Virus resistance mechanisms in plants Three basic ways of Resistance  Healthy planting material and cultural practices  Host plant resistanceVectors, Viruses, Cross protection  Transgenic approach ▪ Pathogen derived genes and other sources Stages of viral infection cycle Co-ordinated expression of viral and plant genome essential for virus Entry, un coating of NA, Virus infection Epidermal translation of viral proteins Replication, cells Asssembly of Replication progeny virus Cell-to-cell movement Mesophyll cells Phloem Systemic movement Other host plants Bundle sheath cells Plant-to-plant movement Phloem Parenchyma Companion cells Prevention Of Viral Uncoating  If a virus is prevented from uncoating, its DNA will not be exposed to cellular replication machinery and the virus will not be able to reproduce shown in tobacco mosaic virus, TMV  Inoculation of a tobacco plant with the treated virus (Ph 8.0 for a brief period of time, which if the virus is not treated,  removes about sixty coat protein subunits ) causes an infection, but no infection results in transgenic coat proten mediated resistant plant This lack of infection is due to the viral mRNA not being translated Two theories exist to explain why this may be so Theory One: TMV has a specific uncoating receptor which is blocked by coat protein High coat protein concentrations would block the uncoating receptor  which also controls encapsidation in a normal replication cycle  In a transgenic plant the large coat protein concentration blocks the uncoating receptor of the therefore preventing infection Theory Two :TMV uncoats when it enters a plant cell due to local physiological changes If a transgenic plant has a high coat protein concentration, it will favour viral coating  This would effectively prevent uncoating of the invading virus viral RNA, CELL-TO-CELL MOVEMENT OF COWPEA MOSAIC VIRUS Movement of virus particle through modified plasmodesmata Viral movement protein Disassembly of viral movement complex Assembly of viral movement complex coat protein Virus disassembly Virus assembly virus particles viral RNA Viral RNA replication, translation, etc Plasmadesmata = junction between plant cells More cell-to-cell movement RESPONSES OF PLANTS TO VIRUS INOCULATION  Immune - no replication at all  Complete resistance in a plant to virus infection is referred to as immunity  The immunity is usually manifested in preventing virus replication  If immunity occurs against all biotypes of a pathogen and in all cultivars or accessions of a particular plant species the situation is re ferred to as non-host resistance  For viruses, this is a largely an unexplored Area unlike in fungi etc  Nonhost resistance  All genotypes within plant species show resistance or fail to be infected by a particular virus Unlike in fungus it is not understoo d against virus  One of the first innate immune responses all plant viruses encounter when Ai) and invading a host consists of antiviral RNA silencing (RN post-transcriptional gene silencing (PTSG)]  RNA silencing is a host response triggered by double stranded (ds)RNA  These molecules thus act as a MAMP/PAMP and in which RNAi can be regarded as PTI  The main difference with pathogens such as fungi and bacteria is that recognition of viral MAMPs/PAMPs occur intra-cellular ly  One of the most common strategies plant viruses use to counteract RNAi is to encode RNA silencing suppressors (RSS), viral proteins that i nterfere with a specific part of the RNAi pathway and thereby reduce its effectiveness  The majority of plant virus RSS proteins exert this activity through binding of small hereby prevent their  uploading into RISC and Dicer-cleavage, respectively In recent years some RSS have also been discovered to inhibit the RNAi pathway in ke AGO1, the core interfering (si)RNAs, or sometimes (also) long dsRNA, and t component of RISC during the antiviral RNAi response other ways, e.g., by binding directly to key-enzyme proteins li RESISTANCE AGAINST VIRUS VECTORS Genes that deter infestations with the vectors and/or block virus transmission have the potential to provide additional scope for genetic resistance Only a few genes conferring resistance to potential virus vectors have been characterized These fall within the NBS-LRR group of plant R genes, underlining the key role of this type The tomato Mi-1 confers resistance to the potato aphid (Macrosiphum euphorbiae), of gene not only against pathogens but also to pests whitefly (Bemisia tabaci ) and the root-knot nematodes (Meloidogyne spp.), The melon Vat gene controls plant colonization by the aphid Aphis gossypii and the transmission of non-persistent viruses Nr, conferring resistance in lettuce against the aphid Nasonovia ribisnigri, shows functional (Acyrthosiphon kondoi ) similarity to Mi-1, and resistance to the blue green aphid Proteases Viruses need express several proteins from a single mRNA –a polyprotein Then processed to produce functional proteins by virus encoded proteases Block Proteases Proteases have two domains Location Domain : identify cutting site Catalytic domain: Actual cutting Strategy a Only catalytic is modified : So block the substrate trap the viral protein b Only over produce location domain in the plant: lock the cutting site of the multiprotein c Produce protoxin which can be cleaved by viral protease to produce toxin Use of Antibodies against virus Active antibodies raised against Antichoke mottled crinkle virus F8 antibodies have highest affinity for the CP in both dissociated and polymerized forms Attach to highly conserved regions in coat protein Similarly antibodies against RNA also