Shanjin Huang

Plasmodesmata (PD)‐mediated intercellular communication is critical for plant stress adaptation, yet its dynamic regulation by the actin cytoskeleton under abiotic stress remains poorly characterised. Here, we show that rice OsFH14 modulates salt‐induced PD permeability to enhance salt tolerance. Through CRISPR‐Cas9 mutagenesis and subcellular localisation analyses, we first demonstrate that OsFH14 specifically localises to PD and is strictly required for salt stress‐induced increases in PD permeability. Osfh14 mutants exhibit reduced PD permeability and enhanced sensitivity to NaCl treatment. Mechanistic investigations show that OsFH14 functions as a specialised actin‐bundling protein that stabilises existing actin filaments without nucleation or capping activity, which suggests that OsFH14 probably regulates salt tolerance by maintaining PD‐associated actin cytoskeletal integrity. Supporting this hypothesis, pharmacological disruption of actin dynamics using Latrunculin B (LatB) phenocopies Osfh14 mutant defects, specifically impairing both PD permeability regulation and salt tolerance. Conversely, pretreatment of Osfh14 mutants with the actin filament stabiliser Jasplakinolide (Jasp) recovers PD permeability after salt treatment and they exhibit salt tolerance phenotypes indistinguishable from wild‐type (WT) plants. These findings establish a causal link between OsFH14‐mediated actin bundling at PD and salt tolerance, highlighting PD‐associated actin dynamics as a key adaptive strategy for crop salt tolerance.

Plants have developed fine-tuned cellular mechanisms to respond to intracellular and extracellular signals, which are crucial for their development and adaptation to the ever-changing environment. These cellular responses are often mediated by the highly dynamic cytoskeletal network, including actin filaments and microtubules in plant cells. The cytoskeleton quickly rearranges its organization, through dynamic disassembly and assembly, in response to intrinsic and extrinsic stimuli, which further impacts a variety of cellular activities. In this review, we summarized the roles and regulatory mechanisms of the cytoskeleton in cell morphogenesis, organogenesis, and plant environmental adaptation. We next discussed the relationship between the cytoskeleton and the upstream signals, as well as the research on the cytoskeleton and its relevance in agriculture. Finally, we highlight the key questions remaining to be investigated and put forward some future perspectives in the field.

Summary: Villins are versatile, multifunctional actin regulatory proteins. They promote actin stabilization and remodeling mainly via their actin bundling and Ca²⁺‐dependent severing activities, respectively. Arabidopsis subclass II and III villins normally coexist in cells, but the biological significance of their coexistence remains unknown. Here we demonstrate that subclass II villin binds to Ca²⁺ with high affinity and exhibits strong severing but weak bundling activity compared to subclass III villin. Subclass II villin plays a dominant role in promoting actin remodeling, which requires its Ca²⁺‐dependent severing activity. Subclass II villin is also strictly required for physiological processes including oriented organ growth and stress tolerance. By comparison, subclass III villin binds to Ca²⁺ with low affinity and exhibits weak severing but strong bundling activity, and acts as the major player in controlling actin stabilization and organization. Thus, we demonstrate that multifunctional villin isovariants have diverged biochemically to ensure exquisite control of the actin cytoskeleton to meet different cellular needs in plants. This study provides new insights into the role of villins in fine‐tuning actin dynamics and plant development.

SUMMARY: Being a bona fide actin bundler, Arabidopsis villin5 (VLN5) plays a crucial role in regulating actin stability and organization within pollen tubes. Despite its significance, the precise mechanism through which VLN5 bundles actin filaments has remained elusive. Through meticulous deletion analysis, we have unveiled that the link between gelsolin repeat 6 (G6) and the headpiece domain (VHP), rather than VHP itself, is indispensable for VLN5‐mediated actin bundling. Further refinement of this region has pinpointed a critical sequence spanning from Val763 to Ser823, essential for VLN5's actin‐bundling activity. Notably, the absence of Val763‐Ser823 in VLN5 results in decreased filamentous decoration within pollen tubes and a diminished ability to rescue actin bundling defects in vln2vln5 mutant pollen tubes compared to intact VLN5. Moreover, our findings highlight that the Val763‐Ser823 sequence harbors a binding site for F‐actin, suggesting that this linker‐based F‐actin binding site, in conjunction with the F‐actin binding site localized in G1–G6, enables a single VLN5 to concurrently bind to two adjacent actin filaments. Therefore, our study unveils a novel mechanism by which VLN5 bundles actin filaments.

