Exploring the potential of engineering polygalacturonase-inhibiting protein as an ecological, friendly, and nontoxic pest control agent.

In plants, polygalacturonase-inhibiting proteins (PGIPs) play critical roles for resistance to fungal disease by inhibiting the pectin-depolymerizing activity of endopolygalacturonases (PGs), one type of enzyme secreted by pathogens that compromises plant cell walls and leaves the plant susceptible to disease. Here, the interactions between PGIPs from Phaseolus vulgaris (PvPGIP1 and PvPGIP2) and PGs from Aspergillus niger (AnPG2), Botrytis cinerea (BcPG1 and BcPG2), and Fusarium moniliforme (FmPG3) were reconstituted through a yeast two hybrid (Y2H) system to investigate the inhibition efficiency of various PvPGIP1 and 2 truncations and mutants. We found that tPvPGIP2_5-8, which contains LRR5 to LRR8 and is only one-third the size of the full length peptide, exhibits the same level of interactions with AnPG and BcPGs as the full length PvPGIP2 via Y2H. The inhibitory activities of tPvPGIP2_5-8 on the growth of A. niger and B. cinerea were then examined and confirmed on pectin agar. On pectin assays, application of both full length PvPGIP2 and tPvPGIP2_5-8 clearly slows down the growth of A. niger and B. cinerea. Investigation on the sequence-function relationships of PGIP utilizing a combination of site directed mutagenesis and a variety of peptide truncations suggests that LRR5 could have the most essential structural feature for the inhibitory activities, and may be a possible target for the future engineering of PGIP with enhanced activity. This study highlights the potential of plant-derived PGIPs as a candidate for future in planta evaluation as a pest control agent.

Novel biotechnological strategies to combat biotic stresses: polygalacturonase inhibitor (PGIP) proteins as a promising comprehensive option.

Global climate change and combinatorial environmental stresses pose grave challenges to food security and agricultural sustainability. This calls for diverse and futuristic approaches for the development of crops with increased resilience to natural vagaries. Though innumerable strategies involving diverse genes/pathways are being deciphered in plants to aid stress mitigation, the hunt is still on. Furthermore, strategies that work to alleviate a combination of stresses are always pertinent. In this review, we discuss polygalacturonase inhibitor (PGIP) proteins as a plausible option to mitigate multiple biotic stresses. These are ubiquitous cell wall proteins that inhibit the pectin-depolymerizing activity of cell wall loosening enzymes, polygalacturonases (PGs). While plant PGs are those responsible for developmental activities like fruit ripening, pollen tube elongation, etc., PGs from various biotic stress factors like insects, fungal and bacterial pathogens aid in invasion by reducing the plant cell wall rigidity. To counteract, plants secrete PGIPs, which inhibit the pectin hydrolyzing activity of PGs from the attacking pests and pathogens. Multiple approaches in diverse crop species have demonstrated PGIP-based protection against pathogens and insect pests. Additionally, effectual interaction between PGs-PGIP is an important aspect for successful utilization of this approach. Molecular strategies leading to improved PG-PGIPs interaction is a highlight to demonstrate the use of PGIPs as an amenable stress mitigation approach. The review focuses on a comprehensive update on phylogeny of PGIPs, natural variation of resistance as well as their emerging translational utility towards mitigation of various biotic stresses.

The Constitutive Endopolygalacturonase TvPG2 Regulates the Induction of Plant Systemic Resistance by Trichoderma virens.

