RNA silencing suppressor p19

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RNA silencing suppressor p19
1R9F tombusvirus p19 dimer.png
The protein dimer formed by two p19 molecules. Each monomer is colored from N-terminus (blue) to C-terminus (red) to illustrate the end-to-end orientation of the dimer. The dotted gray line in the center highlights the dimer interface. From PDB: 1R9F ​. [1]
Identifiers
SymbolTombus_p19
Pfam PF03220
InterPro IPR004905

RNA silencing suppressor p19 (also known as Tombusvirus P19 core protein and 19 kDa symptom severity modulator) is a protein expressed from the ORF4 gene in the genome of tombusviruses. These viruses are positive-sense single-stranded RNA viruses that infect plant cells, in which RNA silencing forms a widespread and robust antiviral defense system. The p19 protein serves as a counter-defense strategy, specifically binding the 19- to 21-nucleotide double-stranded RNAs that function as small interfering RNA (siRNA) in the RNA silencing system. By sequestering siRNA, p19 suppresses RNA silencing and promotes viral proliferation. [1] [2] [3] The p19 protein is considered a significant virulence factor [4] and a component of an evolutionary arms race between plants and their pathogens. [5]

Contents

Structure

The structure of the tomato bushy stunt virus p19 protein bound to double-stranded RNA. The two p19 monomers are shown in blue and green; the RNA backbone is shown in orange. The alpha helices at the top and bottom interact with the ends of the RNA, ensuring that only RNA of the correct length is bound. This has been described as a "molecular caliper". From PDB: 1R9F . 1R9F tombusvirus p19.png
The structure of the tomato bushy stunt virus p19 protein bound to double-stranded RNA. The two p19 monomers are shown in blue and green; the RNA backbone is shown in orange. The alpha helices at the top and bottom interact with the ends of the RNA, ensuring that only RNA of the correct length is bound. This has been described as a "molecular caliper". From PDB: 1R9F ​.

The p19 protein received its name from its size, being approximately 19 kilodaltons. It forms a functional homodimer. The crystal structures are available of p19 proteins from the tomato bushy stunt virus [1] and Carnation Italian ringspot virus; [2] the protein consists of a novel protein fold and exemplifies a previously unknown mechanism for binding RNA, using a binding surface formed by a beta sheet and flanked by alpha helices to interact with double-stranded RNAs of around 21 nucleotides in length in a non-sequence-specific manner. [1] [2] [7]

Function

The p19 protein binds to double-stranded RNAs that function as short interfering RNA (siRNA) and is specialized for the 21-nucleotide product of the enzyme DCL4 (a member of a family of plant enzymes with homology to Dicer). [7] By binding to siRNA, p19 sequesters these species and prevents them from interacting with the RNA-induced silencing complex (RISC), a protein complex that mediates the antiviral RNA silencing mechanism in the cell.

The p19 protein is also capable of binding to microRNA molecules that are endogenous to the host cell, as well as the siRNAs that are ultimately derived from the virus's own genome. Notably, an exception to this pattern is p19's inefficiency in interacting with the microRNA miR-168, a regulatory non-coding RNA that represses expression of argonaute-1 (AGO1). The AGO1 protein is required for RNA silencing, thus selectively sparing its repressor from p19's general sequestration of miRNA has the effect of reducing cellular AGO1 levels and is an additional mechanism by which p19 inhibits silencing. [5] [8] The two mechanisms are independent of one another and can be selectively abrogated by mutations. [9]

Evolution

The gene encoding the p19 protein is an example of an overprinted gene, a genomic arrangement common in viruses in which multiple genes are encoded by the same portion of the genome read in alternate reading frames. [10] [11] The open reading frame ORF4, which encodes p19, is completely contained within the open reading frame of another gene, which is designated ORF3 and encodes the movement protein p22. Both genes, and their relative positions, are conserved within the tombusvirus family. [4] [11] P19 is thought to have originated de novo in this lineage. [11] [12]

Sequestration of dsRNA is a common viral counter-defense strategy against RNA silencing, evolved in a form of evolutionary arms race between virus and host. [5] The p19 protein is not unique in this role; in an example of convergent evolution, this strategy appears to have evolved at least three times in distinct viral lineages using proteins with distinct structures and physical means of binding RNA. [3] [13] [14]

History

The tomato bushy stunt virus, which is the type species of the tombusvirus family, is a long-standing model system for the study of plant viruses. The open reading frame encoding p19 was originally discovered in the late 1980s when the virus's genome was sequenced; it was subsequently demonstrated that the predicted protein was indeed expressed from the gene, although its role in promoting virulence and infectivity was initially underappreciated. Following the elucidation of its role as a suppressor of RNA silencing, p19 has also been used as a tool in molecular biology research on RNA silencing, RNA interference, and related processes. [4] [6]

Related Research Articles

Interferon Signaling proteins released by host cells in response to the presence of pathogens

Interferons are a group of signaling proteins made and released by host cells in response to the presence of several viruses. In a typical scenario, a virus-infected cell will release interferons causing nearby cells to heighten their anti-viral defenses.

