Thomas G, Zelenka P S, Cuthbertson R A, Norman B L, Piatigorsky J
Laboratory of Molecular and Developmental Biology, National Eye Institute, National Institutes of Health, Bethesda, MD 20892.
New Biol. 1990 Oct;2(10):903-14.
Chicken delta-crystallin/argininosuccinate lyase (ASL), a major enzyme-crystallin of the embryonic lens, is encoded by two similar, tandemly arranged genes (delta 1 and delta 2). We show here by the polymerase chain reaction (PCR) and in situ hybridization that although the amount of mRNA for each of the delta-crystallins increases in the lens epithelial and fiber cells during development of the embryonic chicken, the delta 1 mRNA accumulates preferentially in the fiber cells. The delta 1/delta 2 mRNA ratio actually decreased from 16.5 +/- 7 to 6.5 +/- 1 in the central epithelial cells while it increased from 20 +/- 10 to 95 +/- 5 in the fibers between 6 and 14 days of development. By contrast, the heart and brain of 4- to 8-day-old embryonic chickens showed 10(3) to 10(4) times less delta-crystallin mRNA per microgram of total RNA than the lens, with a delta 1/delta 2 mRNA ratio of only 0.20 to 0.30. The tissue-specific differences in the relative expression of the two delta-crystallin genes suggest that the delta 2 polypeptide is principally responsible for ASL activity and that the delta 1 polypeptide is specialized for lens transparency. The trace amounts of delta 1 mRNA in the heart and brain raise the possibility that the delta 1 polypeptide contributes to or modulates ASL activity of the native tetrameric protein. Transfection experiments in which we used the pSVOCAT plasmid demonstrated that both delta-crystallin genes contain an enhancer in their third intron. The promoter and enhancer of each delta-crystallin gene were functionally comparable in the pSVOCAT plasmid when tested in primary embryonic lens epithelial cells, suggesting that the differential expression of the two delta-crystallin genes in the lens requires additional cis-regulatory sequences, post-transcriptional mechanisms, or both.
鸡δ-晶体蛋白/精氨琥珀酸裂解酶（ASL）是胚胎晶状体的一种主要酶晶体蛋白，由两个相似的、串联排列的基因（δ1和δ2）编码。我们通过聚合酶链反应（PCR）和原位杂交在此表明，尽管在胚胎鸡发育过程中，晶状体上皮细胞和纤维细胞中每种δ-晶体蛋白的mRNA量都会增加，但δ1 mRNA优先在纤维细胞中积累。在发育的6至14天期间，中央上皮细胞中δ1/δ2 mRNA的比例实际上从16.5±7降至6.5±1，而在纤维细胞中则从20±10增至95±5。相比之下，4至8日龄胚胎鸡的心脏和大脑每微克总RNA中的δ-晶体蛋白mRNA比晶状体少10³至10⁴倍，δ1/δ2 mRNA的比例仅为0.20至0.30。两个δ-晶体蛋白基因相对表达的组织特异性差异表明，δ2多肽主要负责ASL活性，而δ1多肽专门用于晶状体的透明度。心脏和大脑中微量的δ1 mRNA增加了δ1多肽对天然四聚体蛋白的ASL活性有贡献或调节作用的可能性。我们使用pSVOCAT质粒进行的转染实验表明，两个δ-晶体蛋白基因在其第三个内含子中都含有一个增强子。当在原代胚胎晶状体上皮细胞中进行测试时，每个δ-晶体蛋白基因的启动子和增强子在pSVOCAT质粒中的功能相当，这表明晶状体中两个δ-晶体蛋白基因的差异表达需要额外的顺式调控序列、转录后机制或两者兼而有之。