Trichoderma spp. are opportunistic fungi some of which are commonly present in the rhizosphere. Several species, such as T. virens, are also efficient biocontrol agents against phytopathogenic fungi and exert beneficial effects on plants. These effects are the consequence of interactions between Trichoderma and plant roots, which trigger enhanced plant growth and induce plant resistance. We have previously shown that T. virens I10 expresses two endopolygalacturonase genes, tvpg1 and tvpg2, during the interaction with plant roots; tvpg1 is inducible while tvpg2 is constitutively transcribed. Using the same system, the tomato polygalacturonase-inhibitor gene Lepgip1 was induced at the same time as tvpg1. Here we show by gene disruption that TvPG2 performs a regulatory role on the inducible tvpg1 gene and in triggering the plant immune response. A tvpg2-knockout strain fails to transcribe the inducible tvpg1 gene in neither in vitro in inducing media containing pectin or plant cell walls, nor during the in vivo interaction with tomato roots. Likewise, the in vivo induction of Lepgip1 does not occur, and its defense against the pathogen Botrytis cinerea is significantly reduced. Our data prove the importance of a T. virens constitutively produced endopolygalacturonase in eliciting plant induced systemic resistance against pathogenic fungi.

Pyramiding PvPGIP2 and TAXI-III But Not PvPGIP2 and PMEI Enhances Resistance Against Fusarium graminearum.

Plant protein inhibitors counteract the activity of cell wall-degrading enzymes (CWDEs) secreted by pathogens to breach the plant cell-wall barrier. Transgenic plants expressing a single protein inhibitor restrict pathogen infections. However, since pathogens secrete a number of CWDEs at the onset of infection, we combined more inhibitors in a single wheat genotype to reinforce further the cell-wall barrier. We combined polygalacturonase (PG) inhibiting protein (PGIP) and pectin methyl esterase inhibitor (PMEI), both controlling the activity of PG, one of the first CWDEs secreted during infection. We also pyramided PGIP and TAXI-III, a xylanase inhibitor that controls the activity of xylanases, key factors for the degradation of xylan, a main component of cereal cell wall. We demonstrated that the pyramiding of PGIP and PMEI did not contribute to any further improvement of disease resistance. However, the presence of both pectinase inhibitors ensured a broader spectrum of disease resistance. Conversely, the PGIP and TAXI-III combination contributed to further improvement of Fusarium head blight (FHB) resistance, probably because these inhibitors target the activity of different types of CWDEs, i.e., PGs and xylanases. Worth mentioning, the reduction of FHB symptoms is accompanied by a reduction of deoxynivalenol accumulation with a foreseen great benefit to human and animal health.

Immuno-affinity purification of PglPGIP1, a polygalacturonase-inhibitor protein from pearl millet: studies on its inhibition of fungal polygalacturonases and role in resistance against the downy mildew pathogen.

Polygalacturonase-inhibitor proteins (PGIPs) are important plant defense proteins which modulate the activity of microbial polygalacturonases (PGs) leading to elicitor accumulation. Very few studies have been carried out towards understanding the role of PGIPs in monocot host defense. Hence, present study was taken up to characterize a native PGIP from pearl millet and understand its role in resistance against downy mildew. A native glycosylated PGIP (PglPGIP1) of ~43 kDa and pI 5.9 was immunopurified from pearl millet. Comparative inhibition studies involving PglPGIP1 and its non-glycosylated form (rPglPGIP1; recombinant pearl millet PGIP produced in Escherichia coli) against two PGs, PG-II isoform from Aspergillus niger (AnPGII) and PG-III isoform from Fusarium moniliforme, showed both PGIPs to inhibit only AnPGII. The protein glycosylation was found to impact only the pH and temperature stability of PGIP, with the native form showing relatively higher stability to pH and temperature changes. Temporal accumulation of both PglPGIP1 protein (western blot and ELISA) and transcripts (real time PCR) in resistant and susceptible pearl millet cultivars showed significant Sclerospora graminicola-induced accumulation only in the incompatible interaction. Further, confocal PGIP immunolocalization results showed a very intense immuno-decoration with highest fluorescent intensities observed at the outer epidermal layer and vascular bundles in resistant cultivar only. This is the first native PGIP isolated from millets and the results indicate a role for PglPGIP1 in host defense. This could further be exploited in devising pearl millet cultivars with better pathogen resistance.

Tracing the evolution of angucyclinone monooxygenases: structural determinants for C-12b hydroxylation and substrate inhibition in PgaE.

