While the transcriptional code governing retinal ganglion cell (RGC) type specification begins to be understood, its interplay with neurotrophic signaling is largely unexplored. In mice, the transcription factor Brn3a/Pou4f1 is expressed in most RGCs, and is required for the specification of RGCs with small dendritic arbors. The Glial Derived Neurotrophic Factor (GDNF) receptor Ret is expressed in a subset of RGCs, including some expressing Brn3a, but its role in RGC development is not defined. Here we use combinatorial genetic experiments using conditional knock-in reporter alleles at the Brn3a and Ret loci, in combination with retina- or Ret specific Cre drivers, to generate complete or mosaic genetic ablations of either Brn3a or Ret in RGCs. We then use sparse labelling to investigate Brn3a and Ret gene dosage effects on RGC dendritic arbor morphology. In addition, we use immunostaining and/or gene expression profiling by RNASeq to identify transcriptional targets relevant for the potential Brn3a-Ret interaction in RGC development. We find that mosaic gene dosage manipulation of the transcription factor Brn3a/Pou4f1 in neurotrophic receptor Ret heterozygote RGCs results in altered cell fate decisions and/or morphological dendritic defects. Specific RGC types are lost if Brn3a is ablated during embryogenesis and only mildly affected by postnatal Brn3a ablation. Sparse but not complete Brn3a heterozygosity combined with complete Ret heterozygosity has striking effects on RGC type distribution. Brn3a only mildly modulates Ret transcription, while Ret knockouts exhibit slightly skewed Brn3a and Brn3b expression during development that is corrected by adult age. Brn3a loss of function modestly but significantly affects distribution of Ret co-receptors GFRα1-3, and neurotrophin receptors TrkA and TrkC in RGCs. Based on these observations, we propose that Brn3a and Ret converge onto developmental pathways that control RGC type specification, potentially through a competitive mechanism requiring signaling from the surrounding tissue.

中文翻译：

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视网膜神经节细胞类型规范中转录因子 Pou4f1/Brn3a 与神经营养因子受体 Ret 之间的遗传相互作用

虽然控制视网膜神经节细胞 (RGC) 类型规范的转录代码开始被理解，但它与神经营养信号的相互作用在很大程度上尚未探索。在小鼠中，转录因子 Brn3a/Pou4f1 在大多数 RGC 中表达，并且是规范具有小树突乔木的 RGC 所必需的。胶质源性神经营养因子 (GDNF) 受体 Ret 在 RGC 的一个子集中表达，包括一些表达 Brn3a，但其在 RGC 发育中的作用尚未确定。在这里，我们使用组合遗传实验，使用 Brn3a 和 Ret 基因座的条件敲入报告基因等位基因，结合视网膜或 Ret 特异性 Cre 驱动程序，在 RGC 中生成 Brn3a 或 Ret 的完整或镶嵌遗传消融。然后，我们使用稀疏标记来研究 Brn3a 和 Ret 基因剂量对 RGC 树突乔木形态的影响。此外，我们通过 RNASeq 使用免疫染色和/或基因表达谱来识别与 RGC 发育中潜在的 Brn3a-Ret 相互作用相关的转录靶标。我们发现神经营养受体 Ret 杂合子 RGC 中转录因子 Brn3a/Pou4f1 的镶嵌基因剂量操作导致细胞命运决定和/或形态树突缺陷的改变。如果 Brn3a 在胚胎发生过程中被消融，特定的 RGC 类型会丢失，并且仅受出生后 Brn3a 消融的轻微影响。稀疏但不完全的Brn3a杂合性与完全Ret杂合性相结合对RGC类型分布具有显着影响。Brn3a 仅轻微调节 Ret 转录，而 Ret 基因敲除在发育过程中表现出略微偏斜的 Brn3a 和 Brn3b 表达，并被成年年龄纠正。Brn3a 功能丧失适度但显着影响 Ret 共受体 GFRα1-3 和神经营养因子受体 TrkA 和 TrkC 在 RGC 中的分布。基于这些观察，我们建议 Brn3a 和 Ret 收敛到控制 RGC 类型规范的发育途径，可能通过需要来自周围组织的信号传导的竞争机制。