Increased Lignin Deposition Research Articles

Functional characterization of WsPR-1 reveals its interplay with cytokinin and gibberellin signaling pathways

Pathogenesis-related protein 1 (PR-1) is an antimicrobial protein involved in systemic acquired resistance (SAR) in plants, but its regulatory role and interactions with other pathways remain unclear. In this study, we functionally characterize WsPR-1 gene of Withania somnifera in Nicotiana tabacum to elucidate its role in plant defense, growth, and development. Interestingly, transgenic tobacco plants with increased levels of cytokinin (CK) and decreased gibberellins (GAs) exhibited stunted shoot growth, an underdeveloped root system, modified leaf morphology, reduced seed pod production, and delayed leaf senescence. Transcriptional analysis revealed that WsPR-1 overexpression downregulated the GA 20-oxidase (GA20ox) gene involved in GA biosynthesis while upregulating GA 2-oxidase (GA2ox), a GA catabolic enzyme. Moreover, transcript levels of FRUITFULL (FUL) and LEAFY (NFL2) flowering genes exhibited a decrease in WsPR-1 plants, which could explain the delayed flowering and reduced seed pod development in transgenic plants. Confocal microscopy confirmed increased lignin deposition in stem cross-sections of WsPR-1 transgenic plants, supported by gene expression analysis and lignin content quantification. Additionally, our findings also suggest the involvement of Knotted1-like homeobox (KNOX) gene in enhancing cytokinin levels. This study highlights PR-1's regulatory role in plant growth and development, with potential to boost crop yields and enhance resilience.

Gibberellin signaling regulates lignin biosynthesis to modulate rice seed shattering.

The elimination of seed shattering was a key step in rice (Oryza sativa) domestication. In this paper, we show that increasing the gibberellic acid (GA) content or response in the abscission region enhanced seed shattering in rice. We demonstrate that SLENDER RICE1 (SLR1), the key repressor of GA signaling, could physically interact with the rice seed shattering-related transcription factors quantitative trait locus of seed shattering on chromosome 1 (qSH1), O. sativa HOMEOBOX 15 (OSH15), and SUPERNUMERARY BRACT (SNB). Importantly, these physical interactions interfered with the direct binding of these three regulators to the lignin biosynthesis gene 4-COUMARATE: COENZYME A LIGASE 3 (4CL3), thereby derepressing its expression. Derepression of 4CL3 led to increased lignin deposition in the abscission region, causing reduced rice seed shattering. Importantly, we also show that modulating GA content could alter the degree of seed shattering to increase harvest efficiency. Our results reveal that the "Green Revolution" phytohormone GA is important for regulating rice seed shattering, and we provide an applicable breeding strategy for high-efficiency rice harvesting.

A Higher Lignin Content in ugt72b37 Poplar Mutants Indicates a Role of Monolignol Glycosylation in Xylem Lignification

Plant UDP-glycosyltransferases (UGT) transfer sugars to small acceptor molecules and thereby play key roles in the biosynthesis of secondary metabolites, including phenylpropanoids. Some of those metabolites are involved in the xylem lignification of a broad range of terrestrial plants, particularly trees. Here, we focused on poplar UGT72B37, coding for an enzyme glycosylating monolignols by investigating CRISPR/Cas9 mutant lines. The cell wall characterization revealed a 10% lignin content increase in the xylem of three-month-old mutant lines compared to the wild type. No ectopic lignification was evidenced in the pith of the stems of the mutants, suggesting that the increased lignin deposition is restricted to lignified cell walls. The analysis of the expression level of lignin biosynthesis and polymerization genes did not show significant changes between the WT and the ugt72b37 mutants, except for CINNAMOYL-COA REDUCTASE 2 which was significantly upregulated by 1.2–1.5-fold. Noticeably, UGT72B38, the closest related gene to UGT72B37, is upregulated in mutant lines, suggesting a functional compensation between UGT72B37 and UGT72B38 possibly linked with lignin biosynthesis and accumulation in poplar. Overall, these results reinforce a plausible role of monolignol glycosylation in the cell wall lignification process.