Two functionally distinct homologous flavoprotein hydroxylases, PgaE and JadH, have been identified as branching points in the biosynthesis of the polyketide antibiotics gaudimycin C and jadomycin A, respectively. These evolutionarily related enzymes are both bifunctional and able to catalyze the same initial reaction, C-12 hydroxylation of the common angucyclinone intermediate prejadomycin. The enzymes diverge in their secondary activities, which include hydroxylation at C-12b by PgaE and dehydration at C-4a/C-12b by JadH. A further difference is that the C-12 hydroxylation is subject to substrate inhibition only in PgaE. Here we have identified regions associated with the C-12b hydroxylation in PgaE by extensive chimeragenesis, focusing on regions surrounding the active site. The results highlight the importance of a hairpin-ß motif near the dimer interface, with two nonconserved residues, P78 and I79 (corresponding to Q89 and F90, respectively, in JadH), and invariant residue H73 playing key roles. Kinetic characterization of PgaE variants demonstrates that the secondary C-12b hydroxylation and substrate inhibition by prejadomycin are likely to be interlinked. The crystal structure of the PgaE P78Q/I79F variant at 2.4 Å resolution confirms that the changes do not alter the conformation of the ß-strand secondary structure and that the side chains of these residues in effect point away from the active site toward the dimer interface. The results support a catalytic model for PgaE containing two binding modes for C-12 and C-12b hydroxylations, where binding of prejadomycin in the orientation for C-12b hydroxylation leads to substrate inhibition. The presence of an allosteric network is evident based on enzyme kinetics.

Brassica napus polygalacturonase inhibitor proteins inhibit Sclerotinia sclerotiorum polygalacturonase enzymatic and necrotizing activities and delay symptoms in transgenic plants.

Sclerotinia sclerotiorum releases a battery of polygalacturonases (PGs) during infection, which the host plant may cope with through production of polygalacturonase inhibitor proteins (PGIPs). To study the interaction between S. sclerotiorum PGs and Brassica napus PGIPs, 5 S. sclerotiorum PGs and 4 B. napus PGIPs were expressed in Pichia pastoris. SsPG3, SsPG6, and BnPGIP1 were successfully produced in the yeast system, and BnPGIP1 inhibited SsPG6 enzymatic activity in vitro. SsPG3 and SsPG6 both induced light-dependent necrosis when infiltrated into leaves, which was reduced in an Arabidopsis thaliana line expressing BnPGIP2 and to a lesser extent in a line expressing BnPGIP1. The line expressing BnPGIP2 also exhibited a delay in the onset of symptoms upon S. sclerotiorum inoculation, but no long-term effect on S. sclerotiorum disease progression was observed. The P. pastoris system was found to be suitable for expressing high levels of some S. sclerotiorum PGs, but PGIP interaction studies were best performed in planta. Arabidopsis thaliana forms necrotic lesions upon infiltration of PGs, is susceptible to S. sclerotiorum, and is easily transformed, and thus, is well-suited for the qualitative study of PG-PGIP interactions.

Regulation of the grapevine polygalacturonase-inhibiting protein encoding gene: expression pattern, induction profile and promoter analysis.

Regulation of defense in plants is a complex process mediated by various signaling pathways. Promoter analysis of defense-related genes is useful to understand these signaling pathways involved in regulation. To this end, the regulation of the polygalacturonase-inhibiting protein encoding gene from Vitis vinifera L. (Vvpgip1) was analyzed with regard to expression pattern and induction profile as well as the promoter in terms of putative regulatory elements present, core promoter size and the start of transcription. Expression of Vvpgip1 is tissue-specific and developmentally regulated. Vvpgip1 expression was induced in response to auxin, salicylic acid and sugar treatment, wounding and pathogen infection. The start of transcription was mapped to 17 bp upstream of the ATG and the core promoter was mapped to the 137 bp upstream of the ATG. Fructose- and Botrytis responsiveness were identified in the region between positions -3.1 and -1.5 kb. The analyses showed induction in water when the leaves were submersed and this response and the response to wounding mapped to the region between positions -1.1 and -0.1 kb. In silico analyses revealed putative cis-acting elements in these areas that correspond well to the induction stimuli tested.

