Bankaitis V A, Malehorn D E, Emr S D, Greene R
Department of Microbiology, University of Illinois, Urbana-Champaign 61801.
J Cell Biol. 1989 Apr;108(4):1271-81. doi: 10.1083/jcb.108.4.1271.
We have obtained and characterized a genomic clone of SEC14, a Saccharomyces cerevisiae gene whose product is required for export of yeast secretory proteins from the Golgi complex. Gene disruption experiments indicated that SEC14 is an essential gene for yeast vegetative growth. Nucleotide sequence analysis revealed the presence of an intron within the SEC14 structural gene, and predicted the synthesis of a hydrophilic polypeptide of 35 kD in molecular mass. In confirmation, immunoprecipitation experiments demonstrated SEC14p to be an unglycosylated polypeptide, with an apparent molecular mass of some 37 kD, that behaved predominantly as a cytosolic protein in subcellular fractionation experiments. These data were consistent with the notion that SEC14p is a cytosolic factor that promotes protein export from yeast Golgi. Additional radiolabeling experiments also revealed the presence of SEC14p-related polypeptides in extracts prepared from the yeasts Kluyveromyces lactis and Schizosaccharomyces pombe. Furthermore, the K. lactis SEC14p was able to functionally complement S. cerevisiae sec14ts defects. These data suggested a degree of conservation of SEC14p structure and function in these yeasts species.
我们已获得酿酒酵母基因SEC14的基因组克隆并对其进行了表征,该基因的产物是酵母分泌蛋白从高尔基体复合体输出所必需的。基因破坏实验表明,SEC14是酵母营养生长所必需的基因。核苷酸序列分析显示,SEC14结构基因内存在一个内含子,并预测合成一种分子量为35kD的亲水性多肽。经证实,免疫沉淀实验表明SEC14p是一种未糖基化的多肽,表观分子量约为37kD,在亚细胞分级实验中主要表现为胞质蛋白。这些数据与SEC14p是促进酵母高尔基体蛋白输出的胞质因子这一观点一致。额外的放射性标记实验还揭示了在乳酸克鲁维酵母和粟酒裂殖酵母提取物中存在与SEC14p相关的多肽。此外,乳酸克鲁维酵母的SEC14p能够在功能上互补酿酒酵母sec14ts缺陷。这些数据表明在这些酵母物种中SEC14p的结构和功能有一定程度的保守性。