Molecular Structure and Function of Crystallins

眼睛的组织依赖于严格调控的发育途径和特化蛋白质家族。这些包括晶状体的晶状体蛋白。我们已经证明，在人类和其他物种中，晶体蛋白是通过一种名为基因招募的过程产生的。许多种类的晶体蛋白的起源和功能已被阐明。我们已经证明GS-晶体蛋白是成人晶状体中主要的BG-晶体蛋白。在小鼠中，GS基因是OPJ白内障的基因座。同源重组，GS基因的“敲除”小鼠已经被创造出来。在缺乏GS的小鼠中，晶状体细胞的组织被破坏，晶状体发育的几个特征发生异常。这在很大程度上与晶状体细胞骨架的组织失败有关，表明GS在正常晶状体发育中起着关键的功能作用。其他的g-晶体蛋白与人类的白内障有关。我们已经为一种导致两种人类浑浊的gD突变体制造了重组蛋白。蛋白质表面的微小变化会导致明显的溶解性损失。结构和生物物理研究正在进行中。我们还在研究g-晶体蛋白家族的一个新成员，gn，它的基因结构结合了b-基因和g-基因家族的特征。GN在小鼠晶状体和视网膜中表达，一种新的抗体在猴子晶状体中检测到预期大小的蛋白质。然而，人类的gn基因序列似乎发生了变化，这表明在哺乳动物进化过程中，功能发生了变化。 在所有被研究的哺乳动物晶状体中，GS和Ba2晶体蛋白在RNA水平上都有丰富的表达。然而，Ba2蛋白在人类晶状体中含量很低。这可能是由于翻译控制。我们正在研究人类Ba2转录本5‘非编码区中发夹环的作用。此外，现在有证据表明，GS和Ba2在晶状体发育中的表达比之前认为的要早得多，可能是应激诱导的。我们已经建立了在GS和Ba2基因启动子控制下表达GFP标记蛋白的细胞系和转基因小鼠。

Tissues of the eye depend on closely regulated developmental pathways and families of specialized proteins. These include the crystallins of the lens. We have shown that in humans and other species, crystallins have arisen by a process called Gene Recruitment. The origins and functions of many classes of crystallins have been elucidated. We have shown that gS-crystallin is the major bg-crystallin in the adult human lens. In mouse, the gS gene is the locus of the Opj cataract. Homologous recombination, "knock-out" mice for the gS gene have been created. In mice lacking gS the organization of lens cells is disrupted and several features of lens development are aberrant. Much of this is connected to a failure of organization of the lens cytoskeleton, suggesting a key functional role for gS in normal lens development. Other g-crystallins are associated with cataract in humans. We have made recombinant protein for a gD mutant that causes two kinds of human opacity. A small surface change in the protein leads to a marked loss of solubility. Structural and biophysical studies are in progress. We are also examining a new member of the g-crystallin family, gN, which has a gene structure that combines features of both b- and g- gene families. gN is expressed in mouse lens and retina and a new antibody detects a protein of the expected size in monkey lens. However there appear to have been changes in gN gene sequence in humans that suggest a change in function during mammalian evolution. Both gS and bA2 crystallins are abundantly expressed at the RNA level in lens in all mammals examined. However bA2 protein is very low in abundance in human lens. This may be due to translational control. We are investigating the effect of a hairpin loop in the 5' UTR of the human bA2 transcript. In addition there is now evidence that both gS and bA2 are expressed much early in lens development than previously thought and may be stress inducible. We have created cell lines and transgenic mice expressing the GFP marker protein under the control of the gS and bA2 gene promoters.