Protein trafficking to the cell wall occurs through mechanisms distinguishable from default sorting in tobacco.

The secretory pathway in plants involves sustained traffic to the cell wall, as matrix components, polysaccharides and proteins reach the cell wall through the endomembrane system. We studied the secretion pattern of cell-wall proteins in tobacco protoplasts and leaf epidermal cells using fluorescent forms of a pectin methylesterase inhibitor protein (PMEI1) and a polygalacturonase inhibitor protein (PGIP2). The two most representative protein fusions, secGFP-PMEI1 and PGIP2-GFP, reached the cell wall by passing through ER and Golgi stacks but using distinct mechanisms. secGFP-PMEI1 was linked to a glycosylphosphatidylinositol (GPI) anchor and stably accumulated in the cell wall, regulating the activity of the endogenous pectin methylesterases (PMEs) that are constitutively present in this compartment. A mannosamine-induced non-GPI-anchored form of PMEI1 as well as a form (PMEI1-GFP) that was unable to bind membranes failed to reach the cell wall, and accumulated in the Golgi stacks. In contrast, PGIP2-GFP moved as a soluble cargo protein along the secretory pathway, but was not stably retained in the cell wall, due to internalization to an endosomal compartment and eventually the vacuole. Stable localization of PGIP2 in the wall was observed only in the presence of a specific fungal endopolygalacturonase ligand in the cell wall. Both secGFP-PMEI1 and PGIP2-GFP sorting were distinguishable from that of a secreted GFP, suggesting that rigorous and more complex controls than the simple mechanism of bulk flow are the basis of cell-wall growth and differentiation.

Polygalacturonase-inhibitor proteins in pearl millet: possible involvement in resistance against downy mildew.

Polygalacturonase-inhibitor protein (PGIP) is a defense protein found in plant cell walls. It prevents the degradation of pectin by modulating the endo-polygalacturonase activity. The present study has used heterologous anti-bean PGIP probes to investigate the role of PGIP in pearl millet [Pennisetum glaucum (L) R. Br.] resistance against downy mildew caused by oomycete pathogen Sclerospora graminicola (Sacc.) Schroet. Northern blot analysis using bean pgip2 DNA fragment as probe showed an early and marked induction of transcripts (∼1.2 kb) upon pathogen-inoculation in pearl millet cultivar resistant to downy mildew, with the maximum level observed at 24 and 48 h post-inoculation (h.p.i.). Western blot analysis of pearl millet total cell wall proteins using antibodies against bean PGIP showed the presence of a major band of ∼43 kDa, and several minor ones. The protein accumulation was higher in resistant seedlings than in susceptible seedlings with a differential expression observed only in the case of incompatible interaction. Immunocytochemical localization in epidermal peelings of coleoptiles and tissue-printing showed a similar trend in the PGIP accumulation. PGIP was found to localize in the epidermal as well as in the vascular regions of tissues. Higher accumulation was observed in the stomatal guard cells of resistant cultivar inoculated with the pathogen. PGIP activity of pearl millet total protein extracts when assayed against Aspergillus niger PG displayed differential PG inhibitory activities between the resistant and suceptible cultivars with resistant sample showing the highest inhibition of 16%, post-pathogen treatment. Thus, PGIP appeared to be an important player in pearl millet-S. graminicola interaction leading to host resistance.

Natural resistance against pre- and post-harvest pathogens in Sardinian pears germoplasm.

