Secondary Cell Wall Research Articles

Transcription factor PagERF110 inhibits xylem differentiation by direct regulating PagXND1d in poplar

Ethylene-responsive factor (ERF) transcription factors (TFs) play an important role in plant development and growth, while their underlying function in xylem development is not very clear in poplar. In this study, a nucleus-located ERF gene, PagERF110 was isolated from Populus alba × P. glandulosa, which displayed a relatively high expression in phloem and xylem. The transgenic poplar overexpressing PagERF110 showed less secondary xylem deposition than wild-type (WT). And the contents of cellulose, xylose and lignin in the cell wall of transgenic lines were less than those of WT. Transcriptome analyses showed that PagERF110 could activate the expression of a few downstream genes related to secondary cell wall (SCW) formation. Among them, poplar NAC domain XYLEM NAC DOMAIN1 (PagXND1d) is a key downstream gene, whose overexpression inhibits the development of secondary xylem. And the evidences from yeast one-hybrid (Y1H), luciferase assay and chromatin immunoprecipitation (ChIP) assay all proved that PagERF110 directly regulates the expression of PagXND1d, and overexpression PagXND1d reduces secondary xylem deposition in transgenic poplar. This study enriches the roles of ERFs in SCW formation in poplar, which further provides references for revealing regulatory networks of TFs in xylem development in woody plants.

GhMYB52 Like: A Key Factor That Enhances Lint Yield by Negatively Regulating the Lignin Biosynthesis Pathway in Fibers of Upland Cotton (Gossypium hirsutum L.).

In the context of sustainable agriculture and biomaterial development, understanding and enhancing plant secondary cell wall formation are crucial for improving crop fiber quality and biomass conversion efficiency. This is especially critical for economically important crops like upland cotton (Gossypium hirsutum L.), for which fiber quality and its processing properties are essential. Through comprehensive genome-wide screening and analysis of expression patterns, we identified a particularly high expression of an R2R3 MYB transcription factor, GhMYB52 Like, in the development of the secondary cell wall in cotton fiber cells. Utilizing gene-editing technology to generate a loss-of-function mutant to clarify the role of GhMYB52 Like, we revealed that GhMYB52 Like does not directly contribute to cellulose synthesis in cotton fibers but instead represses a subset of lignin biosynthesis genes, establishing it as a lignin biosynthesis inhibitor. Concurrently, a substantial decrease in the lint index, a critical measure of cotton yield, was noted in parallel with an elevation in lignin levels. This study not only deepens our understanding of the molecular mechanisms underlying cotton fiber development but also offers new perspectives for the molecular improvement of other economically important crops and the enhancement of biomass energy utilization.

Xylans as a promising prebiotic agent: a brief review

Xylans are the most abundant hemicelluloses in nature, constituting components of the secondary cell wall of plant cells. They are polysaccharides with versatile properties, which can be used in various industrial sectors. Several biological activities have been identified in different xylans, such as antioxidant, antitumor, antimicrobial, emulsifying, anticoagulant, and immunomodulatory activities. Among these applications of xylans, its use as a prebiotic stands out, composing functional foods. Xylans form non-digestible dietary fibers that serve as a source of energy for beneficial microorganisms, modifying the intestinal microbiome and bringing a general improvement to the health status of those who consume them. The literature reports the use of xylooligosaccharides as promoters of the growth of probiotic microorganisms, such as species of Lactobacillus and Bifidobacterium, in addition to treating dysbiosis and reducing intestinal inflammation. Due to their structural diversity, abundance in nature, and a wide range of biological properties, xylans are valuable components that can be used to create various products aimed at enhancing human health.

