Rangachar Chetan C, Moran Denice D, Emerson Mark M
Department of Biology, The City College of New York, City University of New York, New York, NY 10031, USA.
Biology PhD Program, Graduate Center, City University of New York, New York, NY, 10016.
bioRxiv. 2025 Jul 31:2025.07.30.667746. doi: 10.1101/2025.07.30.667746.
The retinal development of elasmobranchs--the superclass comprising sharks, skates and rays--is a poorly understood phenomenon. The clade is diverse in retinal phenotypes, with many sharks and rays possessing rods and multiple cone types. In contrast, the little skate () has only a single type of rod photoreceptor, which is reported to have taken on some physiological and anatomical properties of cones. How does the little skate develop this photoreceptor system? To investigate this question, we identified an early stage of embryonic photoreceptor formation based on Otx2 expression. To assess whether cone photoreceptor gene regulatory networks regulated by the Onecut1 transcription factor were intact we developed an electroporation approach. Activation of a Onecut1-dependent reporter was not detected supporting a change in early Onecut1-transcriptional networks associated with cone photoreceptor formation. To assess developmental changes in gene expression, bulk RNA-Seq samples of embryos, juvenile hatchlings and adult retinas were generated. As expected, gene expression changes associated with cone photoreceptors of other species were not identified in the little skate and several cone-enriched genes were found to be pseudogenes. A splice isoform of Onecut1 with an additional 48 amino acid sequence located between the CUT and Homeodomain DNA-binding domains - referred to as "spacer isoform" or LSOC1X2 was identified and exhibited high expression in the embryo compared to juvenile and adult stages. To test if this LSOC1X2 isoform retained its regulatory potential compared to the isoform without the additional sequence, we tested it in a mouse retina reporter assay. Both skate Onecut1 isoforms were able to activate the Onecut1-dependent transcriptional reporter. Thus, the spacer isoform represents a developmentally regulated, novel retinal Onecut1 isoform with regulatory potential. This identifies it as a target for further analysis in the retinal development of the little skate and its elasmobranch relatives.
软骨鱼类（包括鲨鱼、鳐鱼和魟鱼的一个纲）的视网膜发育是一个尚未得到充分理解的现象。该类群在视网膜表型上具有多样性，许多鲨鱼和魟鱼拥有视杆细胞和多种视锥细胞类型。相比之下，小鳐（）只有一种视杆光感受器，据报道这种光感受器具有一些视锥细胞的生理和解剖特性。小鳐是如何发育出这种光感受器系统的呢？为了研究这个问题，我们基于Otx2的表达确定了胚胎光感受器形成的早期阶段。为了评估由Onecut1转录因子调控的视锥光感受器基因调控网络是否完整，我们开发了一种电穿孔方法。未检测到Onecut1依赖性报告基因的激活，这支持了与视锥光感受器形成相关的早期Onecut1转录网络发生了变化。为了评估基因表达的发育变化，我们生成了胚胎、幼年孵化幼体和成年视网膜的大量RNA测序样本。正如预期的那样，在小鳐中未发现与其他物种视锥光感受器相关的基因表达变化，并且发现几个富含视锥细胞的基因是假基因。我们鉴定出一种Onecut1的剪接异构体，其在CUT和同源结构域DNA结合结构域之间有一个额外的48个氨基酸序列，被称为“间隔异构体”或LSOC1X2，与幼年和成年阶段相比，它在胚胎中表现出高表达。为了测试这种LSOC1X2异构体与没有额外序列的异构体相比是否保留了其调控潜力，我们在小鼠视网膜报告基因试验中对其进行了测试。两种鳐鱼的Onecut1异构体都能够激活Onecut1依赖性转录报告基因。因此，间隔异构体代表了一种具有调控潜力的、受发育调控的新型视网膜Onecut1异构体。这使其成为在小鳐及其软骨鱼类亲属的视网膜发育中进行进一步分析的目标。