Endophytic Fusarium clavum confers growth and salt tolerance in Cucumis melo

In the present study, we investigated the in-vitro and in-vivo effects of root endophytic fungal species isolated from squirting cucumber Ecballium elaterium on morphological, physiological and biochemical mechanisms of muskmelon (Cucumis melo) in response to salt stress. A total of 23 endophytic fungal isolates belonging to nine different genera were initially screened for their salt tolerance potential in saline medium. Endophytic Fusarium clavum EeR24 exhibited maximum salt tolerance in screening studies and was selected for further studies. Seedlings of C. melo were inoculated with F. clavum EeR24 and one week later were subjected to two salt (100 and 200 mM NaCl) and control (no salt) treatments in a growth chamber and greenhouse experiments. In growth chamber assays, seedlings treated with F. clavum EeR24 exhibited significant increase in growth and biomass under both controlled and saline conditions. Under greenhouse conditions, F. clavum EeR24 significantly improved morphological and physiological traits including plant height, weight, number of leaves, membrane stability, electrical conductivity, photosynthesis, stomatal conductance and transpiration, in C. melo in response to salt stress. Endophyte colonization also led to decreased absorption of Na+ (36–42 %) and Cl- (23–33 %) ions and an improved K+ (35 %) absorption. Under saline conditions, endophyte colonization improved catalase (24–37 %), guaiacol peroxidase (31–42 %) and superoxide dismutase (16–22 %), chlorophyll and phenolic content by 30–64 %, compared to non-colonized plants. Lipid peroxidation activities significantly decreased by 45 % and proline accumulation increased by 18 % in endophyte-colonized compared to non-colonized plants. Histochemical analysis indicated that under saline conditions, endophytic colonization reduced levels of H2O2, superoxide ion production and cell mortality and increased lignin deposition. The endophytic isolate F. clavum EeR24 also exhibited plant growth promoting activities by producing hydrolytic enzymes, IAA, and phosphate solubilization. Furthermore, a study on spatial colonization of the endophyte, using either conventional Petri-plate counts or RFP-tagging, indicated that F. clavum was able to penetrate the root tissues of C. melo. Conclusively, the endophyte F. clavum EeR24 is an effective salt stress reducer and melon growth promotor; hence can be further taken into account for development as an eco-friendly bio-fertilizer to confer salt stress tolerance in melon.

SCFOsFBK1 E3 ligase mediates jasmonic acid-induced turnover of OsATL53 and OsCCR14 to regulate lignification of rice anthers and roots.

The rice F-box protein OsFBK1, which mediates the turnover of a cinnamoyl CoA-reductase, OsCCR14, has previously been shown to regulate anther and root lignification. Here, we identify OsATL53, a member of the ATL family of RING-H2 proteins that interacts with OsCCR14 in the cytoplasm. OsATL53 was identified in the same yeast two-hybrid library screening as reported previously for OsCCR14, and we show it to have cytoplasmic localization and E3 ligase ubiquitination properties. SCFOsFBK1 mediates turnover of OsATL53 in the cytoplasm and the nucleus, and that of OsCCR14 only in the nucleus, as shown by cell-free degradation assays. Confocal fluorescence lifetime imaging microscopy analyses demonstrate that in presence of jasmonic acid (JA), which plays a role in anther dehiscence, OsATL53-OsCCR14 undergoes conformational changes that trigger the complex to accumulate around the nuclear periphery and signals OsFBK1 to initiate degradation of the proteins in the respective cellular compartments. OsATL53 decreases the enzymatic activity of OsCCR14 and sequesters it in the cytoplasm, thereby regulating the lignification process. Transgenic rice with knockdown of OsATL53 display increased lignin deposition in the anthers and roots compared to the wild type, whilst knockdown of OsCCR14 results in decreased lignin content. Our results show that OsATL53 affects the activity of OsCCR14, and that their JA-induced degradation by SCFOsFBK1 regulates lignification of rice anthers and roots.

MicroRNA828 negatively regulates lignin biosynthesis in stem of Populus tomentosa through MYB targets.

