Cell-to-cell transport of plant viruses occurs through plasmodesmata, the channels interconnecting neighboring cells in plant tissues, and requires dedicated virus-encoded movement proteins (MPs) [1Lucas WJ. Plant viral movement proteins: Agents for cell-to-cell trafficking of viral genomes Virology 2006; 344: 169-84.-5Lucas WJ, Ham BK, Kim JY. Plasmodesmata - bridging the gap between neighboring plant cells Trends Cell Biol 2009; 19: 495-503.]. In positive-stranded RNA viruses the number of encoded MPs can vary between distant virus groups, and they are often unrelated in their primary structure [1Lucas WJ. Plant viral movement proteins: Agents for cell-to-cell trafficking of viral genomes Virology 2006; 344: 169-84.]. Nevertheless, viral MPs which represent dissimilar transport systems share three characteristic features. First, the RNA-binding activity is believed to be responsible for formation of complexes with viral genomic RNAs (viral ribonucleoproteins, vRNPs) which represent the transport form of viral genomes capable of cell-to-cell movement. Second, the ability of MPs to interact with cell membranes, in particular with the endoplasmic reticulum (ER), may direct MPs to specific pathway of intracellular transport to plasmodesmata. Third, MPs are able to accumulate in plasmodesmata or plasmodesmata-associated sites and to increase the size exclusion limit of plasmodesmata pores for efficient passage of macromolecules to neighboring cells [3Taliansky M, Torrance L, Kalinina NO. Role of plant virus movement proteins Methods Mol Biol 2008; 451: 33-54.-5Lucas WJ, Ham BK, Kim JY. Plasmodesmata - bridging the gap between neighboring plant cells Trends Cell Biol 2009; 19: 495-503.].

A number of plant virus genera require for their cell-to-cell movement a ‘triple gene block’ (TGB), conserved gene module encoding three MPs [6Morozov SY, Dolja VV, Atabekov JG. Probable reassortment of genomic elements among elongated RNA-containing plant viruses J Mol Evol 1989; 29: 52-62.]. The TGB position relative to other genes vary in genomes of different viruses, however the order of genes within the TGB and the properties of the encoded proteins are conserved [7Morozov SY, Solovyev AG. Triple gene block: modular design of a multifunctional machine for plant virus movement J Gen Virol 2003; 84: 1351-66., 8Verchot-Lubicz J, Torrance L, Solovyev AG, Morozov SY, Jackson AO, Gilmer D. Varied movement strategies employed by triple gene block-encoding viruses Mol Plant-Microbe Interact 2010; 23: 1231-47.]. Based on differences in the structure of TGB proteins and the requirement of viral capsid protein for cell-to-cell movement two types of TGB are distinguished: ‘hordei-like’ TGB in rod-shaped viruses (family Virgaviridae and genus Benyvirus) and ‘potex-like’ TGB in filamentous viruses (family Flexiviridae) [7Morozov SY, Solovyev AG. Triple gene block: modular design of a multifunctional machine for plant virus movement J Gen Virol 2003; 84: 1351-66., 8Verchot-Lubicz J, Torrance L, Solovyev AG, Morozov SY, Jackson AO, Gilmer D. Varied movement strategies employed by triple gene block-encoding viruses Mol Plant-Microbe Interact 2010; 23: 1231-47.].

