In this article we will discuss about the fungal resistance in plants.
Several agricultural crops are severly damaged by fungal infections, which result in dramatic yield loss. The spread of fungal disease can be combated by conventional strategies like breeding for resistance by variation of the cultural practice and by fungicide treatment.
However, a promising alternation that has emerged recently is based on the strengthening of endogenous defence potentialities of the plants because plants are well known to trigger immune like response i.e., systemic aquired resistance (SAR) to fungal challenge.
Plants accumulate number of pathogenic related proteins (PR) during infection. PR proteins enhanced acquired resistance in plants. There are five families of PR proteins. PR, to PRS. The PR, family is acidic proteins with 15 kD molecular weight, PR2 and PR3 proteins are identified as β-1, 3-glucanase, chitinase etc.
Chitinase:
Different class of plant chitinase accumulation can provide protection against fungal infection. Regulation of chitinase genes takes place during development stage but stress conditions such as fungal attack can trigger its expression. Even in the case of fungal inoculation, enhanced activity was noticed in the defence response targeted towards chitin as a major cell wall component in the fungi.
β-1, 3-Glucanase:
In vitro experiments show that either alone or combination, chitinase and β-1, 3-glucanase inhibited most fungi tested. The pea pathogen. Fusarium solani strongly inhibited by chitinase and β-1, 3-glucanase indicating that these enzymes can be employed as a weapon to combat several pathogenic fungi.
High activity of chitinase and β-1, 3-glucanase is frequently found in higher plants, but substrate for this enzyme is absent in the plant. Similarly, substrate for β-1, 3-glucanase is callose is usually present only in small quantities. Availability of substrate in abundance could be present only in cell wall of many fungi.
Since chitinase are induced upon infection with different kinds of pathogen. It has been proposed that they are participated in active defence response of plants. Atleast five classes of plant chitinases are well characterized on the basis of sequence alignments or amino acid sequence. Class I chitinase contains N-terminal cyst-rich amino acid.
Three of which abundantly found in plants. Similarly, cDNA clones encoding class I chitinase (chtc) isolated from potato leaves. Ancillo et al. (1999) reported that infection of potato leaves with phytopthora infestans resultant in accumulation of both chit. C and chit. B mRNA but encoded protein could not be detected. Further studies reveals that both chitinase proteins and mRNA in young leaves and stems are predominently located to the epidermis.
Transgenic technique provides excellent opportunities for the plants to combat fungal attack. High level expression of chitinase and 1, 3-β-glucanase in transgenic plants resulted in enhanced levels of fungal resistance. Expression of class I catalase gene activate the endogenous homologous gene in tobacco plants resulted in disease resistant in transgenic plants.
Roby et al. (1990) has studied the regulation of chitinase 5B gene from bean plants. Transgenic plants containing a chimeric gene composed of 1.7 kb fragment carrying promoter is activated during attack by fungal pathogen like Rhizoctonia solani, sclerotia rolfi etc.
The chimeric gene construct was made by fusing this gene to GUS gene. Regulation of chitinase gene activation during fungal attack has been well investigated. The results indicated that chitinase 5B-gus A fusion gene construct is used to assess the molecular details of activation of plant defence system during pathogen attack.
PR Proteins:
Xu Hu and Reddy (1997) in a different line of investigation characterised pathogenesis related PR5 like (ATLP-3) proteins also called thaumatin like proteins due to their similarities with sweetening thaumatin protein. PR5 proteins are well documented in dicotyledon and monocotyledonous plants. Cloning and overexpression of PR5-like protein in transgenic potato plants exhibit enhanced resistance to phytopthora infestans.
Hu and Reddy (1997) isolated cDNA of ATLP-3 (Arabidopsis thaumatin like protein) from Arabidopsis which shares high similarities with PR5 proteins from the same plant and its expression studies shows that ATLP-3 may be involved in plant defence response to fungal attack. Another sub-group of PR5-proteins from monocot of molecular weight 17.5 kD are also induced by pathogen and shows considerable level of antifungal activity.
KP or KP4:
Apart from pathogen related protein mediated resistance, several other antifungal system, which are prevalent in plant system could be exploited as a novel gene pool. Such antifungal systems have been described in virus infected Ustilago maydis strain. It was shown that several fungal species (nearly 60) are infected by virus like particle.
It is, however, shows that only Saccharomyces cerevisiae and Ustilago maydis infected with double stranded RNA virus. These strains secrete protein called Killing protein (KP). This KP proteins exhibit antifungal activity. In transgenic experiment cDNA encoding antifungal protein KP4 desired from virus, which infect Ustilago was inserted in maize plant driven under the control of ubiquitin promoter.
These are then genetically transferred to certain susceptible wheat variety. Out of seven transgenic lines, three generations expressing KP4 transgene showed antifungal activity against Ustilago maydis. These transgenic lines also exhibit protection against smut and other bunt disease.
In addition, engineering plants with broad spectrum resistance to fungal phytophagus have been reported. Osusky (2000) expressed synthesic gene encoding a terminus modified cecropin-melittin cationic peptide chimeric (MsrA1) exhibit broad spectrum antimicrobial activity in transgenic potato cultivar.
Plant Defensins against Fungal Pathogens:
Although, significant endeavour has been made towards the identification of antifungal proteins and their efficacy in controlling pathogenic fungus, single antifungal protein gene was found to be unsuccessful in providing protection in the field. Hence, good option for the production of fungal resistance is the utilization of certain plant derived defensin gene.
Defensins are small cysteine-rich peptides exhibits high microbial activity. The alfa alfa antifungal peptide, designated as alfAFP, defensins isolated from seeds to Medicago sativa displays strong antifungal activity against fungal pathogen Verticillum dahline. This alf AFP defensin antifungal peptide (5.6 kDa) inhibits the elongation of pre-germinated spores by 50% at the conc. of 5 µg/ml and completely inhibits at 15 µg/ml.
The full length alf AFP cDNA was cloned in potato plant driven under the control of 35 S promoter of figwort mosaic virus (FMV) with a single peptide provide robust resistance in controlled conditions as well as in the field.
This antifungal protein is also effective against control of other plant fungal pathogen such as Alternaria solani and Fusarium culmorum causal agent of potato early blight and wheat-head scab respectively.
There have been report on the antifungal activity of hevein-like protein from Pharbisis nil. Hevein and hevein-like small molecular weight proteins are antimicrobial proteins (AMPs) which have been reported earlier from several plants.
All of these AMPs are peptides containing 40-43% amino acid residues and possess a characteristic cysteine/glycine rich chitin binding domain. These proteins are able to bind to fungal chitin in the cell wall and disrupt cell wall synthesis.
The hevein-like antimicrobial protein from P nil exhibit broad spectrum antifungal activity. When cDNA of Pn-AMP class gene was constitutively expressed in tomato under the control of CaM35S promoter, the transgenic plants showed enhanced resistance against both the non-chitinous fungus Phytopthera capsici and the chitin containing fungus Fusarium oxysporum.