Nakajima T, Fukiage C, Azuma M, Ma H, Shearer T R
Research Laboratory, Senju Pharmaceutical Company, Ltd., Kobe, Japan.
Biochim Biophys Acta. 2001 May 28;1519(1-2):55-64. doi: 10.1016/s0167-4781(01)00212-3.
The purpose of the present investigation was to compare the expression of ubiquitous and tissue-specific calpains in ocular tissues from the Macaca fascicularis monkey. Calpain isoforms in retina and corneal epithelium from adult M. fascicularis monkeys were characterized by RT-PCR, cDNA cloning and sequencing. Calpain isoform activities in ocular tissues were investigated by fractionation on DEAE-HPLC, immunoblotting, and casein zymography. Capn3 splice variants in the ocular tissues from rat, rabbit and monkey were compared after RT-PCR. RT-PCR analysis revealed that numerous splice variants of Capn3 were expressed in the epithelium from monkey cornea. The variants contained deletions or insertions in or around the IS1, IS2, and NS regions. The cDNAs for Capn3 variants were highly conserved, yet the expression patterns of the Capn3 isoforms were widely different among the mammalian species. In contrast, the expression patterns of ubiquitous calpains in ocular tissues were conserved among the mammalian species, and similarities between monkey and human cDNAs for Capn1 (mu-calpain) and Capn2 (m-calpain) were 98 and 99%, respectively. These results suggested that differences in expression patterns of Capn3 variants might be related to the function of each variant in a particular tissue or species.
本研究的目的是比较食蟹猴眼组织中普遍存在的钙蛋白酶和组织特异性钙蛋白酶的表达情况。通过逆转录聚合酶链反应（RT-PCR）、cDNA克隆和测序对成年食蟹猴视网膜和角膜上皮中的钙蛋白酶同工型进行了表征。通过DEAE-高效液相色谱法（DEAE-HPLC）分级分离、免疫印迹和酪蛋白酶谱法研究了眼组织中的钙蛋白酶同工型活性。在RT-PCR后比较了大鼠、兔子和猴子眼组织中的Capn3剪接变体。RT-PCR分析显示，Capn3的许多剪接变体在猴角膜上皮中表达。这些变体在IS1、IS2和NS区域或其周围存在缺失或插入。Capn3变体的cDNA高度保守，但Capn3同工型的表达模式在哺乳动物物种之间差异很大。相比之下，眼组织中普遍存在的钙蛋白酶的表达模式在哺乳动物物种之间是保守的，并且猴和人Capn1（μ-钙蛋白酶）和Capn2（m-钙蛋白酶）的cDNA之间的相似性分别为98%和99%。这些结果表明，Capn3变体表达模式的差异可能与每个变体在特定组织或物种中的功能有关。