Seed morphology in the genus Matthiola R. Br. (Brassicaceae): Identification of species and systematic implications

AbstractSeed coat morphology of 34 species of Matthiola R.Br. has been studied using light and scanning electron microscopy to assess their investigative value for systematic studies. Macro‐ and micromorphological characters, including seed shape, color, size, wing, and radicle position relative to cotyledons, epidermal cell shape, anticlinal boundaries, and periclinal cell wall are presented. Seed shape, color, and radicle position relative to cotyledons have limited taxonomic significance, as their variation is unhelpful. There against, seed size, wing, and seed coat ornamentation present informative characteristics that can be used professionally in distinguishing the studied taxa. Six different types of seed anticlinal cell wall boundaries are recognized and five different outer periclinal cell wall shapes are described. The sculpture of the secondary cell wall varies from striate, micropapillate to microreticulate, and smooth to fine folds. The taxonomic and phylogenetic implications of seed coat micromorphology were compared with those of the existing gross morphological and molecular data. Seed character analysis offered useful data for evaluating the taxonomy of Matthiola both on subgeneric and sectional levels. Seed micromorphological characters provide evidence that M. odoratissima and M. ovatifolia should be preserved as separate species. In addition, M. ghorana is not conspecific with M. graminea. Furthermore, M. tricuspidata was unique in the seed characters. A key to the identification of the examined taxa based on seed characters is provided.

Identification of the CesA7 Gene Encodes Brittleness Mutation Derived from IR64 Variety and Breeding for Ruminant Feeding

Rice straw presents challenges as livestock feed due to its low digestibility and the presence of chemical residues. One potential solution is to focus on breeding brittle varieties that possess disease-resistance traits. In this study, AZ1803, a brittle mutant line isolated from the IR64 mutant pool, was chosen for gene identification and breeding. The AZ1803 mutant was crossed to the TNG67 variety to generate a mapping population and to the CS11 variety for fine mapping and breeding. The gene was mapped on chr. 10 between RM467 and RM171 SSR markers and was narrowed down to RM271 and RM5392 with 600 kb proximately interval. The AZ1803 and IR64 sequencing results revealed a substitution mutant in the Exon 9th of the OsCesA7 gene, resulting in an amino acid mutation at the end of the transmembrane domain 5th of the CESA7, responsible for cellulose synthesis for the secondary cell wall. The cellulose content of AZ1803 was reduced by 25% compared with the IR64. A new brittle and disease-resistant variety was bred by using developed markers in marker-assisted selection. In addition, bending tests and bacterial blight inoculation were applied. The bacterial lesion length of the bred variety is 64% lower than that of AZ1803. The rice straw of the new variety can be used for livestock feeding, which increases farmer income and reduces pesticide residues and air pollution from straw burning.

VND Genes Redundantly Regulate Cell Wall Thickening during Parasitic Nematode Infection.

Plant parasitic root-knot nematodes are major agricultural pests worldwide, as they infect plant roots and cause substantial damages to crop plants. Root-knot nematodes induce specialized feeding cells known as giant cells (GCs) in the root vasculature, which serve as nutrient reservoirs for the infecting nematodes. Here, we show that the cell walls of GCs thicken to form pitted patterns that superficially resemble metaxylem cells. Interestingly, VASCULAR-RELATED NAC-DOMAIN1 (VND1) was found to be upregulated, while the xylem-type programmed cell death marker XYLEM CYSTEINE PEPTIDASE 1 was downregulated upon nematode infection. The vnd2 and vnd3 mutants showed reduced secondary cell wall pore size, while the vnd1 vnd2 vnd3 triple mutant produced significantly fewer nematode egg masses when compared with the wild type. These results suggest that the GC development pathway likely shares common signaling modules with the metaxylem differentiation pathway and VND1, VND2, and VND3 redundantly regulate plant-nematode interaction through secondary cell wall formation.

Irreversibility of the cell wall modification acts as a limiting factor in desiccation tolerance of Oryza sativa ssp. Indica cv MR303

Desiccation tolerance has emerged as a potential strategy for crop improvement. However, the application of desiccation tolerance in crops, particularly rice, has been challenging and limited. Studies in angiosperms suggest that desiccation in vegetative tissues shares a common set of genes, developed convergently by modifying shared pathways associated with seed desiccation. The drought-tolerant Oryza sativa ssp. indica cv MR303 has demonstrated the capability to withstand desiccation state for a short period of time. Using this variety, we aim to investigate the factors that limit rice plants from achieving desiccation tolerance. The irreversibility of the cell wall emerges as a primary factor hindering rice plants from acquiring desiccation tolerance. Analysis of cell wall composition and transcriptomic data reveals an accumulation of secondary cell wall components and the expression of cell wall-related genes involved in secondary wall formation and modification, suggesting that these factors could be underlying causes for the irreversibility of the cell wall. Such factors contribute to the rigidity and stiffness of the cell wall, leading to its irreversibility during the rehydration process. This, in turn, impedes the recovery of rice after dehydration.

