PEX12, the pathogenic gene of group III Zellweger syndrome:: cDNA cloning by functional complementation on a CHO cell mutant, patient analysis, and characterization of Pex12p

被引:92
作者
Okumoto, K
Shimozawa, N
Kawai, A
Tamura, S
Tsukamoto, T
Osumi, T
Moser, H
Wanders, RJA
Suzuki, Y
Kondo, N
Fujiki, Y
机构
[1] Kyushu Univ, Fac Sci, Dept Biol, Higashi Ku, Fukuoka 8128581, Japan
[2] Gifu Univ, Sch Med, Dept Pediat, Gifu 5008076, Japan
[3] Himeji Inst Technol, Dept Life Sci, Kamigori, Hyogo 6781297, Japan
[4] Japan Sci & Technol Corp, CREST, Tokyo 1700013, Japan
[5] Johns Hopkins Univ, Sch Med, Kennedy Krieger Inst, Baltimore, MD 21205 USA
[6] Univ Amsterdam, Acad Med Ctr, Dept Pediat, NL-1100 DE Amsterdam, Netherlands
关键词
D O I
10.1128/MCB.18.7.4324
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Rat PEX12 cDNA was isolated by functional complementation of peroxisome deficiency of a mutant CHO cell line, ZP109 (K. Okumoto, A. Bogaki, K. Tateishi, T. Tsukamoto, T. Osumi, N. Shimozawa, Y. Suzuki, T. Orii, and Y. Fujiki, Exp. Cell Res. 233:11-20, 1997), using a transient transfection assay and an ectopic, readily visible marker, green fluorescent protein. This cDNA encodes a 359-amino-acid membrane protein of peroxisomes with two transmembrane segments and a cysteine rich zinc finger, the RING motif. A stable transformant of ZP109 with the PEX12 was morphologically and biochemically restored for peroxisome biogenesis. Pex12p was shown by expression of bona fide as well as epitope-tagged Pex12p to expose both N- and C-terminal regions to the cytosol. Fibroblasts derived from patients with the peroxisome deficiency Zellweger syndrome of complementation group LII (CG-III) were also complemented for peroxisome biogenesis with PEX12. Two unrelated patients of this group manifesting peroxisome deficiency disorders possessed homozygous, inactivating PEX12 mutations: in one, Arg180Thr by one point mutation, and in the other, deletion of two nucleotides in codons for (291)Asn and (292)Ser, creating an apparently unchanged codon for Asn and a codon 292 for termination. These results indicate that the gene encoding peroxisome assembly factor Pex12p is a pathogenic gene of CG-m peroxisome deficiency. Moreover, truncation and site mutation studies, including patient PEX12 analysis, demonstrated that the cytoplasmically oriented N- and C-terminal parts of Pex12p are essential for biological function.
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页码:4324 / 4336
页数:13
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