Lignin biosynthesis in the sclerenchyma cells is strictly controlled by a complex network of genetic and environmental signals. In the last decades, the transcriptional regulation of lignin synthesis in woody species has been established. However, the role of microRNA-mediated post-transcriptional modulation in secondary cell wall biosynthesis remains poorly understood. Here, we identified a microRNA, miR828, involved in the regulation specific to lignin biosynthesis during stem development in Populus tomentosa Carr. miR828 is preferentially expressed in the secondary vascular tissues during stem development. Two MYB genes (MYB171 and MYB011) were validated as direct targets of miR828 by degradome analysis and green fluorescent protein signal detection. Overexpression of miR828 in poplar downregulated genes for lignin biosynthesis, resulting in reduced lignin content in cell walls. Conversely, suppression of miR828 in plants by the short tandem target mimics elevated the expression of lignin biosynthetic genes and increased lignin deposition. We further revealed that poplar MYB171, as the most abundant miR828 target in the stem, is a positive regulator for lignin biosynthesis. Transient expression assays showed that both MYB171 and MYB011 activated PAL1 and CCR2 transcription, whereas the introduction of miR828 significantly suppressed their expression that was induced by MYB171 or MYB011. Collectively, our results demonstrate that the miR828-MYBs module precisely regulates lignin biosynthesis during the stem development in P. tomentosa through transcriptional and post-transcriptional manners.

Full-length transcriptomic identification of R2R3-MYB family genes related to secondary cell wall development in Cunninghamia lanceolata (Chinese fir)

BackgroundR2R3-MYB is a class of transcription factor crucial in regulating secondary cell wall development during wood formation. The regulation of wood formation in gymnosperm has been understudied due to its large genome size. Using Single-Molecule Real-Time sequencing, we obtained full-length transcriptomic libraries from the developmental stem of Cunninghamia lanceolata, a perennial conifer known as Chinese fir. The R2R3-MYB of C. lanceolata (hereafter named as ClMYB) associated with secondary wall development were identified based on phylogenetic analysis, expression studies and functional study on transgenic line.ResultsThe evolutionary relationship of 52 ClMYBs with those from Arabidopsis thaliana, Eucalyptus grandis, Populus trichocarpa, Oryza sativa, two gymnosperm species, Pinus taeda, and Picea glauca were established by neighbour-joining phylogenetic analysis. A large number of ClMYBs resided in the woody-expanded subgroups that predominated with the members from woody dicots. In contrast, the woody-preferential subgroup strictly carrying the members of woody dicots contained only one candidate. The results suggest that the woody-expanded subgroup emerges before the gymnosperm/angiosperm split, while most of the woody-preferential subgroups are likely lineage-specific to woody dicots. Nine candidates shared the same subgroups with the A. thaliana orthologs, with known function in regulating secondary wall development. Gene expression analysis inferred that ClMYB1/2/3/4/5/26/27/49/51 might participate in secondary wall development, among which ClMYB1/2/5/26/27/49 were significantly upregulated in the highly lignified compression wood region, reinforcing their regulatory role associated with secondary wall development. ClMYB1 was experimentally proven a transcriptional activator that localised in the nucleus. The overexpression of ClMYB1 in Nicotiana benthamiana resulted in an increased lignin deposition in the stems. The members of subgroup S4, ClMYB3/4/5 shared the ERF-associated amphiphilic repression motif with AtMYB4, which is known to repress the metabolism of phenylpropanoid derived compounds. They also carried a core motif specific to gymnosperm lineage, suggesting divergence of the regulatory process compared to the angiosperms.ConclusionsThis work will enrich the collection of full-length gymnosperm-specific R2R3-MYBs related to stem development and contribute to understanding their evolutionary relationship with angiosperm species.

PmMYB4, a Transcriptional Activator from Pinus massoniana, Regulates Secondary Cell Wall Formation and Lignin Biosynthesis

Wood formation originates in the biosynthesis of lignin and further leads to secondary cell wall (SCW) biosynthesis in woody plants. Masson pine (Pinus massoniana Lamb) is an economically important industrial timber tree, and its wood yield affects the stable development of the paper industry. However, the regulatory mechanisms of SCW formation in Masson pine are still unclear. In this study, we characterized PmMYB4, which is a Pinus massoniana MYB gene involved in SCW biosynthesis. The open reading frame (ORF) of PmMYB4 was 1473 bp, which encoded a 490 aa protein and contained two distinctive R2 and R3 MYB domains. It was shown to be a transcription factor, with the highest expression in semi-lignified stems. We overexpressed PmMYB4 in tobacco. The results indicated that PmMYB4 overexpression increased lignin deposition, SCW thickness, and the expression of genes involved in SCW formation. Further analysis indicated that PmMYB4 bound to AC-box motifs and might directly activate the promoters of genes (PmPAL and PmCCoAOMT) involved in SCW biosynthesis. In addition, PmMYB4-OE(over expression) transgenic lines had higher lignin and cellulose contents and gene expression than control plants, indicating that PmMYB4 regulates SCW mainly by targeting lignin biosynthetic genes. In summary, this study illustrated the MYB-induced SCW mechanism in Masson pine and will facilitate enhanced lignin and cellulose synthesis in genetically engineered trees.

Lignin Composition and Timing of Cell Wall Lignification Are Involved in Brassica napus Resistance to Stem Rot Caused by Sclerotinia sclerotiorum

Sclerotinia stem rot (SSR) is an economically and globally significant disease in oilseed rape (Brassica napus) caused by the necrotrophic ascomycete Sclerotinia sclerotiorum. This study explored the role of cell wall reinforcement by lignin as a relevant factor for effective plant defense against attack by this pathogen. Expression of key genes in the phenylpropanoid pathway and the induced synthesis of lignin in infected stem tissues were investigated in a study comparing a susceptible ('Loras') and a moderately resistant cultivar ('Zhongyou 821' [ZY821]). Data revealed an earlier and more rapid defense activation in ZY821 through upregulation of transcript levels of genes related to key steps in the phenylpropanoid pathway associated with increased lignin deposition in the resistant B. napus genotype. Expression level of BnCAD5, encoding a cinnamyl alcohol dehydrogenase, responsible for conversion of monolignol to lignin, was more rapidly upregulated in ZY821 than 'Loras'. The similar expression pattern of BnCAD5 and the gene BnF5H, encoding for ferulate-5-hydroxylase, which catalyzes the synthesis of syringyl (S) lignin precursors, suggests that BnCAD5 is involved in S lignin formation. Histological observations confirmed these results, showing an earlier increase of S lignin deposition in the infected resistant genotype. Deposition of guaiacyl lignin was detected in both genotypes and is thus considered a component of basal, cultivar-independent defense response of B. napus to stem rot. The results indicate the importance of cell wall modification for quantitative stem rot resistance by responses in the phenylpropanoid metabolism generating distinct lignin types on different temporal scales.

Syringic Acid Alleviates Cesium-Induced Growth Defect in Arabidopsis.

Syringic acid, a phenolic compound, serves a variety of beneficial functions in cells. Syringic acid increases in plants in response to cesium, and exogenous application of syringic acid resulted in a significant attenuation of cesium-induced growth defects in Arabidopsis. In addition, cesium or syringic acid application to plants also resulted in increased lignin deposition in interfascicular fibers. To better understand the role of lignin and syringic acid in attenuating cesium-induced growth defects, two mutants for Arabidopsis REDUCED EPIDERMAL FLUORESCENE 4 (REF4) and fourteen laccase mutants, some of which have lower levels of lignin, were evaluated for their response to cesium. These mutants responded differently to cesium stress, compared to control plants, and the application of syringic acid alleviated cesium-induced growth defects in the laccase mutants but not in the ref4 mutants. These findings imply that lignin plays a role in cesium signaling but the attenuation of cesium stress defects by syringic acid is mediated by regulatory components of lignin biosynthesis and not lignin biosynthesis itself. In contrast, syringic acid did not alleviate any low potassium-induced growth defects. Collectively, our findings provide the first established link between lignin and cesium stress via syringic acid in plants.

Isolation and functional identification of a Botrytis cinerea-responsive caffeoyl-CoA O-methyltransferase gene from Lilium regale wilson

In plants, genes involved in the Phenylpropanoid/monolignol pathway play important roles in lignin biosynthesis and plant immunity. However, their biological function in Lilium remains poorly characterized. Comparative RNA sequencing of the expression profiles of the monolignol pathway genes from fungi-resistant species Lilium regale after inoculation with Botrytis cinerea was performed. One upregulated caffeoyl-CoA O-methyltransferase gene, LrCCoAOMT, was cloned for functional characterization by reverse genetic methods. LrCCoAOMT encodes a putative protein of 246 amino acids and is highly expressed in stem tissues and responsive to salicylic acid (SA) signaling and B. cinerea infection. LrCCoAOMT was largely directed to the cytoplasm. LrCCoAOMT overexpression in Arabidopsis resulted in an increased lignin deposition in vascular tissues and conferred resistance to B. cinerea infection in transgenic plants. Transient transformation of LrCCoAOMT in nonresistant Lilium sargentiae leaves also identified the defense function to B. cinerea. In addition, transcript levels of genes involved in the monolignol and SA-dependent signaling pathways were altered in transgenic Arabidopsis, suggesting that LrCCoAOMT might play vital roles in the resistance of L. regale to B. cinerea related to the levels of lignin and the regulation of SA signaling. This is the first report to functionally characterize a CCoAOMT gene in Lilium, a potential molecular target for lily molecular improvement.

