Eugster Anne, Frigerio Gabriella, Dale Martin, Duden Rainer
Department of Clinical Biochemistry, Cambridge Institute for Medical Research, University of Cambridge, Cambridge CB2 2XY, United Kingdom.
Mol Biol Cell. 2004 Mar;15(3):1011-23. doi: 10.1091/mbc.e03-10-0724. Epub 2003 Dec 29.
Coatomer is required for the retrieval of proteins from an early Golgi compartment back to the endoplasmic reticulum. The WD40 domain of alpha-COP is required for the recruitment of KKTN-tagged proteins into coatomer-coated vesicles. However, lack of the domain has only minor effects on growth in yeast. Here, we show that the WD40 domain of beta'-COP is required for the recycling of the KTKLL-tagged Golgi protein Emp47p. The protein is degraded more rapidly in cells with a point mutation in the WD40 domain of beta'-COP (sec27-95) or in cells lacking the domain altogether, whereas a point mutation in the Clathrin Heavy Chain Repeat (sec27-1) does not affect the turnover of Emp47p. Lack of the WD40 domain of beta'-COP has only minor effects on growth of yeast cells; however, absence of both WD40 domains of alpha- and beta'-COP is lethal. Two hybrid studies together with our analysis of the maturation of KKTN-tagged invertase and the turnover of Emp47p in alpha- and beta'-COP mutants suggest that the two WD40 domains of alpha- and beta'-COP bind distinct but overlapping sets of di-lysine signals and hence both contribute to recycling of proteins with di-lysine signals.
COP包被蛋白对于将蛋白质从早期高尔基体区室回收至内质网是必需的。α-COP的WD40结构域对于将KKTN标记的蛋白质招募到COP包被的小泡中是必需的。然而,该结构域的缺失对酵母生长只有轻微影响。在这里,我们表明β'-COP的WD40结构域对于KTKLL标记的高尔基体蛋白Emp47p的循环利用是必需的。在β'-COP的WD40结构域发生点突变的细胞(sec27-95)或完全缺乏该结构域的细胞中,该蛋白降解得更快,而网格蛋白重链重复序列中的点突变(sec27-1)不影响Emp47p的周转。β'-COP的WD40结构域的缺失对酵母细胞生长只有轻微影响;然而,α-COP和β'-COP的两个WD40结构域都缺失是致死的。双杂交研究以及我们对α-COP和β'-COP突变体中KKTN标记的转化酶成熟和Emp47p周转的分析表明,α-COP和β'-COP的两个WD40结构域结合不同但重叠的双赖氨酸信号集,因此两者都有助于具有双赖氨酸信号的蛋白质的循环利用。