Comparative Transcriptomic Analysis of Gossypium hirsutum Fiber Development in Mutant Materials (xin w 139) Provides New Insights into Cotton Fiber Development.

Cotton is the most widely planted fiber crop in the world, and improving cotton fiber quality has long been a research hotspot. The development of cotton fibers is a complex process that includes four consecutive and overlapping stages, and although many studies on cotton fiber development have been reported, most of the studies have been based on cultivars that are promoted in production or based on lines that are used in breeding. Here, we report a phenotypic evaluation of Gossypium hirsutum based on immature fiber mutant (xin w 139) and wild-type (Xin W 139) lines and a comparative transcriptomic study at seven time points during fiber development. The results of the two-year study showed that the fiber length, fiber strength, single-boll weight and lint percentage of xin w 139 were significantly lower than those of Xin W 139, and there were no significant differences in the other traits. Principal component analysis (PCA) and cluster analysis of the RNA-sequencing (RNA-seq) data revealed that these seven time points could be clearly divided into three different groups corresponding to the initiation, elongation and secondary cell wall (SCW) synthesis stages of fiber development, and the differences in fiber development between the two lines were mainly due to developmental differences after twenty days post anthesis (DPA). Differential expression analysis revealed a total of 5131 unique differentially expressed genes (DEGs), including 290 transcription factors (TFs), between the 2 lines. These DEGs were divided into five clusters. Each cluster functional category was annotated based on the KEGG database, and different clusters could describe different stages of fiber development. In addition, we constructed a gene regulatory network by weighted correlation network analysis (WGCNA) and identified 15 key genes that determined the differences in fiber development between the 2 lines. We also screened seven candidate genes related to cotton fiber development through comparative sequence analysis and qRT-PCR; these genes included three TFs (GH_A08G1821 (bHLH), GH_D05G3074 (Dof), and GH_D13G0161 (C3H)). These results provide a theoretical basis for obtaining an in-depth understanding of the molecular mechanism of cotton fiber development and provide new genetic resources for cotton fiber research.

Precocious cell differentiation occurs in proliferating cells in leaf primordia in Arabidopsis angustifolia3 mutant.

During leaf development, the timing of transition from cell proliferation to expansion is an important factor in determining the final organ size. However, the regulatory system involved in this transition remains less understood. To get an insight into this system, we investigated the compensation phenomenon, in which the cell number decreases while the cell size increases in organs with determinate growth. Compensation is observed in several plant species suggesting coordination between cell proliferation and expansion. In this study, we examined an Arabidopsis mutant of ANGUSTIFOLIA 3 (AN3)/GRF-INTERACTING FACTOR 1, a positive regulator of cell proliferation, which exhibits the compensation. Though the AN3 role has been extensively investigated, the mechanism underlying excess cell expansion in the an3 mutant remains unknown. Focusing on the early stage of leaf development, we performed kinematic, cytological, biochemical, and transcriptome analyses, and found that the cell size had already increased during the proliferation phase, with active cell proliferation in the an3 mutant. Moreover, at this stage, chloroplasts, vacuoles, and xylem cells developed earlier than in the wild-type cells. Transcriptome data showed that photosynthetic activity and secondary cell wall biosynthesis were activated in an3 proliferating cells. These results indicated that precocious cell differentiation occurs in an3 cells. Therefore, we suggest a novel AN3 role in the suppression of cell expansion/differentiation during the cell proliferation phase.

Transcriptional dynamics reveals the asymmetrical events underlying graft union formation in pecan (Carya illinoinensis).

