Overexpression of Pax6 results in microphthalmia, retinal dysplasia and defective retinal ganglion cell axon guidance.
Overexpression of Pax6 results in microphthalmia, retinal dysplasia and defective retinal ganglion cell axon guidance.
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DOI：
10.1186/1471-213x-8-59
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发表时间：
2008-05-28
影响因子：
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通讯作者：
Price DJ
中科院分区：
文献类型：
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作者：
Manuel M;Pratt T;Liu M;Jeffery G;Price DJ
在发育中的眼睛中，转录因子Pax6在许多细胞类型中表达。在纯合子功能丧失的小鼠突变体(Pax6Sey/Sey)中，眼睛不形成，在Pax6Sey/+突变体中，眼睛异常小。除了内源性Pax6外，表达多个人类PAX6拷贝的PAX77小鼠的眼睛也异常小；两个物种的蛋白质序列相同。导致PAX77小鼠小眼球的发育事件尚未得到很好的描述，因此尚不清楚Pax6/PAX6的过度和低表达是否通过类似的机制引起小眼球。在这里，我们研究了过度表达对眼睛及其轴突连接的影响。眼睛在PAX77+/+胚胎中形成，但随后退化。在E12.5，我们没有发现眼形态、视网膜细胞周期参数和视网膜细胞死亡的发生率的异常。从E14.5开始，我们观察到视盘畸形。从胎龄16.5到出生后，视网膜发育不良和小眼球逐渐加重。基因表达模式分析表明，PAX77+/+视网膜产生正常范围的细胞类型，包括视网膜神经节细胞(RGC)。在E14.5和E16.5，与视网膜发育相关的一系列分子的定量RT-PCR显示只有一个显著的变化：在E16.5，编码分泌的形态原Shh的mRNA水平略有下降。在E16.5，用碳菁染料对PAX77+/+胚胎进行的轨迹追踪显示，RGC轴突在视网膜内的导航错误，到达丘脑的RGC轴突数量减少，到达丘脑的RGC轴突中同侧投射的比例增加。对具有不同Pax6/PAX6基因剂量(Pax6Sey/+、Pax6+/+、PAX77+和PAX77+/+)的胚胎的观察表明：(1)视网膜投射的RGC轴突总数和(2)在视交叉位于同侧和对侧视束的比例随基因剂量的不同而不同。增加剂量会增加同侧投射的比例，而不考虑总投射的大小。Pax6的过度表达不会明显损害眼睛的初始形成及其主要细胞类型，但会从E14.5开始阻碍视网膜的正常发育，最终导致出生后生活中严重的视网膜退化。这一序列不同于Pax6+/-杂合子中潜在的小眼球，后者主要是由于晶状体形成初期的缺陷所致。在严重的视网膜发育不良发生之前，Pax6的过度表达会导致视网膜轴突的缺陷，阻碍它们的正常生长和通过视交叉的导航。
The transcription factor Pax6 is expressed by many cell types in the developing eye. Eyes do not form in homozygous loss-of-function mouse mutants (Pax6Sey/Sey) and are abnormally small in Pax6Sey/+ mutants. Eyes are also abnormally small in PAX77 mice expressing multiple copies of human PAX6 in addition to endogenous Pax6; protein sequences are identical in the two species. The developmental events that lead to microphthalmia in PAX77 mice are not well-characterised, so it is not clear whether over- and under-expression of Pax6/PAX6 cause microphthalmia through similar mechanisms. Here, we examined the consequences of over-expression for the eye and its axonal connections. Eyes form in PAX77+/+ embryos but subsequently degenerate. At E12.5, we found no abnormalities in ocular morphology, retinal cell cycle parameters and the incidence of retinal cell death. From E14.5 on, we observed malformations of the optic disc. From E16.5 into postnatal life there is progressively more severe retinal dysplasia and microphthalmia. Analyses of patterns of gene expression indicated that PAX77+/+ retinae produce a normal range of cell types, including retinal ganglion cells (RGCs). At E14.5 and E16.5, quantitative RT-PCR with probes for a range of molecules associated with retinal development showed only one significant change: a slight reduction in levels of mRNA encoding the secreted morphogen Shh at E16.5. At E16.5, tract-tracing with carbocyanine dyes in PAX77+/+ embryos revealed errors in intraretinal navigation by RGC axons, a decrease in the number of RGC axons reaching the thalamus and an increase in the proportion of ipsilateral projections among those RGC axons that do reach the thalamus. A survey of embryos with different Pax6/PAX6 gene dosage (Pax6Sey/+, Pax6+/+, PAX77+ and PAX77+/+) showed that (1) the total number of RGC axons projected by the retina and (2) the proportions that are sorted into the ipsilateral and contralateral optic tracts at the optic chiasm vary differently with gene dosage. Increasing dosage increases the proportion projecting ipsilaterally regardless of the size of the total projection. Pax6 overexpression does not obviously impair the initial formation of the eye and its major cell-types but prevents normal development of the retina from about E14.5, leading eventually to severe retinal degeneration in postnatal life. This sequence is different to that underlying microphthalmia in Pax6+/- heterozygotes, which is due primarily to defects in the initial stages of lens formation. Before the onset of severe retinal dysplasia, Pax6 overexpression causes defects of retinal axons, preventing their normal growth and navigation through the optic chiasm.