Studies of hordei-like TGB carried out in the recent years and employed as models Barley stripe mosaic virus (BSMV, genus Hordeivirus), Poa semilatent virus (genus Hordeivirus) and Potato mop-top virus (PMTV, genus Pomovirus) revealed characteristic biochemical properties and functions of individual TGB proteins, as well as their physical and functional interactions involved in virus cell-to-cell movement [8Verchot-Lubicz J, Torrance L, Solovyev AG, Morozov SY, Jackson AO, Gilmer D. Varied movement strategies employed by triple gene block-encoding viruses Mol Plant-Microbe Interact 2010; 23: 1231-47., 9Jackson AO, Lim HS, Bragg J, Ganesan U, Lee MY. Hordeivirus replication, movement, and pathogenesis Annu Rev Phytopathol 2009; 47: 385-422.]. TGBp1, the largest of TGB proteins with molecular mass of 51 to 63 kDa in hordei-like TGBs, consists of three domains, the N-terminal domain, internal domain and the C-terminal NTPase/helicase domain [10Makarov VV, Rybakova EN, Efimov AE, et al. Domain organization of the N-terminal portion of hordeivirus movement protein TGBp1 J Gen Virol 2009; 90: 3022-2.]. In Poa semilatent virus, the N-terminal domain represents an unfolded protein region which includes two positively charged sequence motifs involved in virus long-distance movement via the phloem, whereas the internal domain is a small structured domain able to interact with both the N-terminal domain and the NTPase/helicase domain [10Makarov VV, Rybakova EN, Efimov AE, et al. Domain organization of the N-terminal portion of hordeivirus movement protein TGBp1 J Gen Virol 2009; 90: 3022-2., 11Kalinina NO, Rakitina DA, Yelina NE, et al. RNA-binding properties of the 63 kDa protein encoded by the triple gene block of poa semilatent hordeivirus J Gen Virol 2001; 82: 2569-78.]. The latter domain has the NTPase and helicase activities and is capable of self-interactions [10Makarov VV, Rybakova EN, Efimov AE, et al. Domain organization of the N-terminal portion of hordeivirus movement protein TGBp1 J Gen Virol 2009; 90: 3022-2.-14Leshchiner AD, Solovyev AG, Morozov SY, Kalinina NO. A minimal region in the NTPase/helicase domain of the TGBp1 plant virus movement protien is responsible for ATPase activity and cooperative RNA binding J Gen Virol 2006; 87: 3087-95.]. Additionally, all three domains of the hordeiviral TGBp1 bind RNA exhibiting either a cooperative (the internal and the NTPase/helicase domains) or non-cooperative (the N-terminal domain) binding [10Makarov VV, Rybakova EN, Efimov AE, et al. Domain organization of the N-terminal portion of hordeivirus movement protein TGBp1 J Gen Virol 2009; 90: 3022-2., 11Kalinina NO, Rakitina DA, Yelina NE, et al. RNA-binding properties of the 63 kDa protein encoded by the triple gene block of poa semilatent hordeivirus J Gen Virol 2001; 82: 2569-78., 13Donald RG, Lawrence DM, Jackson AO. The barley stripe mosaic virus 58-kilodalton beta(b) protein is a multifunctional RNA binding protein J Virol 1997; 71: 1538-46.]. These RNA binding activities are responsible for interaction with viral RNAs and the formation of vRNPs; such RNPs were isolated from BSMV-infected plants and were shown to contain BSMV positive-stranded RNAs and TGBp1 but not other viral or non-viral components [15Lim H-S, Bragg JN, Ganesan U, et al. Triple gene block protein interactions involved in movement of Barley stripe mosaic virus J Virol 2008; 82: 4991-5006.]. The presence of multiple binding sites in TGBp1 and the ability of internal domain for interactions with other protein domains, as well as the proposed ability of the N-terminal domain to conditionally adopt a structured conformation, provide the basis for structural remodeling of vRNPs that would be necessary for different stages of virus movement in plant [10Makarov VV, Rybakova EN, Efimov AE, et al. Domain organization of the N-terminal portion of hordeivirus movement protein TGBp1 J Gen Virol 2009; 90: 3022-2.]. TGBp2 and TGBp3 are small integral membrane proteins required for delivery of hordei-like TGBp1 to plasmodesmata [7Morozov SY, Solovyev AG. Triple gene block: modular design of a multifunctional machine for plant virus movement J Gen Virol 2003; 84: 1351-66., 8Verchot-Lubicz J, Torrance L, Solovyev AG, Morozov SY, Jackson AO, Gilmer D. Varied movement strategies employed by triple gene block-encoding viruses Mol Plant-Microbe Interact 2010; 23: 1231-47.]