Natural resistance against Venturia pirina and Botrytis cinerea was investigated within the Sardinian pears germplasm. The natural occurrence of V. pirina was monitored at the orchard level for 5 years, while resistance against B. cinerea was evaluated by a bioassay using methanolic extracts of the fruit rind or by artificially inoculation. Methanolic extracts of the leaves were employed for the Cladosporium bioassay on TLC plates. Among the accessions, 8 resulted sensitive to V. pirina, vegetation and fruit are severely affected every year. Seven showed an intermediate behaviour and the sole vegetation was affected slightly in two out of the 5 years. Two inhibition spots, in the methanolic extract of the leaves, were clearly evidenced in 55 accessions and a positive correlation was found between the presence of these antifungal compounds in the leaves and the resistance to V. pirina in the field. The artificial inoculation of fruit with B. cinerea evidenced a great variability in resistance, with about 12 accessions that could be considered having a good resistance. The fruit rind methanolic extracts evidenced several inhibition spots in most accessions, but no correlation could be found with fruit resistance to B. cinerea. In addition, the activity of the polygalacturonase inhibiting protein (PGIP), determined with an agarose diffusion bioassay, indicate a positive correlation between the PGIP activity evidenced in the core tissue and the infection degree by B. cinerea.

[The effect of physiologically active compounds on the production of ethylene and the activity of polygalacturonase inhibiting protein in fruits].

The treatment of apple and banana fruits with 2-CEFA and ethacyde induced the production of ethylene and accelerated the ripening and accumulation of ACC in apple fruits. Inhibitors AOA, AVG, and CoCl2 acted at the different steps of ethylene biosynthesis, inhibited the physiological aging process and increased storage longevity. Treatment with astaxantine and BOA delayed the pick of ethylene production by fruits. The content of PGIP was correlated with intensity of ethylene production. The infection of fruits with phytopathogenic microorganisms lowered as the result of the inhibition of pathogen PG. The dynamics of PGIP activity in fruits suggests its important role in the processes of ripening.

Two polygalacturonase-inhibiting proteins (VrPGIP) of Vigna radiata confer resistance to bruchids (Callosobruchus spp.).

Bruchids (Callosobruchus spp.) are destructive storage pests of mung beans (Vigna radiata). Bruchids infest mature seeds during storage and in the field causing heavy losses. Bruchid resistance in mung bean has been characterized as a dominant trait controlled by a single gene. Several independent mapping studies showed that the Br locus on chromosome 5 was a key quantitative trait loci (QTL) involved in bruchid resistance. Two polygalacturonase-inhibitor protein (PGIP) family genes, VrPGIP1 and VrPGIP2, located in the Br locus may be the primary genes responsible for bruchid resistance in mung bean but no experimental proof is available. We isolated the VrPGIP1 and VrPGIP2 genes from bruchid resistant mung bean cultivar V2802 and purified the proteins by prokaryotic expression. Both VrPGIP1 and VrPGIP2 had polygalacturonase inhibitor activity and both of the PGIP proteins conferred resistance to bruchids in an artificial seed test system. VrPGIPs can inhibit the enzyme activity of polygalacturonase present in males, females and fourth instar larvae of C. maculatus. These results demonstrated that VrPGIP1 and VrPGIP2 play a critical role in bruchid resistance probably through inhibiting polygalacturonase activity.

The pro-form of Stereum purpureum endopolygalacturonase I is inactivated by a pro-sequence in the C-terminal region.

The Pro-form (Pro-EndoPG I) of Stereum purpureum endopolygalacturonase I has a unique C-terminal region (pro-sequence) that is lacking in PGs of other origins. Mature EndoPG I purified from the culture filtrate of this fungus does not have the 44-amino-acid pro-sequence present in Pro-EndoPG I. We expressed Pro-EndoPG I in Escherichia coli and examined its activity. It was found that Pro-EndoPG I had no PG activity, but that PG activity was acquired after the degradation of part of the pro-sequence with V8 protease. These results suggest that the pro-sequence inactivates auto-PG activity. No similar characteristic has been reported for any glycoside hydrolase. We then constructed EndoPG I mutants and identified two Glu residues, E364 and E366, that were related to auto-inactivation. A test involving injection of the enzyme into apple trees showed that Pro-EndoPG I induced the same silver-leaf symptoms as mature EndoPG I.

Polygalacturonase inhibitor protein from fruits of anthracnose resistant and susceptible varieties of Chilli (Capsicum annuum L).

