RNA i Stratagies to manage plant viruses

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1 WEL COME

RNA i STRATEGIES TO MANAGE PLANT VIRUSES : 

RNA i STRATEGIES TO MANAGE PLANT VIRUSES MURALI, R PAL 0243 DEPT. OF PLANT PATHOLOGY

INTRODUCTION: : 

RNA interference ( RNAi ) is a process within living cells that moderates the activity of their genes. Historically, it was known by other names, co-suppression , post transcriptional gene silencing (PTGS), and quelling . Using dsRNA to silence gene expression through binding, cleaving and degrading complimentary endogenous mRNA. INTRODUCTION: What is RNAi ?

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In 2006 , Andrew Fire and Craig C. Mello shared the Nobel Prize in Physiology or Medicine for their work on RNA interference in the nematode worm C. elegans , which they published in 1998 .

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RNA silencing occurs in a wide variety of organisms, including protozoa, fungi, plants and animals and involves recognition of a target RNA and initiation of a sequence-specific RNA degradation pathway in the cytoplasm. This mechanism is conceived as a natural antiviral defence system in plants that is activated as a response to double-stranded RNA (dsRNA) formed during virus replication.

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6 Plant Pathogenic Virus RNA virus DNA virus Plant pathogenic viruses - >900 species

Why is it important?: 

Why is it important? Method to study the function of genes Provides a basis of discovering new drugs capable of silencing viral or human genes that cause diseases. (i.e HIV infection)

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8 Post-transcriptional gene silencing (PTGS) Antisense RNA Sense RNA Defective interfering and satellite RNA The siRNA The microRNA (miRNA) Artificial miRNAs (amiRNAs) Ribozymes RNA -mediated resistance

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Two types of small ribonucleic acid (RNA) molecules micro RNA (miRNA) small interfering RNA (siRNA) – Are central to RNA interference. RNAs are the direct products of genes, and these small RNAs can bind to other specific messenger RNA (mRNA) molecules and either increase or decrease their activity, by preventing an mRNA from producing a protein. RNA interference has an important role in defending cells against parasitic genes – viruses and transposons.

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Main component of RNAi machinery DICER A protein with RNAse activity cleaves ds RNAs into small interference RNA (siRNA) RISC (RNA induced silencing complex ) protein A protein complex in which the antisense strand of siRNA duplexes guides endonucleolytic cleavage of target RNAs

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DICER

siRNA: 

siRNA 5’ 3’ 5’ 3’

RISC: 

RISC 2 RNA binding proteins RNA/DNA Helicase Translation Initiation Factor RNA-Dependent RNA Polymerase (RdRP) Transmembrane Protein

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Mechanism of post-transcriptional gene silencing Initiation phase -------- Synthesis of dsRNA and production of small interfering RNA (siRNAs) frgments Maintenance phase -------- Association of siRNA complex protein (RISC) to guide nuclease activity and degradation of target mRNA Signal amplification & -------- siRNA act as promoter for dsRNA Spreading phase polymerization which moves through cell to cell.

How RNAi Works : 

How RNAi Works The entry of long double stranded RNA, such as an introduced transgene, a rogue genetic element or a viral intruder, triggers the RNAi pathway of cells. This results in the recruitment of the enzyme Dicer. Dicer cleaves the dsRNA into short, 20-25 basepairs long, fragments, called small interfering RNA (siRNA). An RNA-induced silencing complex (RISC) then distinguishes between the two siRNA strands as either sense or antisense. The sense strands (with exactly the same sequence as the target gene) are degraded.

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The antisense strands on the other hand are incorporated to the RISC. These are used as guide to target messenger RNAs (mRNA) in a sequence-specific manner. Messenger RNAs (mRNA), which codes for amino acids, are cleaved by RISC. The activated RISC can repeatedly participate in mRNA degradation, inhibiting protein synthesis.

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Mechanism of RNAi

RNA i for Disease and Pathogen Resistance : 

RNA i for Disease and Pathogen Resistance Gene silencing was first used to develop plant varieties resistant to viruses. Engineered antiviral strategies in plants mimic natural RNA silencing mechanisms. This was first demonstrated in Potato virus Y- resistant plants expressing RNA transcripts of a viral proteinase gene. Immunity has since been shown to other viruses such as

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Cucumber Mosaic Virus Tobacco Mosaic Virus Tomato Spotted Wilt Virus Bean Golden Mosaic Virus Banana Bract Mosaic Virus and Rice Tungro Bacilliform Virus

Uses: 

Uses RNAi for Disease and Pathogen Resistance RNAi for Male Sterility RNAi and Plant Functional Genomics

Applications: 

Applications Technological applications Gene knockdown Functional genomics Medicine Biotechnology Genome-scale RNAi screening

Conclusion: 

Conclusion RNAi can also provide broad-spectrum resistance against pathogens with high degree of variability, like viruses. Recent studies have hinted possible roles of RNAi-related processes in plant stress adaptation. Although much progress has been made on the field of RNAi over the past few years, the full potential of RNAi for crop improvement remains to be realized.  The complexities of RNAi pathway, the molecular machineries, and how it relates to plant development are still to be elucidated

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