. While TGBp2, when expressed in the absence of TGBp3, is localized to the cortical endoplasmic reticulum (ER) or numerous ER-associated vesicular bodies, TGBp3 localizes to plasmodesmata or plasmodesmata-associated membrane compartments called peripheral membrane bodies (PMBs) and, moreover, is able to target TGBp2 from the ER structures to plasmodesmata and PMBs [7Morozov SY, Solovyev AG. Triple gene block: modular design of a multifunctional machine for plant virus movement J Gen Virol 2003; 84: 1351-66.-9Jackson AO, Lim HS, Bragg J, Ganesan U, Lee MY. Hordeivirus replication, movement, and pathogenesis Annu Rev Phytopathol 2009; 47: 385-422.]. Furthermore, TGBp2 and TGBp3 direct TGBp1 to PMBs and plasmodesmata, and in the case of BSMV this targeting could involve direct interactions between TGBp1 and TGBp3 [16Zamyatnin AA Jr, Solovyev AG, Savenkov EI, et al. Transient coexpression of individual genes encoded by the triple gene block of potato mop-top virus reveals requirements for TGBp1 trafficking Mol Plant-Microbe Interact 2004; 17: 921-30., 17Lim H-S, Bragg JN, Ganesan U, et al. Subcellular localization of the barley stripe mosaic virus triple gene block proteins J Virol 2009; 83: 9432-48.]. Recent evidences suggest that TGBp3 contains signals of plasmodesmata targeting [18Schepetilnikov MV, Solovyev AG, Gorshkova EN, et al. Intracellular targeting of a hordeiviral membrane-spanning movement protein: sequence requirements and involvement of an unconventional mechanism J Virol 2008; 82: 1284-93., 19Tilsner J, Cowan GH, Roberts AG, et al. Plasmodesmal targeting and intercellular movement of potato mop-top pomovirus is mediated by a membrane anchored tyrosine-based motif on the lumenal side of the endoplasmic reticulum and the C-terminal transmembrane domain in the TGB3 movement protein Virology 2010; 402: 41-51.] and that TGBp3 serves therefore as a ‘driving force’ for the targeting of TGBp2, TGBp1, and most probably TGBp1-containing vRNPs to plasmodesmata [8Verchot-Lubicz J, Torrance L, Solovyev AG, Morozov SY, Jackson AO, Gilmer D. Varied movement strategies employed by triple gene block-encoding viruses Mol Plant-Microbe Interact 2010; 23: 1231-47.].

Previously we analyzed PMTV TGB functions using an experimental system in which intra- and intercellular transport of TGB proteins was reconstructed using individually cloned PMTV TGB genes and a particle bombardment-based delivery of recombinant plasmids into plant cells [16Zamyatnin AA Jr, Solovyev AG, Savenkov EI, et al. Transient coexpression of individual genes encoded by the triple gene block of potato mop-top virus reveals requirements for TGBp1 trafficking Mol Plant-Microbe Interact 2004; 17: 921-30.]. Specifically, this experimental system includes two expression vectors which carry, under the control of the cauliflower mosaic virus 35S promoter, either the PMTV TGBp1 gene fused to the green fluorescent protein (GFP) gene, or the cloned region of PMTV genome comprising overlapping genes of TGBp2 and TGBp3. The latter construct mimics the natural expression of TGBp2 and TGBp3 from a single subgenomic RNA [20Zhou H, Jackson AO. Expression of the barley stripe mosaic virus RNA beta ‘triple gene block’ Virology 1996; 216: 367-79.]. After bombardment of plant leaves with metal particles carrying the GFP-TGBp1 expression vector, the fusion protein expressed in individual epidermal cells was distributed uniformly in the cytoplasm and also found in the nucleus. However, when leaves were bombarded by a mixture of GFP-TGBp1 and TGBp2/TGBp3 expression vectors, GFP-TGBp1 was directed to large PMBs and to cell wall-embedded punctate bodies supposed to correspond to plasmodesmata and, moreover, was transported into cells immediately adjacent to the bombarded cell showing that PMTV TGBp2 and TGBp3 can mediate transport of GFP-TGBp1 to and through plasmodesmata in the absence of other virus-encoded components [16Zamyatnin AA Jr, Solovyev AG, Savenkov EI, et al. Transient coexpression of individual genes encoded by the triple gene block of potato mop-top virus reveals requirements for TGBp1 trafficking Mol Plant-Microbe Interact 2004; 17: 921-30.]. Further experiments revealed that specific interactions between TGBp1 and TGBp2/TGBp3 are required for TGBp1 delivery to plasmodesmata and translocation to adjacent cells as well as allowed us to carry out a preliminary mapping of protein region involved in interaction between TGB proteins [16Zamyatnin AA Jr, Solovyev AG, Savenkov EI, et al. Transient coexpression of individual genes encoded by the triple gene block of potato mop-top virus reveals requirements for TGBp1 trafficking Mol Plant-Microbe Interact 2004; 17: 921-30.]. Therefore, the experimental system based on co-expression of TGB genes by use of particle bombardment proved to be a useful tool for studies of TGB-mediated transport.

