The roles of LIM-Homeodomain Transcription Factors in Retinal Development

LIM-同源域转录因子在视网膜发育中的作用

基本信息

批准号：
10116905
负责人：
Lin Gan
金额：
$ 38.38万
依托单位：
AUGUSTA UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-03-01 至 2022-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10116905
关键词：
Amacrine Cells Axon Blindness Cell Differentiation process Cell Lineage Cells Characteristics Contrast Sensitivity Coupled Data Dendrites Detection Development Dyes Electrophysiology (science)Enterobacteria phage P1 Cre recombinase Event Failure Family Ganglia Ganglion Cell Layer Genes Genetic Genetic Transcription Individual Injections Inner Nuclear Layer Inner Plexiform Layer Knock-in LHX2 gene Link Location Massive Parallel Sequencing Mediating Molecular Morphology Mus Nervous system structure Neuraxis Neurons Pathway interactions Pattern Photoreceptors Play Process Property Proteins Regulatory Pathway Reporter Retina Retinal Cone Retinal Defect Role Shotgun Sequencing Stratification Structure Subgroup Synapses Technology Vision Visual Perception Visual impairment base cell type chromatin immunoprecipitation homeodomain in vivo interest member neural circuit novel null mutation object motion patch clamp public health relevance retinal neuron retinal rods retinogenesis transcription factor transcriptome transcriptome sequencing visual information visual processing

项目摘要

DESCRIPTION (provided by applicant): Our accurate vision depends on the flow of visual information through precisely wired synaptic connections among axons and dendrites of retinal neurons with unique morphological and functional properties. In the vertebrate retina, each of the six neuronal cell types: ganglion, amacrine, bipolar, horizontal, and rod and cone photoreceptor cells are further divided into subtypes based on location, morphology and function. Of all retinal neurons, amacrine cells are the most diverse group with >30 subtypes being identified so far. They represent ~40% of neurons both in the inner nuclear layer (INL) and the ganglion cell layer (GCL), make up a majority of synapses in the inner plexiform layer (IPL), and contribute to a majority of visual processing in the retina. One of the key questions i how the many retinal neuronal subtypes are produced and wired during development. In this proposal, we focus on the amacrine cells associated with the sublaminar layer 3 (S3) of the IPL. The S3 sublamina separates the ON and OFF laminas in the IPL but its cellular makeup and function is poorly understood. Here, we have demonstrated LHX9, a LIM-homeodomain transcription factor, is expressed early in retinogenesis and its expression is tightly confined toa few amacrine cells in the INL and the GCL. In our preliminary study, we have shown that these LHX9+ cells are a subgroup of GABAergic amacrine cells and express GAD67 but not GAD65. LHX9-expressing cells are also LHX2-expressing subgroup of amacrine cells. Targeted deletion of Lhx9 in mice results in a nearly complete loss of these LHX2-expressing amacrine cells and strikingly, in the absence of the S3 sublamina, suggesting that LHX9 could be expressed in and be required for the development of a unique, S3-stratifying amacrine subtype cells. Interestingly, our preliminary data show that bNOS expression is significantly down-regulated in the Lhx9-null retina, suggesting a loss of bNOS-subtype of amacrine cells that are known to project in the S3 sublamina. Being a transcription factor with a known function in neuronal subtype development in the central nervous system, LHX9 likely plays a critic role in amacrine subtype specification and offers us a unique opportunity to ultimately elucidate the genetic pathway governing the formation of the S3 sublamina and its associated neural circuitry. In this proposal, we will fully characterize the subtype identity of these LHX9-expressing amacrine subtypes and will identify its circuitry within the retina. Second, we will analyze the retinal defects of Lhx9-null mutation, particularly the effect on amacrine subtype specification, differentiation of these Lhx9-expressing S3 stratifying amacrine cells, and the change in the functional properties and circuitry of the Lhx9-lineage cells. To elucidate the LHX9 regulatory pathway in the S3-stratifying amacrine cells, we will perform RNA-Seq of control and Lhx9-null retinas and use LHX9 ChIP-Seq to screen for downstream target genes of LHX9 and to identify the transcriptional network. Together, these studies will define the role of LHX9 in regulating the formation and neural circuitry of S3 sublamina and elucidate the transcriptional events that occur downstream of LHX9.

描述（由申请人提供）：我们准确的视力取决于通过视网膜神经元的轴突和树突之间精确有线突触连接的视觉信息流，这些神经元具有独特的形态和功能特性在脊椎动物视网膜中，有六种神经元细胞类型：神经节。根据所有视网膜神经元的位置、形态和功能，将无长突细胞、双极细胞、水平细胞、视杆细胞和视锥细胞进一步分为亚型。无长突细胞是最多样化的群体，迄今为止已识别出超过 30 种亚型，它们代表了内核层 (INL) 和神经节细胞层 (GCL) 神经元的约 40%，构成了内核层的大部分突触。丛状层（IPL），并有助于视网膜的大部分视觉处理，其中一个关键问题是许多视网膜神经亚型在发育过程中是如何产生和连接的。与 IPL 的第 3 层下层 (S3) 相关的无长突细胞 S3 下层将 IPL 中的 ON 层和 OFF 层分开，但其细胞组成和功能知之甚少。因子，在视网膜发生早期表达，其表达严格限制在 INL 和 GCL 中的一些无长突细胞中。 LHX9+ 细胞是 GABA 能无长突细胞的一个亚群，表达 GAD67，但不表达 GAD65，也是表达 LHX2 的无长突细胞亚群，导致小鼠中这些表达 LHX2 的无长突细胞几乎完全丧失。引人注目的是，在缺乏 S3 亚层的情况下，这表明 LHX9 可以在我们的初步数据表明，bNOS 表达在 Lhx9 缺失的视网膜中显着下调，表明已知在 S3 亚层中投射的无长突细胞的 bNOS 亚型丢失。作为一种在中枢神经系统神经元亚型发育中具有已知功能的转录因子，LHX9 可能在无长突亚型规范中发挥着关键作用，并为我们最终提供了独特的机会。阐明控制 S3 亚层形成及其相关神经回路的遗传途径在本提案中，我们将充分表征这些表达 LHX9 的无长突亚型的亚型身份，并确定其在视网膜内的回路。 Lhx9 缺失突变的视网膜缺陷，特别是对无长突亚型规范的影响、这些表达 Lhx9 的 S3 分层无长突细胞的分化，以及 Lhx9 谱系细胞的功能特性和电路的变化，以阐明 S3 分层中的 LHX9 调节途径。无长突细胞，我们将对对照和 Lhx9 缺失的视网膜进行 RNA 测序，并使用 LHX9 ChIP-Seq 筛选 LHX9 的下游靶基因并鉴定转录网络，这些研究将共同确定 LHX9 在调节无长突细胞中的作用。 S3 下层的形成和神经回路，并阐明 LHX9 下游发生的转录事件。