RNA Interference: Promising Approach to Combat Plant Viruses

Abstract

Plant viruses are devastating plant pathogens that severely affect crop yield and quality. Plants have developed multiple lines of defense systems to combat viral infection. Gene silencing/RNA interference is the key defense system in plants that inhibits the virulence and multiplication of pathogens. The general mechanism of RNAi involves (i) the transcription and cleavage of dsRNA into small RNA molecules, such as microRNA (miRNA), or small interfering RNA (siRNA), (ii) the loading of siRNA/miRNA into an RNA Induced Silencing Complex (RISC), (iii) complementary base pairing between siRNA/miRNA with a targeted gene, and (iv) the cleavage or repression of a target gene with an Argonaute (AGO) protein. This natural RNAi pathway could introduce transgenes targeting various viral genes to induce gene silencing. Different RNAi pathways are reported for the artificial silencing of viral genes. These include Host-Induced Gene Silencing (HIGS), Virus-Induced Gene Silencing (VIGS), and Spray-Induced Gene Silencing (SIGS). There are significant limitations in HIGS and VIGS technology, such as lengthy and time-consuming processes, off-target effects, and public concerns regarding genetically modified (GM) transgenic plants. Here, we provide in-depth knowledge regarding SIGS, which efficiently provides RNAi resistance development against targeted genes without the need for GM transgenic plants. We give an overview of the defense system of plants against viral infection, including a detailed mechanism of RNAi, small RNA molecules and their types, and various kinds of RNAi pathways. This review will describe how RNA interference provides the antiviral defense, recent improvements, and their limitations.

Keywords: RNA interference (RNAi); gene silencing; host-induced gene silencing (HIGS); spray-induced gene silencing (SIGS); virus-induced gene silencing (VIGS).

Figure 1

Schematic representation of gene silencing mechanism inside the plant cell. (A) Gene silencing pathway via miRNA. miRNA gene is transcribed by RNA polymerase II. It results in the formation of stem-loop structure, which is termed primary microRNA/pri-miRNA. Dicer enzyme (DCL1) cleaved the stem-loop portion in two steps, forming precursor-microRNA (pre-miRNA) and miRNA/miRNA * duplex. The miRNA/miRNA * duplex is methylated by HEN1 protein. Methylated duplex is detected by Agonuate (AGO1) protein of RNA-Induced Silencing Complex (RISC), which unwinds the duplex. Single-stranded AGO1 guided the methylated miRNA strands, which was hybridized with complementary mRNA. Targeted mRNA undergoes cleavage or translational repression. (B) Gene silencing pathway via siRNA. The inverted repeat sequence of hairpin RNA is transcribed by RNA polymerase III, which generates a double-stranded RNA (dsRNA) structure. dsRNA is cleaved by DCL4 to generate siRNA duplex. siRNA duplex is methylated by HEN1 protein. AGO1 protein detected the methylated duplex, which underwent unwinding of duplex DNA and complementary paired with targeted mRNA. It also results in the degradation/repression of the target gene [38,39,40,41,42,43].