We recently noticed that some figures contained Northern and Western blot images comprising spliced lanes and nonuniform contrast adjustments that were not indicated as such. Upon closer inspection of the published figures and the original material that was still available, we also realized that some lanes of one Western blot and one Northern blot were reused in assembling different figures. In addition, some figures presented signs of inappropriate manipula-tion. The details of these issues are as follows.

Point mutations were introduced into the genes encoding the triple gene bock movement proteins P13 and P15 of beet necrotic yellow vein virus (BNYVV). Mutations which disabled viral cell-to-cell movement in Chenopodium quinoa were then tested for their ability to act as dominant negative inhibiters of movement of wild-type BNYVV when expressed from a co-inoculated BNYVV RNA 3-based replicon. For P13, three types of mutation inhibited the movement function: non-synomynous mutations in the N- and C-terminal hydrophobic domains, a mutation at the boundary between the N-terminal hydrophobic domain and the central hydrophilic domain (mutant P13-A12), and mutations in the conserved sequence motif in the central hydrophilic domain. However, only the 'boundary' mutant P13-A12 strongly inhibited movement of wild-type virus when expressed from the co-inoculated replicon. Similar experiments with P15 detected four movement-defective mutants which strongly inhibited cell-to-cell movement of wild-type BNYVV when the mutants were expressed from a co-inoculated replicon. Beta vulgaris transformed with two of these P15 mutants were highly resistant to fungus-mediated infection with BNYVV.

The subcellular localization of the first triple gene block protein (TGBp1) of peanut clump pecluvirus (PCV) was studied by subcellular fractionation and immunogold cytochemistry using TGBp1-specific antibodies raised against a fusion protein expressed in and purified from bacteria. In the inoculated and apical leaves of virus-infected Nicotiana benthamiana, TGBp1 localized to the cell wall and P30 fractions. Electron microscopy of immunogold-decorated ultrathin sections of the infected leaf tissue revealed TGBp1-specific labeling of the plasmodesmata joining mesophyll cells. In longitudinal sections of the plasmodesmata, the TGBp1-specific labeling was most commonly associated with the plasmodesmal collar region. In transgenic N. benthamiana, which constitutively expressed TGBp1, no TGBp1-specific immunogold labeling of plasmodesmata was observed, but plasmodesmata were gold decorated when the transgenic plants were infected with a TGBp1-defective PCV mutant, indicating that factors induced by the virus infection target and/or anchor the transgene TGBp1 to the plasmodesmata.

Beet necrotic yellow vein virus RNA 1 contains a single long ORF corresponding to the theoretical translation product of 237 kDa which contains the information necessary for replication of the viral genome. This ORF contains a putative papain-like proteinase domain which has been localized, on the basis of sequence alignments, between the helicase and polymerase domains. Here we show that the RNA 1 primary translation product can be cleaved autocatalytically in vitro into two species of 150 kDa and 66 kDa, the latter of which probably contains the entire polymerase domain. A 66 kDa protein was detected immunologically in infected C. quinoa protoplasts using an antiserum specific for the C-terminal region of the RNA 1 primary translation product, confirming that processing also occurs in vivo.

The complete nucleotide sequence of the genomic RNA of beet mild yellowing virus, isolate 2ITB, is reported. The RNA consists of 5722 nucleotides and contains six long open reading frames which conform to the arrangement characteristic of Subgroup 2 luteoviruses. The three 3'-proximal open reading frames, which encode the viral coat protein, a putative movement protein and the Readthrough Domain, are highly homologous to the corresponding genes of beet western yellows luteovirus while the three 5'-proximal open reading frames are more closely related to the corresponding genes of cucurbit aphid borne yellows luteovirus. The sequence data thus indicate that beet mild yellowing virus should be considered a distinct virus rather than a strain of beet western yellows virus.

Long internal deletions were introduced into cloned cDNA of beet necrotic yellow vein virus RNAs 1-4 and transcripts containing the deletions were tested for their ability to inhibit replication of viral RNA in Chenopodium quinoa protoplasts and plants. No inhibition was observed with the deletion mutants based on RNAs 1, 3 and 4 but the RNA 2 deletion mutants all provoked a dramatic inhibition of synthesis of viral RNAs 1 and 2.

RNA 2 of beet necrotic yellow vein virus carries the cistron for the 21 kd coat protein at its 5'-extremity. During translation, the coat protein cistron termination codon is suppressed about 10% of the time so that translation continues into the adjacent open reading frame to produce a 75 kd species, known as P75, which contains the coat protein sequence at its N-terminus. Immunoblotting experiments with a P75-specific antiserum showed that P75 is present in only trace amounts in purified virus preparations. Electron microscopic visualization of immunogold-labelled virions in crude tissue extracts has provided evidence for an association between P75 and at least a fraction of the BNYVV particles, with P75 being predominantly located near one end of the rod-shaped virions. This finding is discussed in the context of the current model for the role of P75 in virus assembly and vector transmission.

Anthrone gives Diels-Alder adducts with dimethyl acetylenedicarboxylate and with maleic anhydride, which have been shown to be 9,10-bridged derivatives of 9-anthrol. The adduct with maleic anhydride rearranges on heating to give 10-anthronylsuccinic anhydride. Reaction of some other dienophiles (tetracyanoethylene, diethyl azodicarboxylate, or chloranil) with anthrone gives bianthron-9-yl.The n.m.r. spectra of these compounds are discussed.

