Human Genome Sequencing Center, Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX 77030, USA.
Verna and Marrs McLean Department of Biochemistry and Molecular Biology, Baylor College of Medicine, Houston, TX, 77030 USA.
Hum Mol Genet. 2021 Oct 13;30(21):1907-1918. doi: 10.1093/hmg/ddab157.
Much of the complexity of the eukaryotic cell transcriptome is due to the alternative splicing of mRNA. However, knowledge on how transcriptome complexity is translated into functional complexity remains limited. For example, although different isoforms of a gene may show distinct temporal and spatial expression patterns, it is largely unknown whether these isoforms encode proteins with distinct functions matching their expression pattern. In this report, we investigated the function and relationship of the two isoforms of Reep6, namely Reep6.1 and Reep6.2, in rod photoreceptor cells. These two isoforms result from the alternative splicing of exon 5 and show mutually exclusive expression patterns. Reep6.2 is the canonical isoform that is expressed in non-retinal tissues, whereas Reep6.1 is the only expressed isoform in the adult retina. The Reep6.1 isoform-specific knockout mouse, Reep6E5/E5, is generated by deleting exon 5 and a homozygous deletion phenotypically displayed a rod degeneration phenotype comparable to a Reep6 full knockout mouse, indicating that the Reep6.1 isoform is essential for the rod photoreceptor cell survival. Consistent with the results obtained from a loss-of-function experiment, overexpression of Reep6.2 failed to rescue the rod degeneration phenotype of Reep6 knockout mice whereas overexpression of Reep6.1 does lead to rescue. These results demonstrate that, consistent with the expression pattern of the isoform, Reep6.1 has rod-specific functions that cannot be substituted by its canonical isoform. Our findings suggested that a strict regulation of splicing is required for the maintenance of photoreceptor cells.
真核细胞转录组的复杂性很大程度上是由于 mRNA 的选择性剪接。然而，关于转录组复杂性如何转化为功能复杂性的知识仍然有限。例如，尽管一个基因的不同异构体可能表现出不同的时空表达模式，但很大程度上不清楚这些异构体是否编码与其表达模式相匹配的具有不同功能的蛋白质。在本报告中，我们研究了 rod 光感受器细胞中 Reep6 的两种异构体（即 Reep6.1 和 Reep6.2）的功能和关系。这两种异构体是由外显子 5 的选择性剪接产生的，表现出相互排斥的表达模式。Reep6.2 是在非视网膜组织中表达的典型异构体，而 Reep6.1 是成年视网膜中唯一表达的异构体。Reep6.1 异构体特异性敲除小鼠 Reep6E5/E5 通过删除外显子 5 产生，纯合缺失表型显示出与 Reep6 完全敲除小鼠相当的 rod 变性表型，表明 Reep6.1 异构体对于 rod 光感受器细胞的存活是必不可少的。与功能丧失实验的结果一致，过表达 Reep6.2 未能挽救 Reep6 敲除小鼠的 rod 变性表型，而过表达 Reep6.1 则导致挽救。这些结果表明，与异构体的表达模式一致，Reep6.1 具有 rod 特异性功能，不能被其典型异构体替代。我们的研究结果表明，严格的剪接调控对于维持光感受器细胞是必需的。