Chilli fruit is highly susceptible to anthracnose infection at the stage of harvest maturity, due to which the fruit yield in the leading commercial variety Byadgi is severely affected. Field studies on screening of several varieties for resistance to anthracnose have shown that a variety of chilli AR-4/99K is resistant to anthracnose infection. In many crops, resistance to fungal attack has been correlated with PGIP activity in developing fruits based on which transgenic varieties have been developed with resistance to fungi. The present study was carried out to determine whether anthracnose resistance in AR-4/99K was due to the increased levels of PGIP alone and/ or due to differences, if any, in the properties of PGIP. Hence, a comparative study of the properties of polygalacturonase inhibitor protein (PGIP) isolated from fruits of anthracnose resistant chilli var AR-4/99K and a susceptible variety Byadgi was conducted with the objective of utilizing the information in genetic transformation studies. Both the PGIPs from anthracnose resistant and susceptible varieties of chilli exhibited similarities in the elution pattern on Sephadex gel, DEAE cellulose, PAGE and SDS-PAGE. The two PGIPs were active over a wide range of pH and temperature. Both PGIPs showed differential inhibitory activity against polygalacturonase (PG) secreted by Colletotrichum gleosporoides, C. capsici, C. lindemuthianum, Fusarium moniliforme and Sclerotium rolfsii. The inhibitory activity of PGIP from both resistant and susceptible varieties was the highest (82% and 76%, respectively) against the PG from Colletotrichum capsici, a pathogen causing anthracnose rot of chilli, while the activity was lower (1.27 to 12.3%) on the other fungal PGs. Although PGIP activity decreased with fruit maturation in both the varieties, the resistant variety maintained a higher activity at 45 days after flowering (DAF) as compared to the susceptible variety which helped it to overcome the infection by anthracnose as against the susceptible variety (Byadgi) in which PGIP activity was drastically reduced at maturity. The molecular mass of PGIP as determined by SDS-PAGE was found to be 37 kDa. N-terminal sequence analysis of the PGIP showed the first six amino acid residues from N-terminal end were Asp-Thr-His-Lys-Ser-Glu (DTHKSE), respectively. The similarities in properties of the two PGIPs support the earlier findings that resistance of AR-4/99K to anthracnose fungus is a result of its higher PGIP activity at maturity.

Preparation of alginate oligosaccharide and its effects on decay control and quality maintenance of harvested kiwifruit.

Alginate oligosaccharide (AOS) is a biological carbohydrate formed from the degradation of sodium alginate. AOS used in this study was enzymatically prepared and had varying degrees of polymerization (2-8). AOS applied to harvested kiwifruit stored at 25 °C inhibited gray mold, blue mold, and black rot. AOS inhibited pectin solubilization, gene expression of pectin methylesterase and polygalacturonase, and the corresponding enzyme activity of their encoded proteins in kiwifruit. In contrast, AOS induced antioxidant gene expression and enzyme activity, including catalase and superoxide dismutase. The level of total phenols and flavonoids in kiwifruit was also elevated. AOS treatment also had a beneficial effect on fruit quality. Collectively, the results indicate that postharvest treatment with AOS inhibits postharvest decay and prolongs fruit quality by suppressing cell wall degradation and eliciting antioxidants in harvested kiwifruit. AOS has the potential to be used to preserve and extend the postharvest quality of kiwifruit.

Three aspartic acid residues of polygalacturonase-inhibiting protein (PGIP) from Phaseolus vulgaris are critical for inhibition of Fusarium phyllophilum PG.

Polygalacturonase-inhibiting proteins (PGIPs) are plant cell wall proteins that specifically inhibit the activity of endopolygalacturonases (PGs) produced by fungi during the infection process. The interaction with PGIPs limits the destructive potential of PGs and may trigger plant defence responses through the release of elicitor active oligogalacturonides. In order to pinpoint the residues of PvPGIP2 from Phaseolus vulgaris involved in the interaction with PGs, we used site-directed mutagenesis to mutate the residues D131, D157 and D203, and tested for the inhibitory activity of the mutant proteins expressed in Pichia pastoris against Fusarium phyllophilum and Aspergillus niger PGs. Here, we report that mutation of these residues affects the inhibition capacity of PvPGIP2 against F. phyllophilum PG.

