mRNA analysis of α- and β-synemin and study of protein involving with α-synemin in brain

联蛋白是一种肌肉中间纤维蛋白，最初在鸟类中发现。最近，人类α-和β-突触蛋白同源物已被克隆，后者以前被称为人类真丝蛋白。由于外显子4和外显子5的不同剪接，两种人类synemin同种异构体源自同一基因，因此β-synemin蛋白的c端短了312个氨基酸。β-突触蛋白mRNA和蛋白在骨骼肌和心肌中均有高表达，而在脑组织中northern blot分析显示为弱双链，说明在mRNA水平上，脑组织中存在两种不同类型的突触蛋白。我们认为β-突触蛋白在维持骨骼肌细胞骨架结构中起作用，而α-突触蛋白也参与神经元和胶质细胞形态的维持。我们认为，阐明与α-突触蛋白有关的脑内蛋白具有重要意义。因此，我们决定利用Matchmaker Two-hybrid系统3，从人脑cDNA文库中寻找一个与α-synemin特异区域相互作用的新序列，共312个氨基酸序列。去年，我们利用原位杂交技术检测了α-和β-突触蛋白mRNA的表达，发现突触蛋白mRNA在脑桥背侧和中脑的大神经元中均有表达。今年，我们通过原位杂交和免疫组织化学分析进一步认识到synemin mRNA及其蛋白在同一大神经元中共定位。

Synemin is a muscle intermediate filament protein that was originally identified in birds. Recently, the human α- and β-synemin orthologues have been cloned, the latter of which was previously termed human desmuslin. Both human synemin isoforms derive from the same gene as a result of differential splicing between exons 4 and 5 such that the β-synemin protein is 312 amino acids shorter at its C-terminus. Both β-synemin mRNA and protein are highly expressed in skeletal and cardiac muscles, while northern blot analysis shows weak doublet in brain, indicating that there are two different types of synemin present in brain on the basis of mRNA level. We consider that β-synemin plays a role in maintaining the cytoskeletal structure in skeletal muscle, while α-synemin is also involving to role of the morphological maintenance of neuron and glia. We think that it is of great importance to elucidate the proteins involving α-synemin in brain. Therefore, we decided to examine human brain cDNA library to find a new sequence that interacts with the specific region to α-synemin, 312 amino acids sequences, using Matchmaker Two-hybrid system 3.Last year, we examined the expression of α- and β-synemin mRNA and found that both synemin mRNAs are expressed in the large neurons of dorsal side of pons and midbrain using in situ hybridization. This year, we further recognized that both synemin mRNA and its protein are co-localized in the same large neurons by in situ hybridization and immunohistochemical analysis.