In this paper we report elaboration of an experimental system based on agrobacteria-mediated transient expression of three PMTV TGB proteins from a single binary vector providing controlled synthesis levels for TGBp1, TGBp2 and TGBp3. Using this system we demonstrate a transient association of GFP-tagged TGBp1 with microtubules and microtubule-associated bodies that preceded TGBp1 targeting to cell wall-embedded structures which are proved to be plasmodesmata. We also show that disruption of cell microtubules with colchicine prevents TGBp1 targeting to plasmodesmata, and TGBp1 mutations which block interactions with microtubules also impede its plasmodesmata targeting.

In this paper we report co-localization of GFP-fused PMTV TGBp1 with MTs. Thus, TGBp1 can be added to a list of plant virus MPs capable of interaction, direct or indirect, with MTs. The most extensively studied MP encoded by TMV is among these proteins. During infection, TMV MP interacts with endoplasmic reticulum (ER) membranes and MTs, as well as localizes to plasmodesmata and induces an increase of their size exclusion limit [3Taliansky M, Torrance L, Kalinina NO. Role of plant virus movement proteins Methods Mol Biol 2008; 451: 33-54., 4Epel BL. Plant viruses spread by diffusion on ER-associated movement-protein-rafts through plasmodesmata gated by viral induced host beta-1, 3-glucanases Semin Cell Dev Biol 2009; 20: 1074-81., 35Heinlein M, Epel BL. Macromolecular transport and signaling through plasmodesmata Int Rev Cytol 2004; 235: 93-164.]. TMV MP also has the capacity to bind single-stranded nucleic acids in vitro, and the movement of viral RNA genome likely occurs in the form of vRNP [3Taliansky M, Torrance L, Kalinina NO. Role of plant virus movement proteins Methods Mol Biol 2008; 451: 33-54.]. Thus TMV MP shares with PMTV TGBp1 at least two activities involved in formation of vRNPs and interaction with cortical MTs [8Verchot-Lubicz J, Torrance L, Solovyev AG, Morozov SY, Jackson AO, Gilmer D. Varied movement strategies employed by triple gene block-encoding viruses Mol Plant-Microbe Interact 2010; 23: 1231-47., 35Heinlein M, Epel BL. Macromolecular transport and signaling through plasmodesmata Int Rev Cytol 2004; 235: 93-164.]. TMV MP sequences required for interaction with MTs are identified by mutation analyses [36Kahn TW, Lapidot M, Heinlein M, et al. Domains of the TMV movement protein involved in subcellular localization Plant J 1998; 15: 15-25., 37Boyko V, van der Laak J, Ferralli J, Suslova E, Kwon MO, Heinlein M. Cellular targets of functional and dysfunctional mutants of tobacco mosaic virus movement protein fused to green fluorescent protein J Virol 2000; 74: 11339-46.]. Moreover, a region within TMV MP is revealed that shares sequence similarity to tubulins, and single amino acid substitutions within this so-called ‘tubulin mimicry domain’ are sufficient to confer temperature-sensitive loss of MT association pointing to the possibility that TMV MP is capable of making binding contacts equivalent to tubulin-tubulin interaction [38Boyko V, Ferralli J, Ashby J, Schellenbaum P, Heinlein M. Function of microtubules in intercellular transport of plant virus RNA Nat Cell Biol 2000; 2: 826-32.]. These data, together with biochemical evidences obtained in vitro [39McLean B, Zupan J, Zambryski P. Tobacco mosaic movement protein associates with the cytoskeleton in tobacco cells Plant Cell 1995; 7: 2101-14.-41Sambade A, Brandner K, Hofmann C, Seemanpillai M, Mutterer J, Heinlein M. Transport of TMV movement protein particles associated with the targeting of RNA to plasmodesmata Traffic 2008; 9: 2073-88.], indicate that TMV MP can interact with tubulin and MTs directly. Although we were unable to find a sequence similar to the ‘tubulin mimicry domain’ in PMTV TGBp1 (data not shown), deletion analysis suggest that the TGBp1 region encompassing residues 86 - 184 is involved in MT association. Assuming that the predicted subdomain organization of PMTV TGBp1 is very similar to that of hordeivirus TGBp1 [10Makarov VV, Rybakova EN, Efimov AE, et al. Domain organization of the N-terminal portion of hordeivirus movement protein TGBp1 J Gen Virol 2009; 90: 3022-2.], one can conclude that among three domains of the hordei-like TGBp1s the structured ID domain is responsible for the protein association with MTs. Taking into account our preliminary data indicating that PMTV TGBp1 can interact with tobacco tubulin A1 in vitro, we presume the direct physical TGBp1 interaction with MTs.

