Molecular analysis of alphaB-crystallin in central nervous system and in pathologic conditions.

α-晶体蛋白是一种主要的晶状体蛋白，存在于中枢神经系统(CNS)中，是星形胶质细胞胞浆内包涵体Rosenthal纤维(RF)的主要蛋白质成分。它在正常中枢神经系统中的表达水平很低，似乎仅限于胶质细胞，包括星形胶质细胞和少突胶质细胞。用抗α-晶体蛋白抗血清免疫组织化学方法观察了大量表现为多种病理改变的人脑组织，发现星形胶质细胞和少突胶质细胞的免疫反应增强，但未见RFs的形成。此外，还用抗α-晶体蛋白抗血清对神经退行性疾病的部分神经元进行了免疫标记。因此，αB-晶体蛋白的积聚似乎是CNS胶质细胞和一些神经元在病理条件下反应过程的一部分。我们还测定了αB-晶体蛋白mRNAs和基因组结构。α-晶体蛋白的两类主要的mRNAs在大鼠脑中表达，它们的5‘前导序列长度不同，长度分别约为0.9kb和1.2kb。较长的mRNAs的转录起始点之前有一个推测的CAAT盒，而较短的mRNAs的转录起始点之前有一个推测的TATA盒。由于α-晶体蛋白基因只有一个拷贝，所以这两类mRNAs是由不同启动子交替转录启动而产生的，它们的表达受到不同的调控。接下来，我们检查了两例经病理证实的亚历山大病患者的α-晶体蛋白的初级结构。用聚合酶链式反应扩增冰冻脑组织基因组DNA并测序。对两名患者的α-晶体蛋白基因启动子和编码区进行了测序，发现它们属于正常序列，这表明α-晶体蛋白在这些大脑中的积累并不是由于蛋白质一级结构的任何异常。

AlphaB-crystallin, a major lens protein, is found in the central nervous system (CNS) and is a major protein component of Rosenthal fibers (RF), intracytoplasmic inclusions within astrocytes. Its level of expression in the normal CNS is low and appear to be confined to glial cells, both astrocytes and oligodendrocytes. A number of human brains displaying a variety of pathologic changes were examine by immunohistochemistry with an anti-alphaB-crystallin antiserum and increased immunoreactivity was found in astrocytes and oligodendrocytes without the formation of RFs. Furthermore, some neurons in neurodegenerative disorders were also immunolabeled with anti-alphaB-crystallin antiserum. Thus, the accumulation of Alpha B-crystallin appears to be part of the repertoire of reactive processes of CNS glial cells and some neurons in pathologic conditions.We also determined the structures of alphaB-crystallin mRNAs and the genomic structure. Two major classes of mRNAs for the alphaB-crystallin, about 0.9 and 1.2 kilobases in length, are expressed in rat brain and they differ in the lengths of their 5' leader sequences. The transcriptional start sites of the longer mRNAs are preceded by a putative CAAT box and that of the shorter mRNA by a putative TATA box. Since there is only a single copy of the alphaB-crystallin gene, the two classes of mRNAs are generated by alternative transcriptional initiation from different promoters and their expressions are regulated differentially. Next, we examined primary structures of alphaB-crystallin in two cases of pathologically confirmed Alexander's disease. The genomic DNAs from frozen brain tissues were amplified by polymerase chain reaction and sequenced. Sequencing of the promoter and coding regions of the alphaB-crystallin genes in two patients revealed them to be of normal sequence, suggesting the accumulation of alphaB-crystallin in those brains is not due to any abnormality of the primary structure of the protein.