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) localizes to endoplasmic reticulum not peroxisomes, and NH2-terminal hydrophobic property determines the subcellular localization of ABC subfamily D proteins
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DOI：
10.1016/j.yexcr.2008.10.031
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发表时间：
2009-01-15
影响因子：
3.7
通讯作者：
Imanaka, Tsuneo
中科院分区：
文献类型：
--
作者：
Kashiwayama, Yoshinori;Seki, Midori;Imanaka, Tsuneo
70-过氧化物酶体膜蛋白相关蛋白（P70 R/ABCD 4）是ATP结合盒（ABC）蛋白亚家族D的成员。ABC亚家族D蛋白也称为过氧化物酶体ABC蛋白。因此，P70 R被认为是一种过氧化物酶体膜蛋白。然而，P70 R的亚细胞定位尚未得到广泛研究。在这项研究中，我们在CHO细胞中瞬时表达P70 R与HA融合（P70 R-HA），并通过免疫荧光检测亚细胞定位。令人惊讶的是，P70 R-HA定位于内质网（ER），而不是过氧化物酶体。为了检测P70 R的ER-taggeting性质，我们表达了P70 R的各种NH 2-末端缺失构建体。在NH 2-末端缺失构建体中，以疏水性跨膜片段（TMS）开始的突变蛋白定位于ER，但含有NH 2-末端亲水性胞质结构域的突变蛋白不定位于ER。ABC亚家族D蛋白具有与TMS 1相邻的NH 2-末端亲水区域，但是，只有P70 R缺乏该区域，并且翻译有NH 2-末端疏水性TMSI。此外，NH 2-末端亲水结构域与P70 R的NH 2-末端的连接将P70 R从ER靶向途径中排除。这些数据表明，P70 R驻留在ER，而不是过氧化物酶体膜，和NH 2-末端区域的疏水性决定了ABC亚家族D蛋白的亚细胞定位。(C)2008年爱思唯尔公司All rights reserved.
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-tatgeting property of P70R, we expressed various NH2-terminal deletion constructs of P70R. Among the NH2-terminal deletion constructs, mutant proteins starting with hydrophobic transmembrane segment (TMS) were localized to ER, but the ones containing the NH2-terminal hydrophilic cytosolic domain were not. ABC subfamily D proteins destined for peroxisomes have NH2-terminal hydrophilic region adjacent to TMS1 However, only P70R lacks the region and is translated with NH2-terminal hydrophobic TMSI. Furthermore, attachment of the NH2-terminal hydrophilic domain to the NH2-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 NH2-terminal region determines the subcellular localization of ABC subfamily D proteins. (C) 2008 Elsevier Inc. All rights reserved.