PbMC1a/1b regulates lignification during stone cell development in pear (Pyrus bretschneideri) fruit

Programmed cell death (PCD) and secondary cell wall (SCW) thickening in pear fruit are accompanied by the deposition of cellulose and lignin to form stone cells. Metacaspase is an important protease for development, tissue renewal and PCD. The understanding of the molecular mechanism whereby pear (Pyrus) metacaspase promotes PCD and cell wall lignification is still limited. In this study, the Metacaspases gene family (PbMCs) from P. bretschneideri was identified. PbMC1a/1b was associated with lignin deposition and stone cell formation by physiological data, semiquantitative real-time polymerase chain reaction (RT-PCR) and quantitative RT-PCR (qRT-PCR). Relative to wild-type (WT) Arabidopsis, the overexpression of PbMC1a/1b increased lignin deposition and delayed growth, thickened the cell walls of vessels, xylary fibers and interfascicular fibers, and increased the expression of lignin biosynthetic genes. Yeast two-hybrid (Y2H), bimolecular fluorescence complementation (BiFC) and GST pull-down assays indicated that the PbMC1a/1b protein physically interacted with PbRD21. Simultaneously, the transient expression of PbMC1a/1b and PbRD21 led to significant changes in the expression of genes and lignin contents in pear fruits and flesh calli. These results indicate that PbMC1a/1b plays an important role in cell wall lignification, possibly by interacting with PbRD21 to increase the mRNA levels of some lignin synthesis-associated genes and promote the formation of stone cells in pear fruit.

Gibberellin Promotes Sweetpotato Root Vascular Lignification and Reduces Storage-Root Formation.

Sweetpotato yield depends on a change in the developmental fate of adventitious roots into storage-roots. The mechanisms underlying this developmental switch are still unclear. We examined the hypothesis claiming that regulation of root lignification determines storage-root formation. We show that application of the plant hormone gibberellin increased stem elongation and root gibberellin levels, while having inhibitory effects on root system parameters, decreasing lateral root number and length, and significantly reducing storage-root number and diameter. Furthermore, gibberellin enhanced root xylem development, caused increased lignin deposition, and, at the same time, decreased root starch accumulation. In accordance with these developmental effects, gibberellin application upregulated expression levels of sweetpotato orthologues of Arabidopsis vascular development regulators (IbNA075, IbVND7, and IbSND2) and of lignin biosynthesis genes (IbPAL, IbC4H, Ib4CL, IbCCoAOMT, and IbCAD), while downregulating starch biosynthesis genes (IbAGPase and IbGBSS) in the roots. Interestingly, gibberellin downregulated root expression levels of orthologues of the Arabidopsis BREVIPEDICELLUS transcription factor (IbKN2 and IbKN3), regulator of meristem maintenance. The results substantiate our hypothesis and mark gibberellin as an important player in regulation of sweetpotato root development, suggesting that increased fiber formation and lignification inhibit storage-root formation and yield. Taken together, our findings provide insight into the mechanisms underlying sweetpotato storage-root formation and provide a valuable database of genes for further research.

Mechanismal analysis of resistance to Verticillium dahliae in upland cotton conferred by overexpression of RPL18A-6 (Ribosomal Protein L18A-6)

Verticillium wilt of cotton, caused by the fungal phytopathogen Verticillium dahliae (V. dahliae), is one of the most devastating diseases of upland cotton. While Verticillium dahliae-resistant phenotypes have been identified in several plant species, the underlying mechanisms of resistance need further clarification. Here, the induction of resistance to V. dahliae through over-expression of the GhARPL18A-6 (Ribosomal protein L18A) in Gossypium hirsutum (G. hirsutum) was demonstrated. GhARPL18A-6 was highly induced in G. hirsutum acc. Zhongzhimian2 (Zz2) after inoculation with Verticillium dahliae. Knock-down of GhARPL18A-6 in Zz2 led to disease-sensitive phenotypes to V. dahliae. Further studies showed that increased lignin deposition, an enhanced reactive oxygen species (ROS) burst, and phenylpropanoid biosynthesis defense response pathways all contributed to a decrease in colonization by V. dahliae. RNA-seq analysis of GhARPL18A-6 over-expression and wild-type cotton revealed differentiate-expression of cell wall biosynthesis, sucrose signaling and metabolism, lignin biosynthesis, and ROS response genes in resistant plants inoculated with V. dahliae. Given the susceptibility of most commercially valuable cotton lines, this research provides insight into the mechanisms of resistance to Verticillium wilt as well as novel germplasm for cotton breeding.