Grafting is a widely used technique for pecan propagation; however, the background molecular events underlying grafting are still poorly understood. In our study, the graft partners during pecan [Carya illinoinensis (Wangenh.) K. Koch] graft union formation were separately sampled for RNA-seq, and the transcriptional dynamics were described via weighted gene co-expression network analysis. To reveal the main events underlying grafting, the correlations between modules and grafting traits were analyzed. Functional annotation showed that during the entire graft process, signal transduction was activated in the scion, while messenger RNA splicing was induced in the rootstock. At 2 days after grafting, the main processes occurring in the scion were associated with protein synthesis and processing, while the primary processes occurring in the rootstock were energy release-related. During the period of 7-14 days after grafting, defense response was a critical process taking place in the scion; however, the main process functioning in the rootstock was photosynthesis. From 22 to 32 days after grafting, the principal processes taking place in the scion were jasmonic acid biosynthesis and defense response, whereas the highly activated processes associated with the rootstock were auxin biosynthesis and plant-type secondary cell wall biogenesis. To further prove that the graft partners responded asymmetrically to stress, hydrogen peroxide contents as well as peroxidase and β-1,3-glucanase activities were detected, and the results showed that their levels were increased in the scion not the rootstock at certain time points after grafting. Our study reveals that the scion and rootstock might respond asymmetrically to grafting in pecan, and the scion was likely associated with stress response, while the rootstock was probably involved in energy supply and xylem bridge differentiation during graft union formation.

Cotton sphingosine kinase GhLCBK1 participates in fiber cell elongation by affecting sphingosine-1-phophate and auxin synthesis

Sphingolipids serve as essential components of biomembrane and possess significant bioactive properties. Sphingosine-1-phophate (S1P) plays a key role in plant resistance to stress, but its specific impact on plant growth and development remains to be fully elucidated. Cotton fiber cells are an ideal material for investigating the growth and maturation of plant cells. In this study, we examined the content and composition of sphingosine (Sph) and S1P throughout the progression of fiber cell development. The content of S1P elevated gradually during fiber elongation but declined during the transition stage. Exogenous application of S1P promoted fiber elongation while using of FTY720 (an antagonist of S1P), and DMS (an inhibitor of LCBK) hindered fiber elongation. Cotton Long Chain Base Kinase 1 (GhLCBK1) was notably expressed during the fiber elongation stage, containing all conserved domains of LCBK protein and localized in the endoplasmic reticulum. Overexpression GhLCBK1 increased the S1P content and promoted fiber elongation while retarded secondary cell wall (SCW) deposition. Conversely, downregulation of GhLCBK1 reduced the S1P levels, and suppressed fiber elongation, and accelerated SCW deposition. Transcriptome analysis revealed that upregulating GhLCBK1 or applying S1P induced the expression of GhEXPANSIN and auxin related genes. Furthermore, the levels of IAA were elevated and reduced in the fibers when up-regulating or down-regulating GhLCBK1, respectively. Our investigation demonstrated that GhLCBK1 and its product S1P facilitated the elongation of fiber cells by affecting auxin biosynthesis. This study contributes novel insights into the intricate regulatory pathways involved in fiber cell elongation, identifying GhLCBK1 as a potential target gene and laying the groundwork for enhancing fiber quality via genetic manipulation.

Analysis of genes related to xylem cell wall development based on transcriptomics in Populus alba ‘Berolinensis’ tension wood

Populus alba ‘Berolinensis’ is a fast-growing, high-yielding species with strong biotic and abiotic stress resistance, and widely planted for timber, shelter belts and aesthetic purposes. In this study, molecular development is explored and the important genes regulating xylem formation in P. alba ‘Berolinensis’ under artificial bending treatments was identified. Anatomical investigation indicated that tension wood (TW) was characterized by eccentric growth of xylem and was enriched in cellulose; the degree of lignification was lower than for normal wood (NW) and opposite wood (OW). RNA-Seq-based transcriptome analysis was performed using developing xylem from three wood types (TW, OW and NW). A large number of differentially expressed genes (DEGs) were screened and 4889 counted. In GO and KEGG enrichment results, genes involved in plant hormone signal transduction, phenylpropanoid biosynthesis, and cell wall and secondary cell wall biogenesis play major roles in xylem development under artificial bending. Eight expansin (PalEXP) genes were identified from the RNA-seq data; four were differentially expressed during tension wood formation. Phylogenetic analysis indicated that PalEXLB1 belongs to the EXPB subfamily and that the other PalEXPs are members of the EXPA subfamily. A transcriptional regulatory network construction showed 10 transcription factors located in the first and second layers upstream of EXP, including WRKY, ERF and bHLH. RT‒qPCR analysis in leaves, stems and roots combined with transcriptome analysis suggests that PalEXPA2, PalEXPA4 and PalEXPA15 play significant regulatory roles in cell wall formation during tension wood development. The candidate genes involved in xylem cell wall development during tension wood formation marks an important step toward identifying the molecular regulatory mechanism of xylem development and wood property improvement in P. alba ‘Berolinensis’.

In silico characterization of a MYB TF (WsMYB46) from the medicinal plant Withania somnifera L. predicting its probable role in secondary cell wall biosynthesis

In silico characterization of a MYB TF (WsMYB46) from the medicinal plant Withania somnifera L. predicting its probable role in secondary cell wall biosynthesis

Cell size and xylem differentiation regulating genes from Salicornia europaea contribute to plant salt tolerance.

Cell wall is involved in plant growth and plays pivotal roles in plant adaptation to environmental stresses. Cell wall remodelling may be crucial to salt adaptation in the euhalophyte Salicornia europaea. However, the mechanism underlying this process is still unclear. Here, full-length transcriptome indicated cell wall-related genes were comprehensively regulated under salinity. The morphology and cell wall components in S. europaea shoot were largely modified under salinity. Through the weighted gene co-expression network analysis, SeXTH2 encoding xyloglucan endotransglucosylase/hydrolases, and two SeLACs encoding laccases were focused. Meanwhile, SeEXPB was focused according to expansin activity and the expression profiling. Function analysis in Arabidopsis validated the functions of these genes in enhancing salt tolerance. SeXTH2 and SeEXPB overexpression led to larger cells and leaves with hemicellulose and pectin content alteration. SeLAC1 and SeLAC2 overexpression led to more xylem vessels, increased secondary cell wall thickness and lignin content. Notably, SeXTH2 transgenic rice exhibited enhanced salt tolerance and higher grain yield. Altogether, these genes may function in the succulence and lignification process in S. europaea. This work throws light on the regulatory mechanism of cell wall remodelling in S. europaea under salinity and provides potential strategies for improving crop salt tolerance and yields.

OsGLP8-7 interacts with OsPRX111 to detoxify excess copper in rice

Lignin is a phenolic biopolymer generated from phenylpropanoid pathway in the secondary cell wall and is required for defense of plants against various stress. Although the fact of stress-induced lignin deposition has been clearly demonstrated, it remains largely elusive how the formation of lignin is promoted under Cu stress. The present study showed that OsGLP8-7, an extracellular glycoprotein of rice (Oryza sativa L.), plays an important function against Cu stress. The loss function of OsGLP8-7 results in Cu sensitivity whereas overexpression of OsGLP8-7 scavenges Cu-induced superoxide anion (O2•ˉ). OsGLP8-7 interacts with apoplastic peroxidase111 (OsPRX111) and elevates OsPRX111 stability when exposed to excess Cu. In OsGLP8-7 overexpressing (OE) lines, the retention of Cu within cell wall limiting Cu uptake into cytoplasm is attributed to the enhanced lignification required for Cu tolerance. Exogenous application of a lignin inhibitor can impair the Cu tolerance of transgenic Arabidopsis lines overexpressing OsGLP8-7. In addition, co-expression of OsGLP8-7 and OsPRX111 genes in tobacco leaves leads to an improved lignin deposition compared to leaves expressing each gene individually or the empty vector. Taken together, our findings provided the convincing evidences that the interaction between OsGLP8-7 and OsPRX111 facilitates effectively lignin polymerization, thereby contributing to Cu tolerance in rice.

Cloning and functional mechanism of the dwarf gene gba affecting stem elongation and cellulose biosynthesis in jute (Corchorus olitorius).

