Ypt1 and Sec4 are essential Rab GTPases that control the early and late stages of the yeast secretory pathway, respectively. A chimera consisting of Ypt1 with the switch I domain of Sec4, Ypt1-SW1^Sec4, is efficiently activated in vitro by the Sec4 exchange factor, Sec2. This should lead to its ectopic activation in vivo and thereby disrupt membrane traffic. Nonetheless early studies found that yeast expressing Ypt1-SW1^Sec4 as the sole copy of YPT1 exhibit no growth defect. To resolve this conundrum, we have analyzed yeast expressing various levels of Ypt1-SW1^Sec4. We show that even normal expression of Ypt1-SW1^Sec4 leads to kinetic transport defects at a late stage of the pathway, with secretory vesicles accumulating near exocytic sites. Higher levels are toxic. Toxicity is suppressed by truncation of Uso1, a vesicle tether required for endoplasmic reticulum–Golgi traffic. The globular head of Uso1 binds to Ypt1 and its coiled-coil tail binds to the Golgi-associated SNARE, Sed5. We propose that when Uso1 is inappropriately recruited to secretory vesicles by Ypt1-SW1^Sec4, the extended coiled-coil tail blocks docking to the plasma membrane. This putative inhibitory function could serve to increase the fidelity of vesicle docking.

中文翻译：

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改造Rab调控网络揭示了囊泡系索Uso1的可能抑制作用

Ypt1和Sec4是必不可少的Rab GTPases，分别控制酵母分泌途径的早期和晚期。由Ypt1和Sec4的开关I结构域Ypt1-SW1 ^Sec4组成的嵌合体在体外被Sec4交换因子Sec2有效激活。这应该导致其在体内的异位激活，从而破坏膜的运输。但是，早期研究发现，表达Ypt1-SW1 ^Sec4作为^YPT1唯一拷贝的酵母没有生长缺陷。为了解决这个难题，我们分析了表达各种水平的Ypt1-SW1 ^{Sec4的酵母}。我们证明Ypt1-SW1 ^Sec4的正常表达在通路的后期导致动力学转运缺陷，分泌性小泡在胞外位点附近积聚。较高的含量是有毒的。截短Uso1可以抑制毒性，Uso1是内质网-高尔基体运输所必需的囊泡系链。Uso1的球形头部与Ypt1结合，其盘绕线圈的尾部与高尔基相关的SNARE Sed5结合。我们建议当Uso1不适当地被Ypt1-SW1 ^Sec4募集到分泌小泡时，扩展的卷曲螺旋尾巴阻隔了质膜。这种推定的抑制功能可用于增加囊泡对接的保真度。