Genetic tuning of ipRGC subtype identity to shape visual behavior

The distinct blend of molecular and cellular features that define neuronal subtype identity are central to shaping how individual subtypes impact animal behavior. The diversity of the mammalian nervous system is vast the retina alone contains over 100 neuronal subtypes. Yet, the genetic processes giving rise to this stunning structural and functional diversity remain poorly understood. Here, we uncover a graded expression pattern of the transcription factor Brn3b that tunes and maintains multiple, subtype-defining transcriptional and morphophysiological features of the melanopsin-expressing, intrinsically photosensitive retinal ganglion cells (ipRGCs). Disruption of this Brn3b gradient causes the transcriptional and morphophysiological identity of ipRGC subtypes to begin to converge, leading to dysfunction in multiple ipRGC-dependent behaviors. These findings show that a single transcription factor gradient can tune a diverse array of features to shape neuronal identity and circuit function to drive behavior. Retina from Brn3b conditional knockout (Brn3bcKO) mice expressing Rpl22-HA in ipRGCs or control mice were subjected to translating ribosome affinity purification (TRAP) followed by library preparation using the Smart-seq3 protocol. RNA libraries were sequenced on the Illumina NextSeq 550 platform to obtain 37 bp paired-end reads. Two to four biological replicates were performed for all conditions.定义神经元亚型身份的独特分子与细胞特征组合，是决定各亚型如何影响动物行为的核心因素。哺乳动物神经系统的多样性极为丰富——仅视网膜就包含超过100种神经元亚型。然而，催生这一惊人结构与功能多样性的遗传过程，目前仍知之甚少。本研究揭示了转录因子Brn3b的分级表达模式，该模式可调控并维持表达黑视蛋白的内在感光视网膜神经节细胞（intrinsically photosensitive retinal ganglion cells，ipRGCs）多种亚型特异性的转录与形态生理特征。若破坏这一Brn3b表达梯度，ipRGC亚型的转录与形态生理特征会趋于趋同，进而导致多种ipRGC依赖性行为出现功能异常。这些研究结果表明，单一转录因子的表达梯度可调控一系列多样的特征，从而塑造神经元身份与神经环路功能，最终驱动动物行为。将在ipRGC中表达Rpl22-HA的Brn3b条件性敲除（Brn3bcKO）小鼠及对照小鼠的视网膜样本，进行翻译核糖体亲和纯化（TRAP），随后采用Smart-seq3技术进行文库制备。使用Illumina NextSeq 550平台对RNA文库进行测序，获取37 bp双端读段。所有实验条件均设置2至4次生物学重复。