Genetic and molecular analysis of the temperature-sensitive mutant un-17 carrying a mutation in the gene encoding poly(A)-polymerase in Neurospora crassa.
The un-17 mutant was originally isolated as an irreparable temperature-sensitive (ts) mutant in Neurospora crassa. Early experiments showed that cells of this mutant immediately stopped growing and died when the temperature of the culture was shifted from a permissive temperature (25 degrees C) to non-permissive temperature (35 degrees C). This ts phenotype is suppressed by addition of cycloheximide or in some conditions of growth repression. Even at the permissive temperature, it shows a female sterile phenotype and is deficient in production of exocellular superoxide dismutase SOD4 (EC 1.15.1.1). By searching for a DNA fragment that complements the ts phenotype of the un-17 mutant from a N. crassa genome library, we found the un-17 gene. The cloned un-17 gene encodes a homolog of the Saccharomyces cerevisiae poly(A) polymerase (PAP). The un-17 mutant had a one-base substitution mutation in the gene. The cloned un-17 genes from the wild-type strain and the un-17 mutant were introduced into both the un-17 mutant and wild-type strain. The un-17 mutant introduced by un-17 DNA from the wild-type strain showed recovery of both the ts and female sterile phenotypes. Moreover, the purified product derived from the wild-type strain showed PAP activity in vitro. These findings indicate that the un-17 mutant carries a ts mutation in the gene encoding PAP.
Tanaka
Shuuitsu
S
Laboratory of Genetics, Department of Regulation-Biology, Faculty of Science, Saitama University, Saitama City, Japan. shtanaka@post.saitama-u.ac.jp
Takayanagi
Noriko
N
Murasawa
Kennichi
K
Ishii
Chizu
C
Inoue
Hirokazu
H
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics; D005800:Genes, Fungal; D008969:Molecular Sequence Data; D009154:Mutation; D009492:Neurospora crassa / Q000201:enzymology / Q000235:genetics*; D010641:Phenotype; D011062:Polynucleotide Adenylyltransferase / Q000235:genetics* / Q000378:metabolism; D013696:Temperature
D004271:DNA, Fungal; D005656:Fungal Proteins; D011062:Polynucleotide Adenylyltransferase