Gene silencing is the regulation of gene expression in a cell to prevent the expression of a certain gene. Gene silencing can occur during either transcription or translation and is often used in research. In particular, methods used to silence genes are being increasingly used to produce therapeutics to combat cancer and other diseases, such as infectious diseases and neurodegenerative disorders.

Small interfering RNA

Small interfering RNA (siRNA), sometimes known as short interfering RNA or silencing RNA, is a class of double-stranded RNA non-coding RNA molecules, typically 20-27 base pairs in length, similar to miRNA, and operating within the RNA interference (RNAi) pathway. It interferes with the expression of specific genes with complementary nucleotide sequences by degrading mRNA after transcription, preventing translation.

Dicer Enzyme that cleaves double-stranded RNA (dsRNA) into short dsRNA fragments

Dicer, also known as endoribonuclease Dicer or helicase with RNase motif, is an enzyme that in humans is encoded by the DICER1 gene. Being part of the RNase III family, Dicer cleaves double-stranded RNA (dsRNA) and pre-microRNA (pre-miRNA) into short double-stranded RNA fragments called small interfering RNA and microRNA, respectively. These fragments are approximately 20-25 base pairs long with a two-base overhang on the 3' ends. Dicer facilitates the activation of the RNA-induced silencing complex (RISC), which is essential for RNA interference. RISC has a catalytic component Argonaute, which is an endonuclease capable of degrading messenger RNA (mRNA).

Plant virus Virus that affects plants

Plant viruses are viruses that affect plants. Like all other viruses, plant viruses are obligate intracellular parasites that do not have the molecular machinery to replicate without a host. Plant viruses can be pathogenic to higher plants.

<i>Tombusviridae</i> Family of viruses

Tombusviridae is a family of single-stranded positive sense RNA plant viruses. There are three subfamilies, 17 genera, and 95 species in this family. The name is derived from Tomato bushy stunt virus (TBSV).

<i>Tomato bushy stunt virus</i> Species of virus

Tomato bushy stunt virus (TBSV) is a virus of the tombusvirus family. It was first reported in tomatoes in 1935 and primarily affects vegetable crops, though it is not generally considered an economically significant plant pathogen. Depending upon the host, TBSV causes stunting of growth, leaf mottling, and deformed or absent fruit. The virus is likely to be soil-borne in the natural setting, but can also transmitted mechanically, for example through contaminated cutting tools. TBSV has been used as a model system in virology research on the life cycle of plant viruses, particularly in experimental infections of the model host plant Nicotiana benthamiana.

The NS1 influenza protein (NS1) is a viral nonstructural protein encoded by the NS gene segments of type A, B and C influenza viruses. Also encoded by this segment is the nuclear export protein (NEP), formally referred to as NS2 protein, which mediates the export of influenza virus ribonucleoprotein (RNP) complexes from the nucleus, where they are assembled.

Protein kinase R

Protein kinase RNA-activated also known as protein kinase R (PKR), interferon-induced, double-stranded RNA-activated protein kinase, or eukaryotic translation initiation factor 2-alpha kinase 2 (EIF2AK2) is an enzyme that in humans is encoded by the EIF2AK2 gene.

Agroinfiltration

Agroinfiltration is a method used in plant biology and especially lately in plant biotechnology to induce transient expression of genes in a plant, or isolated leaves from a plant, or even in cultures of plant cells, in order to produce a desired protein. In the method, a suspension of Agrobacterium tumefaciens is introduced into a plant leaf by direct injection or by vacuum infiltration, or brought into association with plant cells immobilised on a porous support, whereafter the bacteria transfer the desired gene into the plant cells via transfer of T-DNA. The main benefit of agroinfiltration when compared to the more traditional plant transformation is speed and convenience, although yields of the recombinant protein are generally also higher and more consistent.

