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

合成素是一种肌肉中间丝蛋白，最初在鸟类中发现。最近，人类的α-和β-Synmin同源基因已经被克隆，后者以前被称为人去黏蛋白。由于外显子4和5之间的差异剪接，这两种人类同步蛋白亚型都来自同一个基因，因此β-同步蛋白的C末端短了312个氨基酸。β同步素在骨骼肌和心肌中均有高表达，而Northern印迹分析显示在脑中有微弱的二倍体表达，这表明在基因水平上大脑中存在两种不同类型的同步素。我们认为β同步素在维持骨骼肌细胞骨架结构中起重要作用，而α同步素还参与神经元和神经胶质细胞的形态维持。我们认为，阐明脑内涉及α-SYMIN的蛋白质具有重要意义。因此，我们决定利用Matchaker双杂交系统3对人脑cDNA文库进行检测，寻找与α同步素的特异区相互作用的新序列，共312个氨基酸序列。去年，我们用原位杂交的方法检测了α-和β-同步素基因的表达，发现这两种同步素在脑桥背侧和中脑的大神经元中都有表达。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，在相同的大型神经元中，同步蛋白及其蛋白是共定位的。

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.