70-kDa peroxisomal membrane protein related protein (P70R/ABCD4) localizes to endoplasmic reticulum not peroxisomes, and NH2-terminal hydrophobic property determines the subcellular localization of ABC subfamily D proteins.
70-kDa peroxisomal membrane protein related protein (P70R/ABCD4) is a member of ATP-binding cassette (ABC) protein subfamily D. ABC subfamily D proteins are also known as peroxisomal ABC proteins. Therefore, P70R is thought to be a peroxisomal membrane protein. However, the subcellular localization of P70R is not extensively investigated. In this study, we transiently expressed P70R in fusion with HA (P70R-HA) in CHO cells and examined subcellular localization by immunofluorescence. Surprisingly, P70R-HA was localized to the endoplasmic reticulum (ER), not to peroxisomes. To examine the ER-targeting property of P70R, we expressed various NH(2)-terminal deletion constructs of P70R. Among the NH(2)-terminal deletion constructs, mutant proteins starting with hydrophobic transmembrane segment (TMS) were localized to ER, but the ones containing the NH(2)-terminal hydrophilic cytosolic domain were not. ABC subfamily D proteins destined for peroxisomes have NH(2)-terminal hydrophilic region adjacent to TMS1. However, only P70R lacks the region and is translated with NH(2)-terminal hydrophobic TMS1. Furthermore, attachment of the NH(2)-terminal hydrophilic domain to the NH(2)-terminus of P70R excluded P70R from the ER-targeting pathway. These data suggest that P70R resides in the ER but not the peroxisomal membranes, and the hydrophobic property of NH(2)-terminal region determines the subcellular localization of ABC subfamily D proteins.
Kashiwayama
Yoshinori
Y
Department of Biological Chemistry, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, 2630 Sugitani, Toyama 930-0194, Japan.
Seki
Midori
M
Yasui
Akina
A
Murasaki
Yoshiyuki
Y
Morita
Masashi
M
Yamashita
Yukari
Y
Sakaguchi
Masao
M
Tanaka
Yoshitaka
Y
Imanaka
Tsuneo
T
D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D016466:CHO Cells; D019556:COS Cells; D002522:Chlorocebus aethiops; D006224:Cricetinae; D003412:Cricetulus; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006031:Glycosylation; D007425:Intracellular Membranes / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D008297:Male; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase / Q000378:metabolism; D020675:Peroxisomes / Q000378:metabolism*; D011485:Protein Binding; D021382:Protein Sorting Signals / Q000502:physiology*; D021381:Protein Transport / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D017384:Sequence Deletion; D014162:Transfection
D018528:ATP-Binding Cassette Transporters; C489274:Abcd4 protein, mouse; D021382:Protein Sorting Signals; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase