Combinations of two amino acids (Ala40 and Phe75 or Ser40 and Tyr75) in the coat protein of apple chlorotic leaf spot virus are crucial for infectivity.
Amino acid sequences of apple chlorotic leaf spot virus (ACLSV) coat protein (CP) were compared between 12 isolates from apple, plum and cherry, and 109 cDNA clones that were amplified directly from infected apple tissues. Phylogenetic analysis based on the amino acid sequences of CP showed that the isolates and cDNA clones were separated into two major clusters in which the combinations of the five amino acids at positions 40, 59, 75, 130 and 184 (Ala(40)-Val(59)-Phe(75)-Ser(130)-Met(184) or Ser(40)-Leu(59)-Tyr(75)-Thr(130)-Leu(184)) were highly conserved within each cluster. Site-directed mutagenesis using an infectious cDNA clone of ACLSV indicated that the combinations of two amino acids (Ala(40) and Phe(75) or Ser(40) and Tyr(75)) are necessary for infectivity to Chenopodium quinoa plants by mechanical inoculation. Moreover, an agroinoculation assay indicated that the substitution of a single amino acid (Ala(40) to Ser(40) or Phe(75) to Tyr(75)) resulted in extreme reduction in the accumulation of viral genomic RNA, double-stranded RNAs and viral proteins (movement protein and CP) in infiltrated tissues, suggesting that the combinations of the two amino acids at positions 40 and 75 are important for effective replication in host plant cells.
Yaegashi
Hajime
H
The United Graduate School of Agricultural Sciences, Iwate University, Morioka 020-8550, Japan.
Isogai
Masamichi
M
Plant Pathology Laboratory, Faculty of Agriculture, Iwate University, Morioka 020-8550, Japan.|The United Graduate School of Agricultural Sciences, Iwate University, Morioka 020-8550, Japan.
Tajima
Hiroko
H
Plant Pathology Laboratory, Faculty of Agriculture and Life Sciences, Hirosaki University, Hirosaki 036-8561, Japan.
Sano
Teruo
T
Plant Pathology Laboratory, Faculty of Agriculture and Life Sciences, Hirosaki University, Hirosaki 036-8561, Japan.|The United Graduate School of Agricultural Sciences, Iwate University, Morioka 020-8550, Japan.
Yoshikawa
Nobuyuki
N
Plant Pathology Laboratory, Faculty of Agriculture, Iwate University, Morioka 020-8550, Japan.|The United Graduate School of Agricultural Sciences, Iwate University, Morioka 020-8550, Japan.
D000409:Alanine; D000595:Amino Acid Sequence; D017931:DNA Primers; D052677:Flexiviridae / Q000235:genetics* / Q000472:pathogenicity* / Q000502:physiology; D027845:Malus / Q000821:virology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010649:Phenylalanine; D010935:Plant Diseases / Q000821:virology*; D018515:Plant Leaves / Q000821:virology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid; D012694:Serine; D014443:Tyrosine; D014779:Virus Replication
D017931:DNA Primers; D014443:Tyrosine; D012694:Serine; D010649:Phenylalanine; D000409:Alanine