Isolation, characterization and mutation analysis of PEX13-defective Chinese hamster ovary cell mutants.
We isolated peroxisome biogenesis mutants ZP128 and ZP150 from rat PEX2 -transformed Chinese hamster ovary (CHO) cells, by the 9-(1'-pyrene)nonanol/ultraviolet method. The mutants lacked morphologically recognizable peroxisomes and showed a typical peroxisome assembly-defective phenotype such as a high sensitivity to 12-(1'-pyrene)dodecanoic acid/UV treatment. By means of PEX cDNA transfection and cell fusion, ZP128 and ZP150 were found to belong to a recently identified complementation group H. Expression of human PEX13 cDNA restored peroxisome assembly in ZP128 and ZP150. CHO cell PEX13 was isolated; its deduced sequence comprises 405 amino acids with 93% identity to human Pex13p. Mutation in PEX13 of mutant ZP150 was determined by RT-PCR: G to A transition resulted in one amino acid substitution, Ser319Asn, in one allele and truncation of a 42 amino acid sequence from Asp265 to Lys306 in another allele. Therefore, ZP128 and ZP150 are CHO cell lines with a phenotype of impaired PEX13.
Toyama
R
R
Department of Biology, Kyushu University Graduate School of Science, Fukuoka 812-8581, Japan.
Mukai
S
S
Itagaki
A
A
Tamura
S
S
Shimozawa
N
N
Suzuki
Y
Y
Kondo
N
N
Wanders
R J
RJ
Fujiki
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D004252:DNA Mutational Analysis; D005455:Fluorescent Antibody Technique; D005816:Genetic Complementation Test; D006801:Humans; D006822:Hybrid Cells; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D000074428:Peroxisomal Biogenesis Factor 2; D020675:Peroxisomes / Q000235:genetics* / Q000473:pathology; D014162:Transfection; D014170:Transformation, Genetic
D008565:Membrane Proteins; C101884:PEX13 protein, human; C000614230:PEX2 protein, human; C000614232:Pex2 protein, rat; D000074428:Peroxisomal Biogenesis Factor 2