Src1 is a Protein of the Inner Nuclear Membrane Interacting with the Dictyostelium Lamin NE81
Src1 is a Protein of the Inner Nuclear Membrane Interacting with the Dictyostelium Lamin NE81
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DOI：
10.3390/cells5010013
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发表时间：
2016-03-01
期刊：
影响因子：
6
通讯作者：
Graef, Ralph
中科院分区：
文献类型：
--
作者：
Batsios, Petros;Ren, Xiang;Graef, Ralph
核被膜（NE）由外核膜（INM）和内核膜（INM）组成，后者与核纤层结合。Src 1是螺旋-延伸-螺旋蛋白家族的盘基网柄菌同源物，该蛋白家族还包括人类层粘连蛋白结合蛋白MAN 1。内源性Src 1和GFP-Src 1在整个细胞周期中都定位于NE。免疫电子显微镜和光学显微镜后，差异洗涤剂处理表明，Src 1驻留在INM。用GFP-Src 1细胞进行的FRAP实验表明，至少一部分蛋白质可以与网骨藻核纤层蛋白NE 81一起稳定地参与形成核纤层。BioID邻近测定和由GFP-NE 81的故意错误定位突变体组成的胞质簇中可溶性截短mRFP-Src 1的错误定位证实了Src 1和NE 81的相互作用。表达GFP-Src 11 -646，一个片段C-末端截短后的第一个跨膜结构域，破坏核膜与核纤层的相互作用，细胞形成的NE依赖于细胞骨架拉力的突起。突起依赖于完整的微管，但不是肌动蛋白丝。我们的研究结果表明，Src 1所需的NE的完整性和突出Dictyosteoblasts作为一个有前途的模型的核结构的演变。
The nuclear envelope (NE) consists of the outer and inner nuclear membrane (INM), whereby the latter is bound to the nuclear lamina. Src1 is a Dictyostelium homologue of the helix-extension-helix family of proteins, which also includes the human lamin-binding protein MAN1. Both endogenous Src1 and GFP-Src1 are localized to the NE during the entire cell cycle. Immuno-electron microscopy and light microscopy after differential detergent treatment indicated that Src1 resides in the INM. FRAP experiments with GFP-Src1 cells suggested that at least a fraction of the protein could be stably engaged in forming the nuclear lamina together with the Dictyostelium lamin NE81. Both a BioID proximity assay and mis-localization of soluble, truncated mRFP-Src1 at cytosolic clusters consisting of an intentionally mis-localized mutant of GFP-NE81 confirmed an interaction of Src1 and NE81. Expression GFP-Src11-646, a fragment C-terminally truncated after the first transmembrane domain, disrupted interaction of nuclear membranes with the nuclear lamina, as cells formed protrusions of the NE that were dependent on cytoskeletal pulling forces. Protrusions were dependent on intact microtubules but not actin filaments. Our results indicate that Src1 is required for integrity of the NE and highlight Dictyostelium as a promising model for the evolution of nuclear architecture.