What is the possible significance of the targeting of PMTV TGBp1 to the MTs? Recent findings illuminated the role of MTs in the intracellular transport of TMV vRNPs. Using a TMV derivative encoding a temperature-sensitive TMV MP, small mobile MT-proximal particles representing MP-RNA complexes were revealed in cells at the leading front of infection [41Sambade A, Brandner K, Hofmann C, Seemanpillai M, Mutterer J, Heinlein M. Transport of TMV movement protein particles associated with the targeting of RNA to plasmodesmata Traffic 2008; 9: 2073-88., 42Boyko V, Hu Q, Seemanpillai M, Ashby J, Heinlein M. Validation of microtubule-associated tobacco mosaic virus RNA movement and involvement of microtubule-aligned particle trafficking Plant J 2007; 51: 589-603.]. Importantly, the TMV MP-containing particles formed upon MP transient expression in the absence of viral infection are functionally related to the particles observed in virus-infected cells; these particles contain RNA and are thought to be assembled on the MT network; their anchorage to MT sites and release can contribute to the intracellular transport of these specific RNPs to plasmodesmata [41Sambade A, Brandner K, Hofmann C, Seemanpillai M, Mutterer J, Heinlein M. Transport of TMV movement protein particles associated with the targeting of RNA to plasmodesmata Traffic 2008; 9: 2073-88.]. Similarly to TMV MP, PMTV TGBp1 was also found in granular bodies associated with MTs. However, in contrast to the TMV MP, we were unable to detect movement of TGBp1-containing bodies. Nevertheless, association of TGBp1 with MTs and its localization in MT-associated granular bodies can be necessary for the protein targeting to plasmodesmata in the presence of TGBp2 and TGBp3, since treatment with the microtubule-disrupting drug, colchicine, inhibits localization of TGBp1 to plasmodesmata. One possible function of TGBp1 association with MTs is that the MT network may provide a scaffold for assembly of TGBp1-containing granular bodies as an initial step in TGBp1 targeting to plasmodesmata by TGBp2 and TGBp3. It should be noted that the Cauliflower mosaic virus proteins P2, P3 and P6 also form bodies that localize to MTs and, moreover, require intact MTs for their efficient formation [43Harries PA, Palanichelvam K, Yu W, Schoelz JE, Nelson RS. The cauliflower mosaic virus protein P6 forms motile inclusions that traffic along actin microfilaments and stabilize microtubules Plant Physiol 2009; 149: 1005-6., 44Martinière A, Gargani D, Uzest M, Lautredou N, Blanc S, Drucker M. A role for plant microtubules in the formation of transmission-specific inclusion bodies of Cauliflower mosaic virus Plant J 2009; 58: 135-46.], lending support to the possibility that a similar mechanism may be utilized for the formation of MT-associated granular structures containing PMTV TGBp1.

Additionally, localization to MTs could facilitate TGBp1 degradation. The degradation of Potato leafroll virus MP, for example, was found to require intact MTs, and a similar model has been proposed for the MT-associated degradation of TMV MP [32Vogel F, Hofius D, Sonnewald U. Intracellular trafficking of Potato leafroll virus movement protein in transgenic Arabidopsis Traffic 2007; 8: 1205-4., 45Curin M, Ojangu EL, Trutnyeva K, Ilau B, Truve E, Waigmann E. MPB2C, a microtubule-associated plant factor, is required for microtubular accumulation of tobacco mosaic virus movement protein in plants Plant Physiol 2007; 143: 801-11.]. It was suggested that individual MP molecules or their small aggregates could traffic along MTs to stationary sites of the formation of aggresome-like structures [32Vogel F, Hofius D, Sonnewald U. Intracellular trafficking of Potato leafroll virus movement protein in transgenic Arabidopsis Traffic 2007; 8: 1205-4.]. Indeed, a similar model has been proposed for the formation of aggresomes, perinuclear structures formed in animal cells in response to protein overproduction and misfolding [46Garcia-Mata R, Gao YS, Sztul E. Hassles with taking out the garbage: aggravating aggresomes Traffic 2002; 3: 388-96.]. Evidently, transient expression of PMTV TGBp1 alone or together with TGBp2 and TGBp3 induced formation of typical single perinuclear aggresome-like structure in transformed cell. Moreover, colchicine blocked both TGBp1 localization to plasmodesmata and formation of perinuclear aggresomes.