The addition reactions of 9,10-anthraquinodimethane with maleic anhydride, dimethyl acetylenedicarboxylate, p-benzoquinone, and 1,4-naphthoquinone have been studied. Syntheses of benzo[a]pyrene, dibenzo[j,xyz]-heptaphene and some substituted and reduced derivatives of these hydrocarbons are reported, and the preparation of 1,4:11,14-dibenzo[h,rst]pentaphenediquinone. The reaction of 10-hydroxy-10-methylanthrone with dimethyl acetylenedicarboxylate and with diethyl azodicarboxylate has been investigated; the latter gives the novel 7H-di-benzo[de,h]phthalazine system.

The 5,6-dihydro-5,6-dimethylchrysene-5,6-diol obtained from 5,6-chrysenequinone and methylmagnesium iodide is shown to be a trans-racemic form. The diol can be thermally dehydrate to give 5,6-chrysenequinodimethane; Diels–Alder reaction of this with maleic anhydride and with 1,4-naphthaquinone gives derivatives of benzo[g]-chrysene and benzo[c]naphtho[1,2-a]naphthacene, respectively, from which the parent hydrocarbons have been obtained.The reaction of the diol with methanolic hydrogen chloride at 0° has been studied; it gives 6-chloromethyl-5-methylchrysene and 12-chloro-5,6-dimethylchrysene. The mechanism of this reaction and the proton magnetic resonance spectra of several chrysene derivatives are discussed.

R. E. Bowman, P. J. Islip, I. M. Lockhart, K. E. Richards and M. Wright, J. Chem. Soc., 1965, 1080 DOI: 10.1039/JR9650001080

D. E. Ames, D. Evans, T. F. Grey, P. J. Islip and K. E. Richards, J. Chem. Soc., 1965, 2636 DOI: 10.1039/JR9650002636

I. T. Millar and K. E. Richards, Chem. Commun. (London), 1965, 369 DOI: 10.1039/C19650000369

The sequence of the first 59 nucleotides from the 3'-OH terminus of high-molecular-weight eggplant mosaic virus RNA has been determined by standard radiochemical techniques. The fragment was identified among the products of partial Ti RNase digestion by making use of the reverse migration, at pH 2.5, of the 3'-OH terminal oligonucleotide. No abnormal bases were found. A model of secondary structure may be constructed for this fragment, which is known to fix valine in the presence of valyl-tRNA synthetase. Its relation to the structures of genuine tRNAs and to the 3'-OH termini of other viral RNAs that also accept amino acids is discussed. The single-stranded RNA genome of eggplant mosaic virus (EMV), in common with a number of other plant viral RNAs, can accept an amino acid at the 3'-end upon treatment with a specific aminoacyl-tRNA-synthetase (1-6). EMV RNA and the RNA of the related turnip yellow mosaic virus, the first viral RNA shown to serve as an acceptor, are both efficiently charged with valine in the presence of ATP and Escherichia coli valyl-tRNA synthetase [EC 6.1.1.9; Lvaline:tRNAval ligase (AMP-forming)] (1, 5-7). Thus, it had been suggested that these two RNAs bear tRNAval-like structures at their 3' extremities. Tobacco mosaic virus RNA can be enzymatically aminoacylated with histidine at its 3'-end (3). The nucleotide sequence of the 71 3'-terminal residues of that RNA, however, is not such that it can be folded into a cloverleaf-type secondary structure characteristic of all tRNAs (7). Apparently such a structure is not a strict prerequisite for the assumption of a three-dimensional conformation which may be recognized, albeit imperfectly, by a tRNA-specific enzyme. It has been our aim to determine the 3'-terminal sequence of other viral RNAs that can be aminoacylated in order to see if they are more evidently tRNA-like in secondary structure. The present paper describes the sequence of the first 59 nucleotides at the 3'-end of EMV RNA and proposes a model for its secondary structure. Analogies and differences with genuine tRNAs and the amino acid accepting 3'-ends of other plant viral RNAs will be discussed. MATERIALS AND METHODS The growth of 32P-labeled EMV on Datura stramonium, the purification of the virus, and extraction of its RNA have been described elsewhere (7). The specific activity of the purified EMV RNA used in this work was about 0.03 Ci/g. In addition to high-molecular-weight RNA (2.3 X 106), infectious EMV particles contain a 4S RNA species that is able to bind lysine (5, 8). The 4S RNA was eliminated from the preparation of high-molecular-weight RNA by filtration on saccharose-Indubiose (9). Abbreviation: EMV, eggplant mosaic virus. 737 The purified 32P-labeled high-molecular-weight EMV RNA was partially digested with 1 unit of T1 RNase per 250 gg of RNA at 00 in 0.1 M Tris, 0.001 M EDTA, pH 7.4. After 30 min the action of the enzyme was stopped by phenol extraction and the mixture of RNA fragments was fractionated by electrophoresis in a 10% polyacrylamide gel (8). The fragment originating from the 3'-OH terminus of intact RNA (T3'-OH) was identified as described by Guilley et al. (8), with one modification: U2 RNase rather than T1 RNase was used to release the 3'-terminal oligonucleotide. Once identified, fragment T3'-OH was purified by a further electrophoresis in a 20% polyacrylamide gel, and its sequence was characterized by standard radiochemical techniques (10). In the course of this work we noticed that total T1 RNase digestion of the fragment under standard conditions induced additional cuts in large oligonucleotides rich in pyrimidine residues, leading to spots in the fingerprint that did not contain guanosine residues. Such nonspecific degradation could be largely avoided by the use of poly(cytidylic acid) (50 ,g/unit of T1 RNase) rather than tRNA as the carrier. The oligonucleotides of fragment T3'-OH were ordered by examination of T1 RNase partial digestion products. Partial digestion was with 1 unit of T1 RNase per 500 Mg of RNA, for 10 min at 00, in 0.01 M Tris, 0.001 M EDTA, pH 7.4. The resulting fragments were separated from one another by electrophoresis upon a 15% polyacrylamide gel, purified on 20-24% gels, and fingerprinted.