A host factor involved in hypovirus symptom expression in the chestnut blight fungus, Cryphonectria parasitica.
The prototype hypovirus CHV1-EP713 causes virulence attenuation and severe suppression of asexual sporulation and pigmentation in its host, the chestnut blight fungus, Cryphonectria parasitica. We identified a factor associated with symptom induction in C. parasitica using a transformation of C. parasitica strain EP155 with a full-length cDNA clone from a mild mutant virus strain, Cys(72). This was accomplished by using mutagenesis of the transformant fungal strain TCys(72)-1 by random integration of plasmid pHygR, conferring hygromycin resistance. The mutant, namA (after nami-gata, meaning wave shaped), showed an irregular fungal morphology with reduced conidiation and pigmentation while retaining similar levels of virulence and virus accumulation relative to TCys(72)-1- or Cys(72)-infected strain EP155. However, the colony morphology of virus-cured namA (VC-namA) was indistinguishable from those of EP155 and virus-cured TCys(72)-1 [VC-TCys(72)-1]. The phenotypic difference between VC-namA and VC-TCys(72)-1 was found only when these strains infected with the wild type or certain mutant CHV1-EP713 strains but not when infected with Mycoreovirus 1. Sequence analysis of inverse-PCR-amplified genomic DNA fragments and cDNA identified the insertion site of the mutagenic plasmid in exon 8 of the nam-1 gene. NAM-1, comprising 1,257 amino acids, shows sequence similarities to counterparts from other filamentous fungi and possesses the CorA domain that is conserved in a class of Mg(2+) transporters from prokaryotes and eukaryotes. Complementation assays using the wild-type and mutant alleles and targeted disruption of nam-1 showed that nam-1 with an extension of the pHygR-derived sequence contributed to the altered phenotype in the namA mutant. The molecular mechanism underlying virus-specific fungal symptom modulation in VC-namA is discussed.
Faruk
M Iqbal
MI
Agrivirology Laboratory, Research Institute for Bioresources, Okayama University, Kurashiki, Okayama 710-0046, Japan.
Eusebio-Cope
Ana
A
Suzuki
Nobuhiro
N
D001203:Ascomycota / Q000821:virology*; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics / Q000502:physiology*; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D010858:Pigmentation / Q000235:genetics; D010957:Plasmids; D017434:Protein Structure, Tertiary / Q000235:genetics; D012328:RNA Viruses / Q000472:pathogenicity*; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013172:Spores, Fungal / Q000235:genetics; D014774:Virulence / Q000235:genetics
D004271:DNA, Fungal; D005656:Fungal Proteins