S. I. Malyshenko

Reduction of tobacco mosaic virus accumulation in transgenic plants producing non-functional viral transport proteins

Transgenic plants producing the 30K temperature-sensitive transport protein (TP) of tobacco mosaic virus (TMV) mutant Ni2519 (affecting cell-to-cell transport) were found to: (i) be susceptible to wild-type TMV U1 at 24 degrees C (a permissive temperature for Ni2519 TP), (ii) acquire a certain level of resistance to TMV U1 accumulation when maintained at 33 degrees C (a non-permissive temperature for Ni2519 TP) and (iii) lose the resistance to wild-type TMV after their transfer from 33 degrees C to 24 degrees C. It is suggested that reversible temperature-dependent conformational changes in Ni2519 TP are responsible for these phenomena and that production of a TP which is only partially functional in transgenic plants confers on these plants a resistance to the virus owing to reduction of the level of cell-to-cell transport. Transgenic tobacco plants producing the 32K TP of brome mosaic virus (BMV) acquired resistance to TMV U1 suggesting that BMV TP is partially functional in tobacco plants.

Plant virus transport function: complementation by helper viruses is non-specific.

Summary The possibility of complementation of the cell-to-cell spread (within the inoculated leaf) between different related and unrelated plant viruses has been studied. Various tobamoviruses (tobacco mosaic, sunn-hemp mosaic, cucumber green mottle mosaic viruses and ‘tobamovirus from orchids’) can facilitate each others replication in non-permissive hosts or at a temperature non-permissive for transport of one of the virus partners, probably by complementation of transport functions. Complementation of movement also occurred between some, but not all unrelated viruses tested. The complementation in transport function seems to be non-specific: it can occur between viruses even if their putative transport proteins significantly differ in structure. Consequently these viruses were classified tentatively into different ‘transport groups’.

Red Clover Mottle Comovirus B-RNA Spreads between Cells in Tobamovirus-infected Tissues

Summary B component RNA (B-RNA) of red clover mottle comovirus (RCMV) was not transported between cells of inoculated leaf tissue unless it was co-inoculated with the M component. However when host plants were infected with sunn-hemp mosaic tobamovirus (SHMV) or mutant Ni118 of tobacco mosaic virus (TMV) before superinoculation, RCMV B-RNA was transported readily between cells. Plants were infected with SHMV or Ni118 and, 5 days after superinoculation with RCMV B component, protoplasts were isolated from the infected leaves and inoculated with RCMV M component. About 30% of such protoplasts multiplied RCMV (identified by immunofluorescence microscopy) whereas only 3 to 5% of protoplasts from similarly treated plants not initially infected with SHMV or Ni118 multiplied RCMV. Thus B-RNA spread from mixedly infected (SHMV + B-RNA or Ni118 + B-RNA) cells in the absence of M-RNA to the neighbouring cells. RCMV B-RNA spread in leaves infected with the temperature-sensitive coat protein TMV mutant Ni118 and grown at non-permissive temperature. Thus TMV coat protein is not involved in enabling RCMV RNA to be transported.

Production of the tobacco mosaic virus (TMV) transport protein in transgenic plants is essential but insufficient for complementing foreign virus transport: a need for the full-length TMV genome or some other TMV-encoded product

We have reported previously that tobamoviruses enable the transport of red clover mottle comovirus (RCMV) in tobacco plants normally resistant to RCMV. Here we show that RCMV transport does not take place in transgenic tobacco plants (line To-4) producing the 30K transport protein of tobacco mosaic virus (TMV), whereas the transport of the TMV Ls1 mutant, the cell-to-cell movement of which is temperature sensitive, is complemented in these plants. However, RCMV transport is observed when these transgenic plants are infected with both RCMV and TMV Ls1 at the non-permissive temperature (33 degrees C). It is suggested that (i) the hypothetical modification of transgenic plant plasmodesmata by the TMV 30K transport protein can specifically mediate the cell-to-cell movement of the homologous virus (TMV), but is insufficient to mediate RCMV transport; (ii) the presence of the full-length TMV genome or a certain TMV-encoded product(s) besides the 30K protein is essential for complementation of the RCMV transport function. The possibility that line To-4 might provide enough 30K protein to complement TMV Ls1 but not RCMV cannot be ruled out. During double infection the mutant 30K protein may, in concert with the wild-type 30K protein, provide the transport function for RCMV.

Host-dependent Suppression of Temperature-sensitive Mutations in Tobacco Mosaic Virus Transport Gene

Summary Tobacco mosaic virus (TMV) mutants Ls1 and Ni2519, temperature-sensitive (ts) in transport function in tobacco plants, were able to spread at high temperature (33 °C) in Amaranthus caudatus L. plants. On the other hand, TMV ts coat protein mutants retained the ts phenotype in both host plants. The ability of Ls1 and Ni2519 to spread systemically in A. caudatus at high temperature is probably due to the functional stabilization of the transport proteins by a factor(s) provided by the host plant. In accordance with this, Ls1 complemented the transport function of cucumber green mottle mosaic tobamovirus and red clover mottle comovirus; they acquired the ability to spread systemically at 33 °C in the A. caudatus plants preinfected with Ls1.

The Cell to Cell Movement of Viruses in Plants

At the moment of inoculation of a plant with a virus only a negligible number of cells become infected. The virus replicates in these initially-infected cells and then moves to the neighbouring healthy ones. A separate virus-specific function, namely the transport function (TF), is coded by the viral genome; “transport” proteins are encoded in the genomes of various plant viruses. In this paper three main questions will be discussed.

Potato Spindle Tuber Viroid Does Not Complement Tobacco Mosaic Virus Temperature-sensitive Transport Function

Summary In mixed infections, the potato spindle tuber viroid (PSTV) does not complement the transport function of the tobacco mosaic virus (TMV) Ls1 mutant, which has a temperature-sensitive transport function. This appears to be due to the fundamentally different mechanisms of virus and viroid transport. However, PSTV significantly enhances the accumulation of temperature-resistant TMV in mixed infections at a temperature non-permissive for Ls1.