Construction of an infectious cDNA clone of aichi virus (a new member of the family Picornaviridae) and mutational analysis of a stem-loop structure at the 5′ end of the genome

Jun Sasaki, Y. Kusuhara, Yoshimasa Maeno, N. Kobayashi, T. Yamashita, K. Sakae, N. Takeda, K. Taniguchi

Research output: Contribution to journalArticle

50 Citations (Scopus)

Abstract

Aichi virus is the type species of a new genus, Kobuvirus, of the family Picornaviridae. In this study, we constructed a full-length cDNA clone of Aichi virus whose in vitro transcripts were infectious to Vero cells. During construction of the infectious cDNA clone, a novel sequence of 32 nucleotides was identified at the 5′ end of the genome. Computer-assisted prediction of the secondary structure of the 5′ end of the genome, including the novel sequence, suggested the formation of a stable stem-loop structure consisting of 42 nucleotides. The function of this stem-loop in virus replication was investigated using various site-directed mutants derived from the infectious cDNA clone. Our data indicated that correct folding of the stem-loop at the 5′ end of the positive strand, but not at the 3′ end of the negative strand, is critical for viral RNA replication. The primary sequence in the lower part of the stem was also suggested to be crucial for RNA replication. In contrast, nucleotide changes in the loop segment did not so severely reduce the efficiency of virus replication. A double mutant, in which both nucleotide stretches of the middle part of the stem were replaced by their complementary nucleotides, had efficient RNA replication and translation abilities but was unable to produce viruses. These results indicate that the stem-loop at the 5′ end of the Aichi virus genome is an element involved in both viral RNA replication and production of infectious virus particles.

Original languageEnglish
Pages (from-to)8021-8030
Number of pages10
JournalJournal of Virology
Volume75
Issue number17
DOIs
Publication statusPublished - 23-08-2001

Fingerprint

Aichi virus
Kobuvirus
Picornaviridae
Nucleotides
Complementary DNA
Clone Cells
Genome
clones
nucleotides
stems
genome
Viral RNA
Virus Replication
RNA
virus replication
Vero Cells
Aptitude
Virion
mutants
Viruses

All Science Journal Classification (ASJC) codes

  • Microbiology
  • Immunology
  • Insect Science
  • Virology

Cite this

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title = "Construction of an infectious cDNA clone of aichi virus (a new member of the family Picornaviridae) and mutational analysis of a stem-loop structure at the 5′ end of the genome",
abstract = "Aichi virus is the type species of a new genus, Kobuvirus, of the family Picornaviridae. In this study, we constructed a full-length cDNA clone of Aichi virus whose in vitro transcripts were infectious to Vero cells. During construction of the infectious cDNA clone, a novel sequence of 32 nucleotides was identified at the 5′ end of the genome. Computer-assisted prediction of the secondary structure of the 5′ end of the genome, including the novel sequence, suggested the formation of a stable stem-loop structure consisting of 42 nucleotides. The function of this stem-loop in virus replication was investigated using various site-directed mutants derived from the infectious cDNA clone. Our data indicated that correct folding of the stem-loop at the 5′ end of the positive strand, but not at the 3′ end of the negative strand, is critical for viral RNA replication. The primary sequence in the lower part of the stem was also suggested to be crucial for RNA replication. In contrast, nucleotide changes in the loop segment did not so severely reduce the efficiency of virus replication. A double mutant, in which both nucleotide stretches of the middle part of the stem were replaced by their complementary nucleotides, had efficient RNA replication and translation abilities but was unable to produce viruses. These results indicate that the stem-loop at the 5′ end of the Aichi virus genome is an element involved in both viral RNA replication and production of infectious virus particles.",
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Construction of an infectious cDNA clone of aichi virus (a new member of the family Picornaviridae) and mutational analysis of a stem-loop structure at the 5′ end of the genome. / Sasaki, Jun; Kusuhara, Y.; Maeno, Yoshimasa; Kobayashi, N.; Yamashita, T.; Sakae, K.; Takeda, N.; Taniguchi, K.

In: Journal of Virology, Vol. 75, No. 17, 23.08.2001, p. 8021-8030.

Research output: Contribution to journalArticle

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T1 - Construction of an infectious cDNA clone of aichi virus (a new member of the family Picornaviridae) and mutational analysis of a stem-loop structure at the 5′ end of the genome

AU - Sasaki, Jun

AU - Kusuhara, Y.

AU - Maeno, Yoshimasa

AU - Kobayashi, N.

AU - Yamashita, T.

AU - Sakae, K.

AU - Takeda, N.

AU - Taniguchi, K.

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