Fairweather Stephen J, Okada Shoko, Gauthier-Coles Gregory, Javed Kiran, Bröer Angelika, Bröer Stefan
Research School of Biology, Australian National University, Canberra, ACT, Australia.
Research School of Chemistry, Australian National University, Canberra, ACT, Australia.
Front Mol Biosci. 2021 Apr 13;8:646574. doi: 10.3389/fmolb.2021.646574. eCollection 2021.
Amino acid transporters play a vital role in metabolism and nutrient signaling pathways. Typically, transport activity is investigated using single substrates and competing amounts of other amino acids. We used GC-MS and LC-MS for metabolic screening of oocytes expressing various human amino acid transporters incubated in complex media to establish their comprehensive substrate profiles. For most transporters, amino acid selectivity matched reported substrate profiles. However, we could not detect substantial accumulation of cationic amino acids by SNAT4 and ATB in contrast to previous reports. In addition, comparative substrate profiles of two related sodium neutral amino acid transporters known as SNAT1 and SNAT2, revealed the latter as a significant leucine accumulator. As a consequence, SNAT2, but not SNAT1, was shown to be an effective activator of the eukaryotic cellular growth regulator mTORC1. We propose, that metabolomic profiling of membrane transporters in oocytes can be used to test their substrate specificity and role in intracellular signaling pathways.
氨基酸转运蛋白在新陈代谢和营养信号通路中起着至关重要的作用。通常，利用单一底物以及其他氨基酸的竞争量来研究转运活性。我们使用气相色谱-质谱联用仪（GC-MS）和液相色谱-质谱联用仪（LC-MS）对在复杂培养基中孵育的表达各种人类氨基酸转运蛋白的卵母细胞进行代谢筛选，以确定它们的全面底物谱。对于大多数转运蛋白而言，氨基酸选择性与报道的底物谱相符。然而，与之前的报道相反，我们无法检测到SNAT4和ATB对阳离子氨基酸的大量积累。此外，对两种相关的钠中性氨基酸转运蛋白SNAT1和SNAT2的比较底物谱显示，后者是亮氨酸的重要积累者。因此，SNAT2而非SNAT1被证明是真核细胞生长调节因子mTORC1的有效激活剂。我们提出，卵母细胞膜转运蛋白的代谢组学分析可用于测试它们的底物特异性以及在细胞内信号通路中的作用。
