Research School of Biology, Australian National Univ., Canberra, ACT 0200, Australia.
Am J Physiol Cell Physiol. 2010 Nov;299(5):C1047-57. doi: 10.1152/ajpcell.00209.2010. Epub 2010 Aug 25.
The glutamine transporter SNAT3 is involved in the uptake and release of glutamine in the brain, liver, and kidney. Substrate transport is accompanied by Na(+) cotransport and H(+) antiport. In this study, treatment of Xenopus laevis oocytes expressing rat SNAT3 with the phorbol ester PMA resulted in a rapid downregulation of glutamine uptake in less than 20 min. PMA treatment of oocytes coexpressing SNAT3 and the monocarboxylate transporter MCT1 reduced SNAT3 activity only, demonstrating the specificity of the regulatory mechanism. Single or combined mutations of seven putative phosphorylation sites in the SNAT3 sequence did not affect the regulation of SNAT3 by PMA. Expression of an EGFP-SNAT3 fusion protein in oocytes established that the downregulation was caused by the retrieval of the transporter from the plasma membrane. Coexpression of SNAT3 with dominant-negative mutants of dynamin or caveolin revealed that SNAT3 trafficking occurs in a dynamin-independent manner and is influenced by caveolin. Although system N activity was not affected by PMA in cultured astrocytes, a downregulation was observed in HepG2 cells.
谷氨酸转运体 SNAT3 参与脑、肝和肾中谷氨酸的摄取和释放。底物转运伴随着 Na(+)共转运和 H(+)反向转运。在这项研究中，用佛波酯 PMA 处理表达大鼠 SNAT3 的非洲爪蟾卵母细胞，导致谷氨酸摄取在不到 20 分钟的时间内迅速下调。共表达 SNAT3 和单羧酸转运体 MCT1 的卵母细胞的 PMA 处理仅降低了 SNAT3 活性，证明了调节机制的特异性。SNAT3 序列中七个假定磷酸化位点的单个或组合突变不影响 PMA 对 SNAT3 的调节。在卵母细胞中表达 EGFP-SNAT3 融合蛋白表明，下调是由于转运蛋白从质膜中回收引起的。与 dynamin 或 caveolin 的显性负突变体共表达表明，SNAT3 转运以 dynamin 非依赖性方式发生，并受 caveolin 影响。尽管 PMA 在培养的星形胶质细胞中对系统 N 活性没有影响，但在 HepG2 细胞中观察到下调。
