A spontaneous red clover necrotic mosaic virus mutant with a truncated movement protein.
A spontaneous red clover necrotic mosaic virus mutant, TpM-341, was isolated by multiple passage of Czechoslovakian isolate TpM-34 in beans, followed by three cycles of single lesion isolation in cowpea and in Chenopodium quinoa. The symptoms induced in cowpea by TpM-34 and TpM-341 differed. TpM-34 gave rise to chlorotic lesions which expanded with time, often becoming confluent with adjacent lesions, and developed necrotic margins; the plants became systemically infected. TpM-341 induced necrotic lesions which, once developed, did not expand further; plants did not become systemically infected. Analysis of pseudorecombinants formed between the RNAs of TpM-34 and TpM-341 showed that RNA 2 determined the difference in symptoms. Comparison of the nucleotide sequence of the open reading frames (ORFs) encoding the P2 movement proteins of the two isolates revealed only one difference, a deletion of an A residue in a sequence of four A residues (nucleotides 790 to 793). Construction of full-length TpM-34 and TpM-341 RNA 2 cDNA clones, from which infectious RNA 2 could be transcribed in vitro, and in vitro mutagenesis of a cDNA clone of TpM-341, confirmed that the difference in the symptoms induced by TpM-341 was caused by the loss of this A residue. This single base deletion was predicted to cause a translational frame-shift in the P2 ORF causing the loss of 88 amino acids at the C terminus which were replaced by a sequence of 34 different amino acids, producing a truncated P2 protein. In vitro translation of RNA 2 transcribed from cDNA clones showed that RNA with four or three A residues starting at nucleotide 790 produced proteins of Mr 36K and 30K respectively, in agreement with the predictions based on the nucleotide sequence.
Osman
T A
TA
Department of Biology, Imperial College of Science, Technology and Medicine, London, U.K.
Ingles
P J
PJ
Miller
S J
SJ
Buck
K W
KW
D001483:Base Sequence; D002465:Cell Movement / Q000235:genetics; D003001:Cloning, Molecular; D004279:DNA, Viral; D004587:Electrophoresis, Agar Gel; D007887:Fabaceae / Q000382:microbiology; D008969:Molecular Sequence Data; D009029:Mosaic Viruses / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D009154:Mutation; D016366:Open Reading Frames; D010935:Plant Diseases; D010946:Plants, Medicinal; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D014764:Viral Proteins / Q000235:genetics*; D014779:Virus Replication
D004279:DNA, Viral; D012367:RNA, Viral; D014764:Viral Proteins