Interaction with Btn2p Is Required for Localization of Rsg1p: Btn2p-Mediated Changes in Arginine Uptake in Saccharomyces cerevisiae

Btn2p, a novel coiled-coil protein, is up-regulated in btn1Δ yeast strains, and this up-regulation is thought to contribute to maintaining a stable vacuolar pH in btn1Δ strains (D. A. Pearce, T. Ferea, S. A. Nosel, B. Das, and F. Sherman, Nat. Genet. 22:55-58, 1999). We now report that Btn2p interacts biochemically and functionally with Rsg1p, a down-regulator of the Can1p arginine and lysine permease. Rsg1p localizes to a distinct structure toward the cell periphery, and strains lacking Btn2p (btn2Δ strains) fail to correctly localize Rsg1p. btn2Δ strains, like rsg1Δ strains, are sensitive for growth in the presence of the arginine analog canavanine. Furthermore, btn2Δ strains, like rsg1Δ strains, demonstrate an elevated rate of uptake of [(14)C]arginine, which leads to increased intracellular levels of arginine. Overexpression of BTN2 results in a decreased rate of arginine uptake. Collectively, these results indicate that altered levels of Btn2p can modulate arginine uptake through localization of the Can1p-arginine permease regulatory protein, Rsg1p. Our original identification of Btn2p was that it is up-regulated in the btn1Δ strain which serves as a model for the lysosomal storage disorder Batten disease. Btn1p is a vacuolar/lysosomal membrane protein, and btn1Δ suppresses both the canavanine sensitivity and the elevated rate of uptake of arginine displayed by btn2Δ rsg1Δ strains. We conclude that Btn2p interacts with Rsg1p and modulates arginine uptake. Up-regulation of BTN2 expression in btn1Δ strains may facilitate either a direct or indirect effect on intracellular arginine levels.

Btn2p是一种新型卷曲螺旋蛋白（coiled-coil protein），在btn1Δ酵母菌株中表达上调，且该上调被认为有助于维持btn1Δ菌株的稳定液泡pH（D. A. Pearce、T. Ferea、S. A. Nosel、B. Das及F. Sherman，《自然·遗传学》，22卷：55-58，1999年）。本研究首次报道，Btn2p可与Can1p型精氨酸赖氨酸通透酶的负调控因子Rsg1p发生生化与功能层面的相互作用。Rsg1p定位于细胞外周的特定结构，而缺失Btn2p的菌株（btn2Δ菌株）无法正确定位Rsg1p。与rsg1Δ菌株类似，btn2Δ菌株在精氨酸类似物刀豆氨酸（canavanine）存在时生长受抑制。此外，与rsg1Δ菌株类似，btn2Δ菌株的[14C]精氨酸摄取速率升高，进而导致细胞内精氨酸水平上升。过表达BTN2可降低精氨酸摄取速率。综合上述结果可知，Btn2p水平的改变可通过调控Can1p型精氨酸赖氨酸通透酶的调控蛋白Rsg1p的定位，来调节精氨酸摄取过程。我们最初对Btn2p的鉴定显示，其在btn1Δ菌株中表达上调，而该菌株可作为溶酶体贮积症巴顿病（Batten disease）的模型。Btn1p是一种液泡/溶酶体膜蛋白，且btn1Δ可抑制btn2Δ rsg1Δ菌株所表现出的刀豆氨酸敏感性与升高的精氨酸摄取速率。综上，我们得出结论：Btn2p与Rsg1p相互作用并调控精氨酸摄取。在btn1Δ菌株中BTN2表达上调，可能对细胞内精氨酸水平产生直接或间接的影响。