Plant height (PH) is an important factor affecting bast fiber yield in jute. Here, we report the mechanism of dwarfism in the 'Guangbaai' (gba) of jute. The mutant gba had shorter internode length and cell length compared to the standard cultivar 'TaiZi 4' (TZ4). Exogenous GA3 treatment indicated that gba is a GA-insensitive dwarf mutant. Quantitative trait locus (QTL) analysis of three PH-related traits via a high-density genetic linkage map according to re-seq showed that a total of 25 QTLs were identified, including 13 QTLs for PH, with phenotypic variation explained ranging from 2.42 to 74.16%. Notably, the functional mechanism of the candidate gene CoGID1a, the gibberellic acid receptor, of the major locus qPHIL5 was evaluated by transgenic analysis and virus-induced gene silencing. A dwarf phenotype-related single nucleotide mutation in CoGID1a was identified in gba, which was also unique to the dwarf phenotype of gba among 57 cultivars. Cogid1a was unable to interact with the growth-repressor DELLA even in the presence of highly accumulated gibberellins in gba. Differentially expressed genes between transcriptomes of gba and TZ4 after GA3 treatment indicated up-regulation of genes involved in gibberellin and cellulose synthesis in gba. Interestingly, it was found that up-regulation of CoMYB46, a key transcription factor in the secondary cell wall, by the highly accumulated gibberellins in gba promoted the expression of cellulose synthase genes CoCesA4 and CoCesA7. These findings provide valuable insights into fiber development affected by endogenous gibberellin accumulation in plants.

Alfalfa Responses to Intensive Soil Compaction: Effects on Plant and Root Growth, Phytohormones and Internal Gene Expression.

The perennial legume alfalfa (Medicago sativa L.) is of high value in providing cheap and high-nutritive forages. Due to a lack of tillage during the production period, the soil in which alfalfa grows prunes to become compacted through highly mechanized agriculture. Compaction deteriorates the soil's structure and fertility, leading to compromised alfalfa development and productivity. However, the way alfalfa responses to different levels of soil compaction and the underlying molecular mechanism are still unclear. In this study, we systematically evaluated the effects of gradient compacted soil on the growth of different cultivars of alfalfa, especially the root system architecture, phytohormones and internal gene expression profile alterations. The results showed that alfalfa growth was facilitated by moderate soil compaction, but drastically inhibited when compaction was intensified. The inhibition effect was universal across different cultivars, but with different severity. Transcriptomic and physiological studies revealed that the expression of a set of genes regulating the biosynthesis of lignin and flavonoids was significantly repressed in compaction treated alfalfa roots, and this might have resulted in a modified secondary cell wall and xylem vessel formation. Phytohormones, like ABA, are supposed to play pivotal roles in the regulation of the overall responses. These findings provide directions for the improvement of field soil management in alfalfa production and the molecular breeding of alfalfa germplasm with better soil compaction resilience.

Reducing nitrogen application at high planting density enhances secondary cell wall formation and decreases stem lodging in rapeseed

Reducing the rate of nitrogen fertilizer (N rate) and achieving high plant density is an effective and economically beneficial approach to enhance rapeseed productivity. However, it is unclear how N rate affects stem lodging in rapeseed directly-sown at high densities. In this study, the effect of different N rates on stem lodging at high plant density was examined in field trial and pot experiments using two modern hybrid rapeseed varieties, Fengyou520 (FY520) and Huayouza62 (HYZ62). Increasing N fertilizer from low level to adequate level significantly increased yield by 39% in FY520 and 43% in HYZ62. However, with elevated N yield increases were limited by an increase in stem lodging. At T1 (the first inflorescence branch below the apex has the first flowers open) phloroglucinol-HCl staining for lignin was weaker under high N (N4: 4 g N pot−1) compared with adequate N (N2: 2 g N pot−1), and was correlated with weaker stem strength. Differences in stem lignin were also associated with variations in gene expression as revealed by transcriptomic profiling of middle stems at T1. Differentially expressed genes indicated significant enrichment for cell wall. Several genes encoding known transcriptional regulators of secondary cell walls (SCWs) biosynthesis and deposition were down-regulated with high N compared with adequate N. Many genes putatively involved in lignin biosynthesis, cellulose biosynthesis and assembly and xylan biosynthesis are down-regulated with high N, corresponding to lower accumulation of lignin, cellulose and hemicellulose in these plants. These findings not only provide us with gene candidates potentially affecting stem lodging in direct sown rapeseed, but also reveal that reducing N at high plant density enhances biosynthesis of stem cell walls to alleviate stem lodging.