Current view of TMV MP intracellular transport is based on the fact that ER membrane proteins can laterally diffuse within the membrane [47Runions J, Brach T, Kühner S, Hawes C. Photoactivation of GFP reveals protein dynamics within the endoplasmic reticulum membrane J Exp Bot 2006; 57: 43-50.] and move from cell to cell [4Epel BL. Plant viruses spread by diffusion on ER-associated movement-protein-rafts through plasmodesmata gated by viral induced host beta-1, 3-glucanases Semin Cell Dev Biol 2009; 20: 1074-81., 46Garcia-Mata R, Gao YS, Sztul E. Hassles with taking out the garbage: aggravating aggresomes Traffic 2002; 3: 388-96.]. Since the ER is tightly associated with the actin network [49Boevink P, Oparka K, Sant Cruz S, Martin B, Betteridge A, Hawes C. Stacks on tracks: The plant Golgi apparatus traffics on an actin/ER network Plant J 1998; 15: 441-7.], microfilaments could play an important role in supporting the ER-mediated targeting of MP and/or vRNA to plasmodesmata. Assuming that in plant cells MTs and actin cables may co-align at cell periphery [50Barton DA, Overall RL. Cryofixation rapidly preserves cytoskeletal arrays of leaf epidermal cells revealing microtubule co-alignments between neighbouring cells and adjacent actin and microtubule bundles in the cortex J Microsc 2010; 237: 79-88.] it can be proposed that the MP or viral transport RNP, initially anchored to MT-proximal sites, may be able to detach and undergo movements by an ER-driven mechanism [41Sambade A, Brandner K, Hofmann C, Seemanpillai M, Mutterer J, Heinlein M. Transport of TMV movement protein particles associated with the targeting of RNA to plasmodesmata Traffic 2008; 9: 2073-88.]. Our data indirectly support applicability of a similar model to the PMTV transport system. After the colchicine treatment GFP-TGBp1 was localized in the cytoplasm not diffusely but in association with an irregular network closely resembling the cortical ER, suggesting its potential ER binding ability. Although the nature of these ER-like structures remains to be established, one can speculate that TGBp1 association with the ER could be due to integral membrane proteins TGBp2 and/or TGBp3 associated with the ER or ER-derived membrane compartments [7Morozov SY, Solovyev AG. Triple gene block: modular design of a multifunctional machine for plant virus movement J Gen Virol 2003; 84: 1351-66.].

To conclude, we suggest the model of PMTV TGBp1 functioning emphasizing the important role of MTs. Early in infection TGBp1 can laterally associate with MTs to form MT-associated granular bodies. After some amounts of TGBp2 integrated into ER membranes are accumulated in infected cell, TGBp1-containing granular bodies, due to the suggested TGBp1 ability to interact with TGBp2 [16Zamyatnin AA Jr, Solovyev AG, Savenkov EI, et al. Transient coexpression of individual genes encoded by the triple gene block of potato mop-top virus reveals requirements for TGBp1 trafficking Mol Plant-Microbe Interact 2004; 17: 921-30.], are directed to the sites of TGBp2 localization at close contacts between MTs and ER. It can be speculated that the TGBp1-containing granular bodies include also RNA because of RNA binding activities of TGBp1 and TGBp2 [8Verchot-Lubicz J, Torrance L, Solovyev AG, Morozov SY, Jackson AO, Gilmer D. Varied movement strategies employed by triple gene block-encoding viruses Mol Plant-Microbe Interact 2010; 23: 1231-47.]. Increase of TGBp3 amount results in a rapid ER-driven movement of TGBp1/TGBp2/RNA complexes to plasmodesmata and through plasmodesmata to neighboring cells. Further increase of concentration of MT-bound TGBp1 results in its targeting to aggresomes for degradation that can be important at the later steps of infection cycle when virions are forming, and excessive amounts of RNA-binding protein TGBp1 can compete with the coat protein for viral genomic RNAs. Thus, we propose that TGBp1 interaction with MTs may be important for both the formation of vRNP bodies early in infection and degradation of the excessive TGBp1 at late infection stages.