SLC35B1 significantly contributes to the uptake of UDPGA into the endoplasmic reticulum for glucuronidation catalyzed by UDP-glucuronosyltransferases.
SLC35B1 significantly contributes to the uptake of UDPGA into the endoplasmic reticulum for glucuronidation catalyzed by UDP-glucuronosyltransferases.
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DOI：
10.1016/j.bcp.2020.113916
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发表时间：
2020-03
影响因子：
5.8
通讯作者：
Kyoko Ondo;H. Arakawa;Masataka Nakano;T. Fukami;M. Nakajima
中科院分区：
文献类型：
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作者：
Kyoko Ondo;H. Arakawa;Masataka Nakano;T. Fukami;M. Nakajima
UDP-葡萄糖醛酸 (UDPGA)（UDP-葡萄糖醛酸基转移酶 (UGT) 的共底物）转运至内质网 (ER) 的腔内侧是外源和内源化合物葡萄糖醛酸化的重要步骤。根据之前的一项研究，重组 SLC35B1、SLC35B4 或 SLC35D1（核苷酸糖转运蛋白）在 V79 细胞中的表达具有将 UDPGA 转运到微粒体腔中的潜力。本研究的目的是检查这些转运蛋白对 UDPGA 的转运是否会显着影响 UGT 活性。由于稳定表达 UGT1A1 的 HEK293 细胞（HEK/UGT1A1 细胞）中 UDP-葡萄糖 6-脱氢酶（UDPGA 的合成酶）的敲低导致 4-甲基伞形酮 (4-MU) 葡萄糖醛酸基转移酶活性显着降低，因此需要补充足够量的 UDPGA 来维持 UGT 活性。通过使用 21 个人类肝脏样本的 cDNA 样本进行 qRT-PCR，我们观察到 SLC35B1 和 SLC35D1 mRNA 的水平分别比 SLC35B4 mRNA 的水平高 15 倍和 14 倍，并且 SLC35B1 显示出最大的个体间变异性（37 倍）。有趣的是，在HEK/UGT1A1细胞中敲低SLC35B1后，4-MU葡萄糖醛酸基转移酶活性显着降低，并且在HepaRG细胞中也观察到这种现象。使用针对 23 个不同 SLC35 亚家族的 siRNA，SLC35B1 和 SLC35E3 的敲低降低了 HEK/UGT1A1 细胞中的 4-MU 葡萄糖醛酸基转移酶活性。然而，HepaRG 细胞中 SLC35E3 敲低并未改变 4-MU 葡萄糖醛酸基转移酶活性，这表明 SLC35B1 是 UDPGA 进入人肝脏内质网的主要转运蛋白。 In conclusion, SLC35B1 is a key modulator of UGT activity by transporting UDPGA to the intraluminal side of the ER.
The transport of UDP-glucuronic acid (UDPGA), a co-substrate of UDP-glucuronosyltransferase (UGT), to the intraluminal side of the endoplasmic reticulum (ER) is an essential step in the glucuronidation of exogenous and endogenous compounds. According to a previous study, the expression of recombinant SLC35B1, SLC35B4, or SLC35D1, nucleotide sugar transporters, in V79 cells has the potential to transport UDPGA into the lumen of microsomes. The purpose of this study is to examine whether the transport of UDPGA by these transporters substantially affects UGT activity. Since the knockdown of UDP-glucose 6-dehydrogenase, a synthetase of UDPGA, in HEK293 cells stably expressing UGT1A1 (HEK/UGT1A1 cells) resulted in a significant decrease in 4-methylumbelliferone (4-MU) glucuronosyltransferase activity, supplementation of a sufficient amount of UDPGA is required for UGT activity. By performing qRT-PCR using cDNA samples from 21 human liver samples, we observed levels of the SLC35B1 and SLC35D1 mRNAs that were 15- and 14-fold higher, respectively, than the levels of the SLC35B4 mRNA, and SLC35B1 showed the largest (37-fold) interindividual variability. Interestingly, 4-MU glucuronosyltransferase activity was significantly decreased upon the knockdown of SLC35B1 in HEK/UGT1A1 cells, and this phenomenon was also observed in HepaRG cells. Using siRNAs targeting 23 different SLC35 subfamilies, the knockdown of SLC35B1 and SLC35E3 decreased 4-MU glucuronosyltransferase activity in HEK/UGT1A1 cells. However, the 4-MU glucuronosyltransferase activity was not altered by SLC35E3 knockdown in HepaRG cells, suggesting that SLC35B1 was the main transporter of UDPGA into the ER in the human liver. In conclusion, SLC35B1 is a key modulator of UGT activity by transporting UDPGA to the intraluminal side of the ER.