Keenan K A, Weiss R L
Department of Chemistry and Biochemistry, University of California, Los Angeles, Los Angeles, California 90095-1569, USA.
Fungal Genet Biol. 1997 Dec;22(3):177-90. doi: 10.1006/fgbi.1997.1009.
Treatment of Neurospora crassa mycelia with cupric ion has been shown to permeabilize the plasma and mitochondrial membranes. Permeabilized mycelia were shown to take up arginine into the vacuoles. Uptake was ATP-independent and appeared to be driven by an existing K+-gradient. The kinetic characteristics of the observed uptake were similar to those observed using vacuolar membrane vesicles: the Km for arginine uptake was found to be 4.2-4.5 mM. Permeabilized mycelia were used to study the regulation of arginine uptake into vacuoles. The results suggest that uptake is relatively indifferent to the contents of the vacuoles and is not affected by growth of mycelia in amino acid-supplemented medium. Efflux of arginine, lysine, and ornithine from vacuoles was also measured using mycelia permeabilized with cupric ion. Arginine release was shown to be specifically enhanced by cytosolic ornithine and/or increases in the vacuolar pool of arginine or ornithine. Lysine efflux was shown be indifferent to the presence of other amino acids. These observations emphasize the importance of vacuolar compartmentation in controlling arginine and ornithine metabolism and suggest that vacuolar compartmentation may play an important role in nitrogen homeostasis of filamentous fungi.
已证明用铜离子处理粗糙脉孢菌菌丝体会使质膜和线粒体膜通透性增加。通透性增加的菌丝体被证明能将精氨酸吸收到液泡中。吸收过程不依赖ATP，似乎是由现有的钾离子梯度驱动的。观察到的吸收的动力学特征与使用液泡膜囊泡观察到的相似：精氨酸吸收的米氏常数为4.2 - 4.5 mM。通透性增加的菌丝体被用于研究精氨酸向液泡内吸收的调节。结果表明，吸收对液泡内容物相对不敏感，且不受在补充氨基酸的培养基中菌丝体生长的影响。还使用用铜离子通透化的菌丝体测量了精氨酸、赖氨酸和鸟氨酸从液泡中的流出。结果表明，胞质中的鸟氨酸和/或液泡中精氨酸或鸟氨酸池的增加会特异性增强精氨酸的释放。赖氨酸的流出对其他氨基酸的存在不敏感。这些观察结果强调了液泡区室化在控制精氨酸和鸟氨酸代谢中的重要性，并表明液泡区室化可能在丝状真菌的氮稳态中起重要作用。
