Recruitment of Rod Photoreceptors from Short Wavelength Sensitive Cones during the Evolution of Nocturnal Vision in Mammals

Vertebrate ancestors had only cone-like photoreceptors. The duplex retina evolved in jawless vertebrates with the advent of highly photosensitive rod-like photoreceptors. Despite cones being the arbiters of high-resolution color vision, rods emerged as the dominant photoreceptor in mammals during a nocturnal phase early in their evolution. We investigated the evolutionary and developmental origins of rods in two divergent vertebrate retinae. In mice, we discovered genetic and epigenetic vestiges of short wavelength cones in developing rods and cell lineage tracing validated the genesis of rods from S-cones. Curiously, rods did not derive from S-cones in zebrafish. Our study illuminates several questions regarding the evolution of duplex retina and supports the hypothesis that, in mammals, the S-cone lineage was recruited via the Maf-family transcription factor NRL to augment rod photoreceptors. We propose that this developmental mechanism allowed the adaptive exploitation of scotopic niches during the nocturnal bottleneck early in mammalian evolution. GFP positive cells from Nrlp-GFP or Nrlp-GFP;Nrl-KO mouse retinas at post-natal ages P2, P4, P6, P10, P14, and P28 were isolated by flow sorting by FACSAria II (Becton Dickinson). Total RNA was extracted by Trizol LS (Life Technologies) and analyzed by 2100 Bioanalyzer (Agilent Technologies Genomics). High quality of total RNA (RIN: >7.0) was subjected to sequencing library construction using 20 ng of total RNA as input. Libraries were constructed using a stranded modification of the Illumina TruSeq mRNA (Brooks, et al. Meth Mol Biol 2012). Each library was single-end sequenced in an independent lane of a GAIIx at a length of 76 bases. Fastq files were generated from reads passing chastity filter.脊椎动物的祖先仅拥有视锥类感光受体（cone-like photoreceptors）。随着高感光性视杆类感光受体（rod-like photoreceptors）的出现，双感光视网膜（duplex retina）在无颌脊椎动物（jawless vertebrates）中演化形成。尽管视锥细胞是高分辨率色觉的核心介导者，但在哺乳动物演化早期的夜行性阶段，视杆细胞成为了其主导的感光受体。本研究针对两种演化分歧的脊椎动物视网膜，探究了视杆细胞的演化与发育起源。在小鼠中，我们于发育中的视杆细胞内发现了短波视锥（short wavelength cones, S-cones）的遗传与表观遗传遗迹，且细胞谱系追踪实验证实视杆细胞由S-视锥细胞分化而来。令人意外的是，斑马鱼体内的视杆细胞并非由S-视锥细胞分化而来。本研究厘清了数个关于双感光视网膜演化的关键问题，并支持下述假说：在哺乳动物中，S-视锥细胞谱系通过Maf家族转录因子NRL（Maf-family transcription factor NRL）被招募，以扩充视杆感光受体的数量。我们提出，该发育机制使得哺乳动物在演化早期的夜行性瓶颈阶段，得以适应性地占据暗视觉生态位（scotopic niches）。从产后第2、4、6、10、14及28天（P2、P4、P6、P10、P14及P28）的Nrlp-GFP或Nrlp-GFP;Nrl-KO小鼠视网膜中分离得到的GFP阳性细胞，通过FACSAria II流式细胞仪（Becton Dickinson）完成流式分选。采用Trizol LS（Life Technologies）提取总RNA，并通过2100生物分析仪（Agilent Technologies Genomics）进行质量检测。选取RNA完整性指数（RIN）＞7.0的高质量总RNA，以20 ng总RNA为起始量构建测序文库。文库构建采用Illumina TruSeq mRNA的链特异性修饰方案（Brooks等人，《Methods in Molecular Biology》，2012）。每个文库均在GAIIx测序仪的独立泳道中进行76个碱基的单端测序，通过纯度过滤的测序读段被用于生成Fastq格式文件。