EARLY EXPRESSION OF THE GENE FOR INTERPHOTORECEPTOR RETINOL-BINDING PROTEIN DURING PHOTORECEPTOR DIFFERENTIATION SUGGESTS A CRITICAL ROLE FOR THE INTERPHOTORECEPTOR MATRIX IN RETINAL DEVELOPMENT
EARLY EXPRESSION OF THE GENE FOR INTERPHOTORECEPTOR RETINOL-BINDING PROTEIN DURING PHOTORECEPTOR DIFFERENTIATION SUGGESTS A CRITICAL ROLE FOR THE INTERPHOTORECEPTOR MATRIX IN RETINAL DEVELOPMENT
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DOI：
10.1083/jcb.111.6.2775
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发表时间：
1990-12-01
影响因子：
7.8
通讯作者：
HEALY, JI
中科院分区：
文献类型：
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作者：
GONZALEZFERNANDEZ, F;HEALY, JI
光受体视黄醇结合蛋白(IRBP)是视网膜下间隙的主要蛋白质组分，在调节视网膜色素上皮(RPE)和神经视网膜之间的细胞相互作用中处于重要地位。虽然IRBP似乎参与了成年人视觉周期中维生素A的运输，但这种蛋白在眼睛发育过程中的作用尚未确定。作为了解IRBP在视网膜发育中作用的第一步，我们研究了这种糖脂蛋白在大鼠光感受器分化过程中的mRNA表达。建立了大鼠视网膜神经DNA文库，并从文库中分离到一个IRBP克隆。该克隆含有一个开放阅读框和一个以10个腺苷残基结尾的3‘非编码序列。该编码区与人和牛IRBP的核苷酸序列同源性分别为83.9%和82.5%。大鼠(Spraogue-Dawley、Wistar和皇家外科医师学院粉眼对照)有6.4和5.2 kb的IRBP信使核糖核酸，它们以1：4的比例存在，因此是唯一已知的确定具有不止一种主要形式的IRBP信息的脊椎动物。基因组Southern blotts与IRBP基因只有一个等位基因的假设一致，表明这两种形式是通过对mRNA的交替处理而产生的。为了制备用于分子滴定分析和Northern杂交的反义RNA探针，将编码区的Eco RI-Bam HI片段亚克隆到Sp6和T7启动子之间。总RNA是从出生后P0-P22天未解剖的大鼠球体中提取的。Northern印迹法检测IRBP基因的表达，溶液杂交法检测转录本的表达水平。出生时大约有10(5)个IRBP mRNA转录本/mU-g眼睛总RNA存在。这增加到p9的最终水平3.1X10(6)转录本/mU-g总RNA。在视网膜下间隙IRBP的一半最高水平出现在p4.2，即在视网膜下间隙达到一半最大水平之前2周(Gonzalez-Fernandez，F.，R.A.Landers，P.A.Glazebrook，S.-L.Fong，G.I.Liou，D.M.K.Lam，C.D.B.Bridges)。1984年。J.细胞生物学。99：2092-2098)。IRBP基因的表达反映了作为视网膜内重要结构的光感受器间基质的发育过程。IRBP的绝对水平与视网膜下间隙的体积有关，在发育过程中，由于外节的延长，视网膜下间隙会扩张。IRBP在视网膜发育过程中出人意料的早期表达表明，光感受器间基质不仅在视觉周期中对成年人很重要，而且在视网膜发育过程中也发挥着关键作用，可能有助于疏水性营养物质和形态原物从RPE脉络膜复合体扩散到生长和分化的神经视网膜。
Interphotoreceptor retinol-binding protein (IRBP), the major protein component of the subretinal space, is in a strategic position to mediate cellular interactions between the retinal pigmented epithelium (RPE) and the neural retina. While IRBP appears to be involved in vitamin A transport during the visual cycle in the adult, the role of this protein during eye development has not been determined. As a first step to understanding the role of IRBP during the retinal development, we have studied the expression of the mRNA for this glycolipoprotein during photoreceptor differentiation in the rat. A rat neural retina cDNA library was prepared from which an IRBP clone was isolated. The clone contains an open reading frame followed by a 3' noncoding sequence ending in 10 adenosine residues. The coding region has an identity of 83.9 and 82.5% with the nucleotide sequence of human and bovine IRBP, respectively. Rats (Sprague-Dawley, Wistar, and Royal College of Surgeon pink-eyed controls) have a 6.4 and 5.2-kb mRNA for IRBP which are present in a 1:4 ratio and thus are the only vertebrate known to definitely have more than one major form of the IRBP message. Genomic Southern blots are consistent with the hypothesis that there is only one allele of the IRBP gene, suggesting that the two forms are produced by alternative processing of the mRNA. To generate an antisense RNA probe for use in molecular titration assays and Northern blots, an Eco RI-Bam HI fragment from the coding region was subcloned in between flanking Sp6 and T7 promoters. Total RNA was prepared from undissected rat globes from postnatal days p0-p22. The expression of the mRNA for IRBP was studied by Northern blots and the level of the transcripts determined by solution hybridization assays. Approximately 10(5) IRBP mRNA transcripts/mu-g total eye RNA are present at birth. This increases to a final level of 3.1 X 10(6) transcripts/mu-g total RNA by p9. The one-half maximal level of the mRNA occurs at p4.2 which is 2 wk before the one-half maximal level of IRBP is reached in the subretinal space (Gonzalez-Fernandez, F., R. A. Landers, P. A. Glazebrook, S.-L. Fong, G. I. Liou, D. M. K. Lam, and C. D. B. Bridges. 1984. J. Cell Biol. 99:2092-2098). The expression of the mRNA for IRBP reflects the developmental emergence of the interphotoreceptor matrix as an important structure within the retina. The absolute level of IRBP is related to the volume of the subretinal space, which expands during development due to the elongation of the outer segments. The unexpected early expression of the mRNA for IRBP during retinal development suggests that the interphotoreceptor matrix is important not only to the adult during the visual cycle, but also plays a critical role during retinal development, perhaps facilitating the diffusion of hydrophobic nutrients and morphogens such as vitamin A from the RPE-choroid complex to the growing and differentiating neural retina.