Depression of Fungal Polygalacturonase Activity in Solanum lycopersicum Contributes to Antagonistic Yeast-Mediated Fruit Immunity to Botrytis.

The acquisition of susceptibility to necrotrophy over the course of ripening is one of the critical factors limiting shelf life. In this study, phytopathology and molecular biology were employed to explore the roles of pectinase in fruit susceptibility and ripening. Solanum lycopersicum fruit softened dramatically from entirely green to 50% red, which was accompanied by a continuously high expressed SlPG2 gene. The necrotrophic fungus Botrytis cinerea further activated the expression of SlPGs and SlPMEs to accelerate cell wall disassembly, while most of the polygalacturonase inhibitor proteins encoding genes expression were postponed in ripe fruit following the pathogen attack. Pectin induced the antagonistic yeast to secrete pectinolytic enzymes to increase fruit resistance against gray mold. The activities of pathogenic pectinase of B. cinerea were correspondingly depressed in the pectin-inducible yeast enzyme elicited ripe fruit. These data suggest that pectinase is a molecular target for regulation of disease resistance during fruit ripening.

A single amino acid substitution in highly similar endo-PGs from Fusarium verticillioides and related Fusarium species affects PGIP inhibition.

Endo-polygalacturonase (PG) may be a critical virulence factor secreted by several fungi upon plant invasion. The single-copy gene encoding PG in Fusarium verticillioides and in eight other species of the Gibberella fujikuroi complex (F. sacchari, F. fujikuroi, F. proliferatum, F. subglutinans, F. thapsinum, F. nygamai, F. circinatum, and F. anthophilum) was functionally analyzed in this paper. Both the nucleotide and amino acid sequences were highly similar among the 12 strains of F. verticillioides analyzed, as well as among those from the G. fujikuroi complex. The PGs were not inhibited by the polygalacturonase-inhibiting proteins (PGIPs) from the monocot asparagus and leek plants, but were inhibited to variable extents by bean PGIP. PGs from F. verticillioides, F. nygamai and one strain of F. proliferatum were barely inhibited. Residue 97 within PG was demonstrated to contribute to the different levels of inhibition. Together these findings provide new insights into the structural and functional relationships between the PG from the species of the G. fujikuroi complex and the plant PGIP.

Polygalacturonase-inhibiting protein is a structural component of plant cell wall.

It is generally believed that plants "evolved a strategy of defending themselves from a phytopathogen attack" during evolution. This metaphor is used frequently, but it does not facilitate understanding of the mechanisms providing plant resistance to the invasion of foreign organisms and to other unfavorable external factors, as well as the role of these mechanisms in plant growth and development. Information on processes involving one of the plant resistance factors--polygalacturonase-inhibiting protein (PGIP)--is considered in this review. The data presented here indicate that PGIP, being an extracellular leucine-rich repeat-containing protein, performs important functions in the structure of plant cell wall. Amino acid residues participating in PGIP binding to homogalacturonan in the cell wall have been determined. The degree of methylation and the mode of distribution of homogalacturonan methyl groups are responsible for the formation of a complex structure, which perhaps determines the specificity of PGIP binding to pectin. PGIP is apparently one of the components of plant cell wall determining some of its mechanical properties; it is involved in biochemical processes related to growth, expansion, and maceration, and it influences plant morphology. Polygalacturonase (PG) is present within practically all plant tissues, but the manifestation of its activity varies significantly depending on physiological conditions in the tissue. Apparently, the regulation of PG functioning in apoplast significantly affects the development of processes associated with the modification of the structure of plant cell wall. PGIP can regulate PG activity through binding to homogalacturonan. The genetically determined structure of PGIP in plants determines the mode of its interaction with an invader and perhaps is one of the factors responsible for the set of pathogens causing diseases in a given plant species.