Cortical microtubule dynamics during reaction wood formation ensures context-appropriate cellulose microfibril angle in woody trees

Key messageCortical microtubule arrays are the primary mechanism for guiding the re-orientation of cellulose microfibrils and determining MFA in secondary cell wall of wood fibre and tracheid cells in reaction wood.Microtubules are directly and indirectly involved in guiding cellulose synthase complexes (CSCs) through the plasma membrane. The angle of cellulose deposition is a critical response to environmental signals and/or stress conditions, and particularly crucial during reaction wood formation, a process in which woody plants deposit additional cell wall material to counteract gravitational forces. Tubulin genes are upregulated in response to gravitational stimulus during reaction wood formation, which can result in changes to microtubule assembly. In this study, microtubules were visualised in three woody tree species (two angiosperms: Eucalyptus globulus Labill., Populus alba L., and one gymnosperm: Pinus radiata D. Don.) using immunofluorescence to quantitatively evaluate microtubule organisation during reaction wood formation. Our results suggest that reorientation of the cortical microtubule array affects secondary cell wall deposition, even across different types of reaction wood, by ensuring context-appropriate orientation of cellulose microfibrils and determining MFA in wood cells. Pharmacological studies conducted on in vitro cultured stem segments or in vivo during reaction wood formation corroborated these important roles for microtubules during wood development. This study starts to unveil the role of tubulins during wood formation by exploring cortical microtubule array organisation in trees subjected to gravitational stimulus and it sheds light on cellular and molecular mechanisms behind cellulose deposition in tree species.

Imaging plant cell walls using fluorescent stains: The beauty is in the details.

Plants continuously face various environmental stressors throughout their lifetime. To be able to grow and adapt in different environments, they developed specialized tissues that allowed them to maintain a protected yet interconnected body. These tissues undergo specific primary and secondary cell wall modifications that are essential to ensure normal plant growth, adaptation and successful land colonization. The composition of cell walls can vary among different plant species, organs and tissues. The ability to remodel their cell walls is fundamental for plants to be able to cope with multiple biotic and abiotic stressors. A better understanding of the changes taking place in plant cell walls may help identify and develop new strategies as well as tools to enhance plants' survival under environmental stresses or prevent pathogen attack. Since the invention of microscopy, numerous imaging techniques have been developed to determine the composition and dynamics of plant cell walls during normal growth and in response to environmental stimuli. In this review, we discuss the main advances in imaging plant cell walls, with a particular focus on fluorescent stains for different cell wall components and their compatibility with tissue clearing techniques. Lay Description: Plants are continuously subjected to various environmental stresses during their lifespan. They evolved specialized tissues that thrive in different environments, enabling them to maintain a protected yet interconnected body. Such tissues undergo distinct primary and secondary cell wall alterations essential to normal plant growth, their adaptability and successful land colonization. Cell wall composition may differ among various plant species, organs and even tissues. To deal with various biotic and abiotic stresses, plants must have the capacity to remodel their cell walls. Gaining insight into changes that take place in plant cell walls will help identify and create novel tools and strategies to improve plants' ability to withstand environmental challenges. Multiple imaging techniques have been developed since the introduction of microscopy to analyse the composition and dynamics of plant cell walls during growth and in response to environmental changes. Advancements in plant tissue cleaning procedures and their compatibility with cell wall stains have significantly enhanced our ability to perform high-resolution cell wall imaging. At the same time, several factors influence the effectiveness of cleaning and staining plant specimens, as well as the time necessary for the process, including the specimen's size, thickness, tissue complexity and the presence of autofluorescence. In this review, we will discuss the major advances in imaging plant cell walls, with a particular emphasis on fluorescent stains for diverse cell wall components and their compatibility with tissue clearing techniques. We hope that this review will assist readers in selecting the most appropriate stain or combination of stains to highlight specific cell wall components of interest.