PbrMYB169 positively regulates lignification of stone cells in pear fruit.

Stone cells negatively affect fruit quality because of their firm and lignified cell walls, so are targets for reduction in pear breeding programmes. However, there is only limited knowledge of the molecular mechanisms underlying the formation of stone cells. Here, we show that PbrMYB169, an R2R3 MYB transcription factor, of Pyrus bretschneideri positively regulates lignification of stone cells in pear fruit. PbrMYB169 was shown to be co-expressed with lignin biosynthesis genes during pear fruit development, and this co-expression pattern was coincident with stone cell formation in the fruit of Pyrus bretschneideri 'Dangshansuli'. The PbrMYB169 expression level was also positively correlated with stone cell content in 36 pear cultivars tested. PbrMYB169 protein significantly activated the promoter of lignin genes C3H1, CCR1, CCOMT2, CAD, 4CL1, 4CL2, HCT2, and LAC18 via binding to AC elements [ACC(T/A)ACC] in these promoters. Furthermore, overexpression of PbrMYB169 in transgenic Arabidopsis plants enhanced the expression of lignin genes, and increased lignin deposition and cell wall thickness of vessel elements, but did not change the ratio of syringyl and guaiacyl lignin monomers. In conclusion, PbrMYB169 appears to be a transcriptional activator of lignin biosynthesis and regulates secondary wall formation in fruit stone cells. This study advances the understanding of the regulation of lignin biosynthesis and provides valuable molecular genetic information for reducing stone cell content in pear fruit.

The basis of pod dehiscence: anatomical traits of the dehiscence zone and expression of eight pod shatter-related genes in four species of Brassicaceae

Members of the Brassicaceae family disperse their seeds through a mechanism commonly referred to as fruit dehiscence or pod shatter. Pod shatter is influenced by variations in valve margin structure and by the molecular control pathways related to valve development. Anatomical patterns of the dehiscence zone from Brassica napus L., Brassica rapa L., Brassica carinata L., and Sinapis alba L., representing fruit types differing in pod shatter resistance, were compared using histological staining. The pod shatter-susceptible plant B. napus showed an increased lignin deposition at the vascular bundle of the replum as well as an increased separation of cell layers. In pod shatter-resistant plants S. alba, B. rapa, and B. carinata, we observed two layers of lignified valve margin cells. From these four species, we isolated and identified homologs of SHATTERPROOF (SHP1, SHP2), INDEHISCENT (IND), ALCATRAZ (ALC), FRUITFULL (FUL), AGAMOUS (AG), NAC SECONDARY WALL THICKENING PROMOTING FACTOR1 (NST1), and SEEDSTICK (STK) genes involved in fruit development and pod shatter in Arabidopsis. Transcriptional analysis of these eight genes was performed by real-time polymerase chain reaction and the results demonstrate that differences in the expression patterns of the eight genes may be associated with dehiscence variation within these four species.

INDUCTION OF RESISTANCE AGAINST VERTICILLIUM DAHLIAE IN CACAO BY A CRINIPELLIS PERNICIOSA SUSPENSION

SUMMARY This study deals with the potential of reducing Verticillium wilt in susceptible cacao using a fungal suspension compared to a commercial resistance inducer. Plants were sprayed with acibenzolar-S-methyl (ASM; Bion® 50 WG (0.2 g l -1 ) and a heterogeneous chitosan suspension (MCp) from Crinipellis perniciosa mycelium. Plants were challenged five days later with a virulent isolate of Verticillium dahliae, under greenhouse conditions. A reduction of Verticillium wilt severity was observed in plants treated by the substances tested, with MCp reaching 80% of ASM protection performance. To evaluate local and systemic activation of defences, cacao leaves and hypocotyls exposed to ASM and MCp were assayed for peroxidase, polyphenol oxidase and lignin deposition. Local induction of resistance was confirmed by the increase of chitinase and b-1,3-glucanase activities in the leaves, 4-18 days after spraying. Treated and inoculated plants showed, in the leaves and hypocotyls, increased lignin deposition, which is associated with the cacao defence strategy against Verticillium wilt.

