Biotechnological Approaches to Disease Resistance

Biotechnological Approaches to Disease Resistance

 Aim: to enhance plant defence systems
 Approaches:

(1) Protection against pathogens
 PR protein show some activity at inhibiting disease, but have limited activity, generally do not switch on disease resistance pathway
 The PR proteins: eg chitinase, glucanase and ribosome-inactivating proteins (RIPs)
 Provide protection and tolerance over fungal infection
 chitinase, glucanase degrade cell wall of many fungi
 RIP – enzyme that remove an adenine residue from a specific site in the large rRNA of eukaryote & prokaryote ribosome à inhibit protein synthesis
 Isolated chitinase, glucanase genes fr plant (rice, barley), bacteria (Serratia marcescens), fungi (Trichoderma harzianum)
 Common promoters used: 35s promotor, wound-inducible promotor (potato prp1-1 promotor – mediate rapid & localized expression in response to pathogen, restrict gene expression to where the protein is required & may reduce any potential hazards associated with the protein)
 Advantage: gene expression only turn on during infection à limiting gene expression à no serious drain on plant biosynthetic capability & yield reduction
 Common promoters used: 35s, wound-inducible
 Most widely used in fruit trees
 Challenge?
Greenhouse  field

(2) Antimicrobial proteins
 Antifungal and antibacterial, eg lysozyme, defensins
A. Lysozyme
 Degrade chitin and peptidoglycan
B. Defensins
 Small antimicrobial peptides
 Display lytic activity difficult to develop resistance
 Eg alfalfa defensin, cecropin, melittin, gallerimycin, temporins etc
 Problem? Limited spectrum

Case study:
Combating the bacterial disease fireblight
 Cause by Erwinia amylovora
 Solution? Transgenic crop containing gene code for antimicrobial protein
 2 Approaches:
(1) Lytic approach – produce transgenic fruit trees contain gene for T4 lysozyme + insect antimocrobial pt attacin E + cecropin
(2) Bovine lactoferrin gene (to compete with microbial siderophores) + depolymerase gene (degrade specific exopolysaccharides)
Siderophores - small, high-affinity iron chelating compounds secreted by grasses and microorganisms such as bacteria and fungi,
- amongst the strongest soluble Fe3+ binding agents known.

(3) Transgenic crops for food safety
 How? By reduce amount of mycotoxins
 Use to combat the effect of fungal infection
 Example:
 Bt genes into Bt crops  minimize insect damage  less wound sites due to insect infection  lower possibility for fungal infection  reduce amount of mycotoxins  ensure safety of livestock and human

(4) Introduction of HR and SAR in transgenic plant
 Aim: use elicitor receptor system to switch on general resistance pathway

 Approach 1: the Guard hypothesis (tomato)
 R protein  increase its production will increase the expression of resistance systems
 Outcomes:
 Overexpression of R protein  genes were induced differently
 Not pathogen-specific

 Approach 2: (tobacco) switch on the HR-related defence system
 crytogein (a Phytophthora cryptogea gene that codes for highly active elicitor) fuse with pathogen inducible promoter (from tobacco) à introduce into transgenic tobacco
 Non-induced condition (eg no pathogen)àno elicitor
 Once infected by virulent fungus (Phytophthora parasitica var. nicotianae) à crytogein gene is expressed à elicitor-induced defense gene expressed
 Broad-spectrum resistance

Other approaches
 Trigger expression of a range of genes by using different stimuli, eg introduction of Arabidopsis gene NPR1 into tomato, wheat, rice etc.
 The product of this gene lies at a node in the regulatory pathways that link SAR, induced systemic resistance, R-gene mediated resistance, ethylene, salicylic acid, jasmonic acid etc
 it activates gene expression by interacting with members of the TGA family of basic leucine zipper (bZIP) transcription factors
 The introduction of the gene enhanced resistance to a range of plant pathogens in tomato, but in rice the gene had a detrimental effect
 The mode of action for the transgene seems to be a priming function: the plant responds more rapidly to attack rather than having its defence system switched on constitutively, which would have an effect on yield
 Switching on cascade of defense genes (sunflower)
 Involve of O sp eg H2O2 as a messenger in HR reaction has been tested by transforming plant with oxalate oxidase from wheat
oxalate oxidase
 oxalic acid+ O2 -----------------------------------> CO2+ H2O2
 transgenic plant could degrade oxalic acid & generate H2O2, and an HR-like response along with salicylic acid synthesis & defence-gene expression occur
 The genes were identified as being PR & defensin protein
 Transgenic sunflower exhibited enhanced resistance against the oxalic acid-generating fungus Sclerotinia sclerotiorum