RNAi Technology for Improving Disease Resistance
Authors: PRIYANKA CHANDRA & POONAM JASROTIA

RNAi interference (RNAi) is becoming one of the major plant functional genomics tool. It is a biological process in which RNA molecules inhibit gene expression, typically by causing the destruction of specific mRNA molecules. The other names of RNAi are co-suppression, post-transcriptional gene silencing (PTGS), and quelling. It requires production of double-stranded RNA (dsRNA) homologous to the gene being targeted for post transcriptional gene silencing (PTGS). This dsRNA is processed into approximately 21-nucleotide RNAs, known as small interfering RNAs (siRNAs), by the enzyme Dicer. These siRNAs then provide specificity to endonuclease-containing, RNA induced silencing complex (RISC), which targets homologous RNAs for degradation [1-3] .

RNAi can be applied for crop improvement by two methods:

1. Silencing of specific plant genes are known to improve agronomic traits or increase plant fitness against stress. The method used for generation of plants with stable gene silencing is called host gene silencing -hair pin RNAi (HGS hpRNAi). In this method, genetically engineering is done in host plant to alter its own gene expression for improvement for agronomic superiority or for improvement against stresses, both biotic and abiotic.

2. Other method is hdRNAi, in which gene(s) are silenced only in target organism (e.g. insect or pathogen or plant parasite) that damages crop. In this method, the plant is provided with ability to silence the gene of target organism to defend against several biotic stresses. This hdRNAi approach can be sub- divided into two subtypes:

• hdRNAi-1: In this approach, silencing of genes is done in organisms that externally feed on the plant and cause damage. This hdRNAi-1 can be used to silence genes in biotic stress causal agents like insect pests, nematodes and parasitic weeds.

• hdRNAi-2: Second method is targeting gene silencing in viruses that enter into the host plant cell.

HGS-hpRNAi method: This can be used to improve plant disease resistance against bacterial and fungal pathogens. In Arabidopsis, bacterial component, flagellin, induces expression of a specific miRNA, which in turn leads to down-regulation of signaling pathways that increases the plant's resistance. This study clearly indicated the potential of gene down-regulation to control disease resistance in plants.

Application of RNAi-mediated oncogene silencing to control crown gall disease has been reported, and tobacco plants silenced for glutathione S-transferase gene transcripts showed resistance to black shank disease.

Further, RNAi-mediated knockdown of gene OsSSI2 markedly enhanced resistance in rice against leaf blight bacterium Xanthomonas oryzae and blast fungus Magnaporthe grisea. Disease resistance against Magnaporthe grisea in rice can be created by suppression of two fatty acid desaturases (OsFAD7 and OsFAD8 genes). The studies carried out by several researchers proved that genes related to fatty acid metabolism can be the targets for developing disease resistant RNAi transgenic crop plants.

hdRNAi method: This method is not explored in the aspect of silencing of bacterial or fungal pathogen genes through hdRNAi-1 method.

Potential target gene for hdRNAi-1 can be from one or either all of three types of pathogen genes namely
(i) essential genes involved in pathogen metabolism;
(ii) genes which cause pathogen to be resistant to plant toxins;
(iii) genes that encode effectors that are involved in pathogenicity.

hdRNAi-2 based strategy has been used to engineer plants to impart virus resistance. In this case RNAi vector carrying viral target sequence in transgenic plants produce dsRNA that eventually silences virus multiplying in the cell. Transgenic tomato plants producing dsRNA against Potato spindle tuber viroid (PSTVd) sequences exhibited resistance to PSTVd infection. Similarly, cassava plants were successfully engineered to resist African cassava mosaic virus (ACMV).

dsRNA spray: This method used to control virus infection in plants. dsRNA derived from viral sequences is inoculated on plant leaves which prevents virus infection. Mass production of dsRNA can be done either through in vitro-transcription or in vivo expression in bacteria. Although, field trial of feasibility of dsRNA spray method is yet to be tested

Conclusion:

RNA interference (RNAi) is one such tool widely used to analyze gene function. RNAi is also proved to be a tool for plant researchers to produce improved crop varieties. RNAi based concepts have potential applications in plant functional genomics and agriculture.

References:

1. Agrawal, N., Dasaradhi, P. V. N., Mohmmed, A., Malhotra, P., Bhatnagar, R. K. & Mukherjee, S. K. (2003). RNA interference: biology, mechanism, and applications.Microbiology and Molecular Biology Reviews 67, 657-685.
2. Qi, Y. & Hannon, G. J. (2005). Uncovering RNAi mechanisms in plants: Biochemistry enters the foray. FEBS Letters 579, 5899-5903.
3. Watson, J. M., Fusaro, A. F., Wang, M. & Waterhouse, P. M. (2005). RNA silencing platforms in plants. FEBS Letters 579, 5982-5987.


About Author / Additional Info:
I am working as Scientist in ICAR-Indian Institute of Wheat and Barley Research . My specialization is in Agricultural Microbiology. The Co-author of the article is Poonam Jasrotia is agricultural entomologist in the same institute.