Reinforcement of cell wall in roots of Lycopersicon esculentum through induction of phenolic compounds and lignin by elicitors

Changes in phenolic metabolism and lignin deposition have been studied in roots of tomato plants after elicitation with four elicitors which are Fusarium mycelium extract (FME), chitosan (CHT), Fusarium culture filtrate (FCF) and Trichoderma mycelium extract (TME). Most profound effect of elicitors was observed on ferulic acid among the phenolic compounds. After 24 h elicitation, the increase in ferulic acid content of root cell wall was 3.71 and 3.30 times by FME and CHT, respectively. The increase of 4-hydroxybenzoic acid was 2.71 and 2.16 times by these two elicitors. The level of 4-coumaric acid was little more than double by these two elicitors after 24 h elicitation. Most pronounced increase in lignin synthesis was also effected by FME followed by CHT. Lignin deposition in the root cell wall was increased 3.6, 5.4 and 7.1 times by FME during 12, 24 and 36 h after elicitation, respectively. Similarly, CHT increased lignin deposition by 2.8, 5.1 and 6.8 times at 12, 24 and 36 h after elicitation, respectively. FCF and TME also increased lignin deposition significantly in the cell walls of tomato roots during the above time periods of elicitation. Activity of phenylalanine ammonia lyase reached highest level at 24 h post elicitation under the influence of the elicitors. Peroxidase activity registered a sharp increase at 24 h post elicitation. Markedly increased level of polyphenol oxidase activity was found at 12 h post elicitation. Cinnamyl alcohol dehydrogenase activity was observed to reach highest level at 48 h post elicitation. Cell wall strengthening, through the deposition of lignin, preceded by the induction of the synthesizing enzymes appears to play an important role in the defense response of Lycopersicon esculentum in reaction to elicitors, including one derived from Fusarium oxysporum f. sp. lycopersici, the causal organism of Fusarium wilt of tomato.

The knotted1-like homeobox gene BREVIPEDICELLUS regulates cell differentiation by modulating metabolic pathways.

Members of the KNOX gene family have important roles in plant meristems by regulating cell division and differentiation. BREVIPEDICELLUS (BP), one of seven KNOX genes in Arabidopsis, has a primary role in internode patterning. We carried out a comparison of RNA expression profiles between wild-type seedlings and bp mutants at a developmental stage prior to a visible phenotypic difference. Transcript differences were found for a number of genes in cell wall biosynthesis, especially genes in the lignin pathway. The regulation of lignin biosynthesis by BP was demonstrated by observing increased lignin deposition in bp mutants following bolting, decreased lignification in plants overexpressing BP, and aberrant lignin deposition in discrete regions of the bp stem. Furthermore, we showed that BP binds promoters of some genes in the lignin pathway. Our results provide a metabolic fingerprint for BP and identify the lignin pathway as one of the coordinate processes that BP regulates.

Induction of defence responses in cultured tobacco cells by elicitors from Phytophthora nicotanae

Black Sbank disease of tobacco, Nicodanae tabacum, caused by Pkytopathora nicotianae, is one of the most eve of soil borne diseases. The present investiption was initiated to study the relative importance of various inducible mechanisms operative in this plant:pathogen interaction. Tobacco cell is culture were treated with beat-released soluble cell-wall elicitors from myceiial walls of the pathogen. The timing and intensity of the induced defence responses of the cells with regard to the synthesis of pbytoalexins, lignin and defence hydrolases were evaluated. An elicitor concentration of 40 μg ml −1 resulted in the optimal induction of sesguiterpenoid pbytoslexin synthesis which was detectable 3–4 hr after elicitation and reached a maximum at 20–28 hr post-elicitation. Increased lignin deposition was detectable 4 hr after elicitation with maximal synthesis between 12 and % hr. Induction of extraceffrdar and cellular β-1,3-glucauases was observed within 4 hr, exhibiting a biphasic response with an initial peak at 8–12 h. Increased cellular and echltimse activities were detected from 8 and 16 hr onwards, respect ively. These results were supported by enzyme staining of electropboretk separated isoforms. The rests obtained indicate that the elicitation of tobacco cells by P. nicotianae derived sigioal molecules exhibits the properties of a multicomponent dynamic system with different protective mechanisms wing complementary roles in the overall expression of the defence response.