Argonaute Protein that plays a role in RNA silencing process

The Argonaute protein family plays a central role in RNA silencing processes, as essential components of the RNA-induced silencing complex (RISC). RISC is responsible for the gene silencing phenomenon known as RNA interference (RNAi). Argonaute proteins bind different classes of small non-coding RNAs, including microRNAs (miRNAs), small interfering RNAs (siRNAs) and Piwi-interacting RNAs (piRNAs). Small RNAs guide Argonaute proteins to their specific targets through sequence complementarity, which then leads to mRNA cleavage or translation inhibition.

RNA-dependent RNA polymerase

RNA-dependent RNA polymerase ( RdRp) or RNA replicase is an enzyme that catalyzes the replication of RNA from an RNA template. Specifically, it catalyzes synthesis of the RNA strand complementary to a given RNA template. This is in contrast to typical DNA-dependent RNA polymerases, which all organisms use to catalyze the transcription of RNA from a DNA template.

RNA silencing or RNA interference refers to a family of gene silencing effects by which gene expression is negatively regulated by non-coding RNAs such as microRNAs. RNA silencing may also be defined as sequence-specific regulation of gene expression triggered by double-stranded RNA (dsRNA). RNA silencing mechanisms are highly conserved in most eukaryotes. The most common and well-studied example is RNA interference (RNAi), in which endogenously expressed microRNA (miRNA) or exogenously derived small interfering RNA (siRNA) induces the degradation of complementary messenger RNA. Other classes of small RNA have been identified, including piwi-interacting RNA (piRNA) and its subspecies repeat associated small interfering RNA (rasiRNA).

Sir David Charles Baulcombe is a British plant scientist and geneticist. As of 2017 he is a Royal Society Research Professor and Regius Professor of Botany in the Department of Plant Sciences at the University of Cambridge.

Rice hoja blanca tenuivirus (RHBV), meaning "white leaf rice virus", is a plant virus in the family Phenuiviridae. RHBV causes Hoja blanca disease (HBD), which affects the leaves of the rice plant Oryza sativa, stunting the growth of the plant or killing it altogether. RHBV is carried by an insect vector, Tagosodes orizicolus, a type of planthopper. The virus is found in South America, Mexico, throughout Central America, the Caribbean region, and the southern United States. In South America, the disease is endemic to Colombia, Venezuela, Ecuador, Peru, Suriname, French Guiana and Guyana.

ADAR

Double-stranded RNA-specific adenosine deaminase is an enzyme that in humans is encoded by the ADAR gene.

RNA polymerase IV is an enzyme that synthesizes small interfering RNA (siRNA) in plants, which silence gene expression. RNAP IV belongs to a family of enzymes that catalyze the process of transcription known as RNA Polymerases, which synthesize RNA from DNA templates. Discovered via phylogenetic studies of land plants, genes of RNAP IV are thought to have resulted from multistep evolution processes that occurred in RNA Polymerase II phylogenies. Such an evolutionary pathway is supported by the fact that RNAP IV is composed of 12 protein subunits that are either similar or identical to RNA polymerase II, and is specific to plant genomes. Via its synthesis of siRNA, RNAP IV is involved in regulation of heterochromatin formation in a process known as RNA directed DNA Methylation (RdDM).

Antiviral protein

Antiviral proteins are proteins that are induced by human or animal cells to interfere with viral replication. These proteins are isolated to inhibit the virus from replicating in a host's cells and stop it from spreading to other cells. The Pokeweed antiviral protein and the Zinc-Finger antiviral protein are two major antiviral proteins that have undergone several tests for viruses, including HIV and influenza.

RNA interference Biological process of gene regulation

RNA interference (RNAi) is a biological process in which RNA molecules are involved in sequence-specific suppression of gene expression by double-stranded RNA, through translation or transcriptional repression. Historically, RNAi was known by other names, including co-suppression, post-transcriptional gene silencing (PTGS), and quelling. The detailed study of each of these seemingly different processes elucidated that the identity of these phenomena were all actually RNAi. Andrew Fire and Craig C. Mello shared the 2006 Nobel Prize in Physiology or Medicine for their work on RNA interference in the nematode worm Caenorhabditis elegans, which they published in 1998. Since the discovery of RNAi and its regulatory potentials, it has become evident that RNAi has immense potential in suppression of desired genes. RNAi is now known as precise, efficient, stable and better than antisense therapy for gene suppression. Antisense RNA produced intracellularly by an expression vector may be developed and find utility as novel therapeutic agents.

