Golgi localization of Syne-1
Golgi localization of Syne-1
复制标题
DOI：
10.1091/mbc.e02-07-0446
复制
发表时间：
2003-06-01
影响因子：
3.3
通讯作者：
Beck, KA
中科院分区：
文献类型：
--
作者：
Gough, LL;Fan, J;Beck, KA
我们以前已经确定了高尔基体本地血影蛋白异构体通过使用抗体的红细胞血影蛋白的β-亚基。在这项研究中，我们表明，筛选的一个residagt 11表达文库中分离出一个类似于5-kb的部分cDNA从Madin-Darby牛肾（MDBK）细胞系，它编码一个多肽的1697个氨基酸的低，但可检测到，序列同源性血影蛋白（37%）。blast搜索显示，该克隆与最近鉴定的血影蛋白家族成员Syne-1B/Nesprin-1 β的5'端重叠，该基因是一种交替转录的基因，具有结合乙酰胆碱受体并与核膜相关的肌肉特异性形式。通过将MDBK克隆的序列与人类基因组数据库的序列数据进行比较，我们确定该cDNA代表了一个非常大的基因（约500 kb）的中心部分，编码一个约25 kb的转录物，我们称之为Syne-1。Syne-1编码一种大多肽（8406个氨基酸），具有多个血影蛋白重复序列，其氨基末端的一个区域与传统血影蛋白的肌动蛋白结合结构域具有高度同源性。高尔基体定位的血影蛋白样蛋白表现为表位标记的片段在MDBK和COS细胞，确定两个不同的高尔基体结合位点，并通过免疫荧光显微镜使用几种不同的抗体制剂。Syne-1上的高尔基体结合结构域之一作为显性负抑制剂，改变高尔基体复合体的结构，其在转染的上皮细胞中在中心体附近塌陷成浓缩结构。细胞我们的结论是，Syne-1，基因，表达在多种形式，是多功能的，能够在高尔基体和核膜的功能，也许连接两个细胞器在肌肉分化。
We have previously identified a Golgi-localized spectrin isoform by using an anti-body to the beta-subunit of erythrocyte spectrin. In this study, we show that a screen of a lambdagt11 expression library resulted in the isolation of an similar to5-kb partial cDNA from a Madin-Darby bovine kidney (MDBK) cell line, which encoded a polypeptide of 1697 amino acids with low, but detectable, sequence homology to spectrin (37%). A blast search revealed that this clone overlaps with the 5' end of a recently identified spectrin family member Syne-1B/Nesprin-1beta, an alternately transcribed gene with muscle-specific forms that bind acetylcholine receptor and associate with the nuclear envelope. By comparing the sequence of the MDBK clone with sequence data from the human genome database, we have determined that this cDNA represents a central portion of a very large gene (similar to500 kb), encoding an similar to25-kb transcript that we refer to as Syne-1. Syne-1 encodes a large polypeptide (8406 amino acids) with multiple spectrin repeats and a region at its amino terminus with high homology to the actin binding domains of conventional spectrins. Golgi localization for this spectrin-like protein was demonstrated by expression of epitope-tagged fragments in MDBK and COS cells, identifying two distinct Golgi binding sites, and by immunofluorescence microscopy by using several different antibody preparations. One of the Golgi binding domains on Syne-1 acts as a dominant negative inhibitor that alters the structure of the Golgi complex, which collapses into a condensed structure near the centrosome in transfected epithelial. cells. We conclude that the Syne-1, gene is, expressed in a variety of forms that are multifunctional and are capable of functioning at both the Golgi and the nuclear envelope, perhaps linking the two organelles during muscle differentiation.