Isolation, characterization and mutation analysis of PEX13-defective Chinese hamster ovary cell mutants

Abstract

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.