Transferred to N benthamiana and results promising Generation of Single chain variable fragment (ScFv) antibodies, Synthestic ScFV libraries may improve the ability of this concept RNA-MEDIATED RESISTANCE This post-transcriptional gene silencing (PTGS) process, occurring in plants is also known as RNA interference (RNAi) or RNA silencing Expression levels of transgene-encoded viral proteins often did not correlate with the level of virus resistance Plants expressing the lowest or even undetectable levels of protein often displayed the highest Subsequent expression of untranslatable transgenes formally demonstrated the involvement of resistance transgenic RNA in resistance Post-transcriptional down regulation of endogenous genes by transgenes of identical sequence Si RNA concept led to constructing inverted repeat (IR) transgenes from which long double-stranded (ds) RNA precursors of siRNAs were generated Utilization of such IR transgene constructs yielded a marked increase in the efficiency of this approach Single-stranded sense or antisense approaches yielded resistance in 5–20% of the transgenic plants IR transgenes that produce dsRNAs proved to yield up to 90% of all transgenic plants resistant to the homologous virus Colud be due to the dsRNAs being fed into a later step in the silencing pathway Drawbacks of RNA-mediated resistance is that it is ineffective against transgene by more than 10% viruses whose sequence differs from that of the RNA-mediated resistance against DNA viruses DNA viruses like caulimoviruses and geminiviruses are also targets of RNA silencing Sense and antisense RNA in transgenic plants have been employed successfully against Tomato golden mosaic virus(TGMV),TYLCSV and TYLCV confirming the suggestion that RNA silencing can be harnessed for antiviral defence Resistant to TYLCV by targeting the CP gene with an IR construct However, these plants often showed significant delays in symptom development, particularly at low inoculum dosage In sharp contrast to the situation with RNA viruses, completely immune lines were not observed The success of the strategy depends on the relevance of the targeted gene product in the systemic spread of the virus Decoys (Traps) Strategy is Replication control sequences Expression Control sequences Satellite RNA Some strains of CMV encapsidate satellite RNA (sat RNA) in addition to the tripartite single-stranded RNA genome CMV sat RNA depends on its helper virus (HV) CMV for replication, movement within the plant, encapsidation and transmission The presence of sat-RNA modulates the symptoms induced by the HV and often depresses HV accumulation in different host species Transgenic tobacco plants expressing multiple or partial copies of CMV sat-RNA showed attenuated symptoms when challenged with CMV28 In addition, tobacco plants transformed with anti-sense sat-RNA also showed delayed symptom development with the cognate virus29 Tobacco Plants with DNA copies of CMV satellites Resistant to CMV and TAV In case of CMV viral multiplication was reduced while in TAV it is only symptoms In some cases it has increased the symptoms Defective-interfering viral nucleic acids DI’s ( defective Interfering Sequences ) DI’s are although rare found in + sense strand RNA viruses DI’s are sequences which are related to parent virus but require parent virus for multiplication In the presence of DI RNA there is reduced accumulation of Viral RNA DI interfere with the replication of the genomic components Expression of DI’s : Cymbidium ring spot tombusvirus in N benthamiana reduce symptoms Transgenics with non-pathogen derived resistance The strategy utilizing genes derived from either the host plant or any other The other nonpathogen derived strategies are the utilization of plant disease resistance genes, the ribosome-inactivating proteins, plant proteinase inhibitors, human interferon-like systems, antiviral antibodies expressed in plants, systemic acquired resistance and secondary metabolite engineering non-pathogenic source Resistance due to the expression of non-pathogen - derived antiviral agents DNA binding proteins •Zinc finger proteins are high affinity for the “Rep–specific direct repeats “ in the “virion-ori “ of different geminiviruses •Block the binding of “Rep” to “virion- ori” of geminivirus •Transgenically expressed artificially designed Zinc finger protein provide resistant against geminiviruses Resistance due to the expression of non-pathogen - derived antiviral agents GroEL – mediated resistance •Chaparon •Homologue of GroEL produced by endosymbiotic bacteria from B tabaci •Higher affinity to TYLCV coat protein Eg B tabaci GroEL gene expressed in transgenic tomatoes under phloem specific promoter, protected the plants from the TYLCV infection Resistance due to the expression of non-pathogen - derived antiviral agents Peptide aptamers •Short recombinant protein, ~ 20 amino acid length •Strongly binds with target protein and destructs the function •Transgenic N benthamiana - protected from Tomato spotted wilt Virus ... and/or companion cells Host Plant Resistance  Resistance Mechanisms to Plant Viruses   Dominant resistance to plant viruses Recessive resistance to plant viruses Resistance Mechanisms Virulence... result in recessively inherited resistance  Recessive resistances are more frequent for potyviruses than for viruses of other families RECESSIVE RESISTANCE   Viruses are dependent on the host... defense that involves resistance genes (HOST PLANT RESISTANCE) Most of these are triggered by and confer resistance to a specific virus only Some act against several (related) viruses The major class