Peptide/receptor-like kinase-mediated signaling plays a pivotal role in male-female interactions, particularly in guiding the growth of pollen tubes, which is essential for fertilization in flowering plants. However, the downstream cellular events within the pollen tube guidance signaling network remain largely unexplored. In this study, actin dynamics in pollen tubes growing semi-in vitro were directly visualized, and it was discovered that AtLURE1s-induced pollen tube growth guidance is intricately regulated by actin assembly and disassembly. The observations revealed that apical actin filaments undergo a process of rapid depolymerization and repolymerization at the leading side, which in turn facilitates the reconstruction of the apical actin structure, enabling the pollen tube to grow in a new direction. Notably, it was observed that actin depolymerization and subsequent repolymerization occur on the leading side before any morphological changes in the pollen tube. Furthermore, it was found that the loss of function of ADF10 results in the accumulation of apical actin filaments, altering the turning behavior of the pollen tube. These findings suggest that rapid actin depolymerization at the leading side and the subsequent reconstruction of the apical structure occur early and are crucial for successful pollen tube turning.

Center dot Grain notching is a common deformation that decreases rice (Oryza sativa) quality; however, the underlying molecular basis causing grain notching remains unclear. center dot We report mechanisms underlying grain notching in Small and notched grain (Sng) mutants, which contained an arginine to histidine substitution at amino acid position 422 (R422H) of the alpha-tubulin protein OsTUBA3. The R422H mutation decreased cell length and increased cell width/height of glumes and caryopses, but led to elongated caryopses compressed within shortened glumes, thus giving rise to notched and small grains. Glume and caryopsis cells had different dimensional orientations relative to the directions of organ elongation. Thus, the abnormal cell expansion induced in glumes and caryopses by the R422H mutation had different effects on elongation of these organs. center dot The R422H mutation in OsTUBA3 compromised beta-tubulin binding and led to formation of defective heterodimers. This in turn affected tubulin incorporation and microtubule (MT) nucleation and regrowth, consequently leading to MT instability and reducing the transverse orientation. The defective MT dynamics affected cell expansion and shape, causing different alterations in glume and caryopsis dimensions and resulting in grain notching. center dot These data indicate that Arg422 in OsTUBA3 is crucial for MT dynamics and that substitution with His causes grain notching, reducing grain quality and yield. These findings offer valuable insights into the molecular regulation underlying grain development in rice.

The actin cytoskeleton is one of the targets of the pH gradient in tip-growing cells, but how cytosolic pH regulates the actin cytoskeleton remains largely unknown. We here demonstrate that Arabidopsis ADF7 and ADF10 function optimally at different pH levels when disassembling actin filaments. This differential pH sensitivity allows ADF7 and ADF10 to respond to the cytosolic pH gradient to regulate actin dynamics in pollen tubes. ADF7 is an unusual actin-depolymerizing factor with a low optimum pH in in vitro actin depolymerization assays. ADF7 plays a dominant role in promoting actin turnover at the pollen tube apex. ADF10 has a typically high optimum pH in in vitro assays and plays a dominant role in regulating the turnover and organization of subapical actin filaments. Thus, functional specification and cooperation of ADF isovariants with different pH sensitivities enable the coordination of the actin cytoskeleton with the cytosolic pH gradient to support pollen tube growth.

As the stimulus-responsive mediator of actin dynamics, actin-depolymerizing factor (ADF)/cofilin is subject to tight regulation. It is well known that kinase-mediated phosphorylation inactivates ADF/cofilin. Here, however, we found that the activity ofArabidopsisADF7 is enhanced by CDPK16-mediated phosphorylation. We found that CDPK16 interacts with ADF7 bothin vitroandin vivo, and it enhances ADF7-mediated actin depolymerization and severingin vitroin a calcium-dependent manner. Accordingly, the rate of actin turnover is reduced incdpk16pollen and the amount of actin filaments increases significantly at the tip ofcdpk16pollen tubes. CDPK16 phosphorylates ADF7 at Serine128 bothin vitroandin vivo, and the phospho-mimetic mutant ADF7^S128Dhas enhanced actin-depolymerizing activity compared to ADF7. Strikingly, we found that failure in the phosphorylation of ADF7 at Ser128 impairs its function in promoting actin turnoverin vivo, which suggests that this phospho-regulation mechanism is biologically significant. Thus, we reveal that CDPK16-mediated phosphorylation up-regulates ADF7 to promote actin turnover in pollen.