A novel pex2 mutant: catalase-deficient but temperature-sensitive PTS1 and PTS2 import.
We searched for Chinese hamster ovary (CHO) cell mutants defective in peroxisome biogenesis by using peroxisome targeting sequence (PTS) of Pex3p (amino acid residues 1-40)-fused enhanced green fluorescent protein (EGFP). From mutagenized wild-type CHO-K1 cells stably expressing rat Pex2p and Pex3p(1-40)-EGFP, cell colonies resistant to the 9-(1(')-pyrene)nonanol/ultraviolet treatment were examined for intracellular location of peroxisomal proteins, including EGFP chimera, catalase, and matrix proteins with PTS types 1 and 2. One clone, ZPEG309, showed a distinct phenotype: import defect of catalase, but normal transport of PTS1 and PTS2 proteins at 37 degrees C. PTS1 and PTS2 import was abrogated when ZPEG309 was cultured at 39 degrees C. Genetic defect of ZPEG309 was a nonsense point mutation in a codon for Arg50 in CHO PEX2 and a mutation resulting in a C-terminal truncation of the introduced rat Pex2p. Therefore, ZPEG309 is a novel pex2, catalase-deficient mutant with temperature-sensitive PTS1 and PTS2 import.
Akiyama
Noriko
N
Department of Biology, Faculty of Sciences, Kyushu University Graduate School, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan.
Ghaedi
Kamran
K
Fujiki
Yukio
Y
D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D004252:DNA Mutational Analysis; D005816:Genetic Complementation Test; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D009154:Mutation; D000074428:Peroxisomal Biogenesis Factor 2; D000074437:Peroxisomal Targeting Signal 2 Receptor; D000074435:Peroxisome-Targeting Signal 1 Receptor; D010641:Phenotype; D010957:Plasmids / Q000378:metabolism; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D051381:Rats; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013696:Temperature; D014162:Transfection
D008164:Luminescent Proteins; D008565:Membrane Proteins; D000074437:Peroxisomal Targeting Signal 2 Receptor; D000074435:Peroxisome-Targeting Signal 1 Receptor; C000614232:Pex2 protein, rat; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D000074428:Peroxisomal Biogenesis Factor 2; D049452:Green Fluorescent Proteins