We studied subcellular distribution of green fluorescent protein (GFP)-tagged movement proteins encoded by the second and the third genes of poa semilatent hordeivirus (PSLV) triple gene block (TGB), 15K TGBp2 and 18K TGBp3. GFP-15K transiently expressed in Nicotiana benthamiana leaf epidermal cells was associated with the endomembrane system elements. GFP-18K appeared in the membrane bodies at cell periphery. Mutation analysis demonstrated that subcellular targeting of GFP-15K depended on the protein transmembrane segment(s), whereas the TGBp3 central hydrophilic region was responsible for targeting of GFP-18K. Coexpression of GFP-15K with the intact 18K protein induced drastic changes in the TGBp2 localization: GFP-15K appeared in the cell peripheral bodies similar to those in the cells expressing GFP-18K alone. Coexpression experiments with mutant forms of both proteins argue against involvement of direct interaction between small TGB proteins in the TGBp3-assisted targeting of TGBp2 to the cell peripheral compartments. This conclusion was further confirmed by similar effects on the PSLV 15K TGBp2 localization induced by TGBp3 proteins of PSLV and potato virus X, which have no detectable sequence similarity to each other.

The 63 kDa ' 63K ' movement protein encoded by the triple gene block of poa semilatent virus (PSLV) comprises the C-terminal NTPase/helicase domain and the N-terminal extension domain, which contains two positively charged sequence motifs, A and B. In this study, the in vitro RNAbinding properties of PSLV 63K and its mutants were analysed. Membrane-immobilized 63K and N-63K (isolated N-terminal extension domain) bound RNA at high NaCl concentrations. In contrast, C-63K (isolated NTPase/helicase domain) was able to bind RNA only at NaCl concentrations of up to 50 mM. In gel-shift assays, C-63K bound RNA to form complexes that were unable to enter an agarose gel, whereas complexes formed by N-63K could enter the gel. Full-length 63K formed both types of complexes. Visualization of the RNA-protein complexes formed by 63K, N-63K and C-63K by atomic force microscopy demonstrated that each complex had a different shape. Collectively, these data indicate that 63K has two distinct RNA-binding activities associated with the NTPase/helicase domain and the N-terminal extension domain. Mutations in either of the positively charged sequence motifs A and B had little effect on the RNA binding of the N-terminal extension domain, whereas mutations in both motifs together inhibited RNA binding. Hybrid viruses with mutations in motifs A and B were able to infect inoculated leaves of Nicotiana benthamiana plants, but were unable to move systemically to uninoculated leaves, suggesting that the RNA-binding activity of the N-terminal extension domain of PSLV 63K is associated with virus long-distance movement.

The subcellular localization of the hydrophobic TGBp3 protein of Poa semilatent virus (PSLV, genus Hordeivirus) was studied in transgenic plants using fluorescent microscopy to detect green fluorescent protein (GFP)-tagged protein and immunodetection with monoclonal antibodies (mAbs) raised against the GFP-based fusion expressed in E. coli. In Western blot analysis, mAbs efficiently recognized the wild-type and GFP-fused PSLV TGBp3 proteins expressed in transgenic Nicotiana benthamiana, but failed to detect TGBp3 in hordeivirus-infected plants. It was found that PSLV TGBp3 and GFP-TGBp3 had a tendency to form large protein complexes of an unknown nature. Fractionation studies revealed that TGBp3 represented an integral membrane protein and probably co-localized with an endoplasmic reticulum-derived domain. Microscopy of epidermal cells in transgenic plants demonstrated that GFP-TGBp3 localized to cell wall-associated punctate bodies, which often formed pairs of opposing discrete structures that co-localized with callose, indicating their association with the plasmodesmata-enriched cell wall fields. After mannitol-induced plasmolysis of the leaf epidermal cells in the transgenic plants, TGBp3 appeared within the cytoplasm and not at cell walls. Although TGBp3-induced bodies were normally static, most of them became motile after plasmolysis and displayed stochastic motion in the cytoplasm.

The combination of interictal and ictal MEG is a valuable tool for identification of the epileptogenic tuber/tubers in presurgical work-up in patients with TS.

FURYL(ARYL)METHANES AND THEIR ANALOGS. (REVIEW) A. V. Butin, T. A. Stroganova, and V. G. Kul'nevich The literature data on furyl(arv. Omethanes are svstematized and analyzed for the first time. Results obtained by the authors on the synthesis, reactions, and application of these compounds are reviewed.~ "XHgCI + ~CH,CI ~ O~~OO I A convenient method for synthesis of difurylmethane is the reaction of furyllithium with furfurol and subsequent reduction of the intermediate carbinol with . It was noted that this is the optimal method for synthesis of difurylmethane because the product yield is 64% and the undesirable side products are