RNA therapeutics are a class of medications based on ribonucleic acid (RNA).

References

  1. 1 2 3 4 5 Ye K, Malinina L, Patel DJ (December 2003). "Recognition of small interfering RNA by a viral suppressor of RNA silencing". Nature. 426 (6968): 874–8. Bibcode:2003Natur.426..874Y. doi:10.1038/nature02213. PMC   4694583 . PMID   14661029.
  2. 1 2 3 Vargason JM, Szittya G, Burgyán J, Hall TM (December 2003). "Size selective recognition of siRNA by an RNA silencing suppressor". Cell. 115 (7): 799–811. doi:10.1016/S0092-8674(03)00984-X. PMID   14697199. S2CID   12993441.
  3. 1 2 Lakatos L, Szittya G, Silhavy D, Burgyán J (February 2004). "Molecular mechanism of RNA silencing suppression mediated by p19 protein of tombusviruses". The EMBO Journal. 23 (4): 876–84. doi:10.1038/sj.emboj.7600096. PMC   381004 . PMID   14976549.
  4. 1 2 3 Scholthof HB (May 2006). "The Tombusvirus-encoded P19: from irrelevance to elegance". Nature Reviews. Microbiology. 4 (5): 405–11. doi:10.1038/nrmicro1395. PMID   16518419. S2CID   30361458.
  5. 1 2 3 Pumplin N, Voinnet O (November 2013). "RNA silencing suppression by plant pathogens: defence, counter-defence and counter-counter-defence". Nature Reviews. Microbiology. 11 (11): 745–60. doi:10.1038/nrmicro3120. PMID   24129510. S2CID   205498691.
  6. 1 2 Danielson DC, Pezacki JP (April 2013). "Studying the RNA silencing pathway with the p19 protein". FEBS Letters. 587 (8): 1198–205. doi: 10.1016/j.febslet.2013.01.036 . PMID   23376479.
  7. 1 2 Ding SW, Voinnet O (August 2007). "Antiviral immunity directed by small RNAs". Cell. 130 (3): 413–26. doi:10.1016/j.cell.2007.07.039. PMC   2703654 . PMID   17693253.
  8. ↑ Várallyay E, Válóczi A, Agyi A, Burgyán J, Havelda Z (October 2010). "Plant virus-mediated induction of miR168 is associated with repression of ARGONAUTE1 accumulation". The EMBO Journal. 29 (20): 3507–19. doi:10.1038/emboj.2010.215. PMC   2964164 . PMID   20823831.
  9. ↑ Várallyay É, Oláh E, Havelda Z (January 2014). "Independent parallel functions of p19 plant viral suppressor of RNA silencing required for effective suppressor activity". Nucleic Acids Research. 42 (1): 599–608. doi:10.1093/nar/gkt846. PMC   3874164 . PMID   24062160.
  10. ↑ Scholthof HB, Scholthof KB, Kikkert M, Jackson AO (November 1995). "Tomato bushy stunt virus spread is regulated by two nested genes that function in cell-to-cell movement and host-dependent systemic invasion". Virology. 213 (2): 425–38. doi: 10.1006/viro.1995.0015 . PMID   7491767.
  11. 1 2 3 Rancurel C, Khosravi M, Dunker AK, Romero PR, Karlin D (October 2009). "Overlapping genes produce proteins with unusual sequence properties and offer insight into de novo protein creation". Journal of Virology. 83 (20): 10719–36. doi:10.1128/JVI.00595-09. PMC   2753099 . PMID   19640978.
  12. ↑ Sabath N, Wagner A, Karlin D (December 2012). "Evolution of viral proteins originated de novo by overprinting". Molecular Biology and Evolution. 29 (12): 3767–80. doi:10.1093/molbev/mss179. PMC   3494269 . PMID   22821011.
  13. ↑ Lakatos L, Csorba T, Pantaleo V, Chapman EJ, Carrington JC, Liu YP, Dolja VV, Calvino LF, López-Moya JJ, Burgyán J (June 2006). "Small RNA binding is a common strategy to suppress RNA silencing by several viral suppressors". The EMBO Journal. 25 (12): 2768–80. doi:10.1038/sj.emboj.7601164. PMC   1500863 . PMID   16724105.
  14. ↑ Voinnet O (March 2005). "Induction and suppression of RNA silencing: insights from viral infections". Nature Reviews. Genetics. 6 (3): 206–20. doi:10.1038/nrg1555. PMID   15703763. S2CID   26351712.

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