in plants. To adapt to drought stress, plants have developed intricate mechanisms, one of which involves regulating the opening and closing of stomata (Saradadevi et al., 2017). Stomata are pores found on the epidermis of aerial parts of plants and are responsible for absorbing carbon dioxide and releasing water vapor (Jiang et al., 2012). The stomatal aperture is finely tuned to prevailing environmental conditions by a pair of guard cells surrounding each pore (Schroeder et al., 2001). Stomatal movement mediated by ABA is particularly important for plant adaptations to drought conditions. Microfilament dynamics play a crucial role in regulating the opening and closing of the stomata, involving the alteration of the radial orientation of the actin filaments during open stomata changes to a longitudinal orientation characteristic of closed stomata during stomatal closure (Zhao et al., 2011).

In Arabidopsis, AtADF1 overexpression leads to disorganized microfilament bundles in guard cells, in turn resulting in abnormal stomatal closure following ABA treatment (Dong et al., 2001). Arabidopsis casein kinase 1-like protein 2 (CKL2) plays an important regulatory role in ABA- and drought-induced stomatal closure. CKL2 can inhibit the depolymerization activity of AtADF4 via phosphorylation, thereby rendering the microfilament cytoskeleton more stable in guard cells and regulating stomatal opening or closing (Zhao et al., 2016). ABA and drought stress also induce AtADF5 expression in Arabidopsis seedlings. Compared with the wild type, an atadf5 mutant showed reduced microfilament bundles in cells, delayed stomatal closure, intensified leaf dehydration, and decreased survival rates under drought conditions (Qian et al., 2019). Further biochemical experiments demonstrated that DPBF3, an ABA-responsive element-binding factor (ABF/AREB), can activate AtADF5 expression via ABA-responsive core elements in its promoter region. AtADF5 regulates stomatal movement by modulating the rearrangement of microfilament structures through its F-actin bundling activity, which improves plants’ adaptability to drought stress. Therefore, AtADF5 is an important player in the ABF/AREB-mediated pathway facilitating plant responses to drought stress (Qian et al., 2019). In Populus euphratica, PeABF3 is a transcription factor involved in ABA signaling response and its expression is induced by drought and ABA. PeABF3 can activate the expression of PeADF5, which facilitates ABA-induced stomatal movement by promoting actin cytoskeletal rearrangement and enhancing drought resistance (Yang et al., 2020). Furthermore, in rice the exogenous application of ABA or various stress conditions induces OsADF3 expression in the root tips and lateral roots. Moreover, the heterologous expression of OsADF3 in Arabidopsis enhanced its drought tolerance, as evidenced by improved germination rate, primary root length, and survival rate. In addition, several drought-tolerance responsive genes are upregulated under drought stress, suggesting that OsADF3 may exert regulatory effects upstream of these genes (Huang et al., 2012). The above studies indicate that plant ADF genes are important factors in regulating stomatal movement and play an important role in enhancing plant drought resistance. Further exploration and research on these drought resistant ADF genes and homologous ADF genes in other plants, especially crops, will provide important genetic resources for the cultivation of drought resistant crops.