Molecular control of neuronal shape and connectivity in the developing retina

 描述(由申请人提供)：神经元表现出不同的形态，影响信息如何通过复杂的连接网络传播和调制。电路功能的一个关键决定因素是树枝的数量和排列。例如，局部中间神经元从细胞体对称地延伸出多个树突，而小脑Purkinje神经元则形成局限于分子层的单个巨大的树突。像轴突一样，树突是从分化神经元的细胞体延伸而来的全能神经突起。一个轴突变成一个轴突。剩下的神经突起要么被收回，要么被保留下来，发育成树突。在活体中，这些事件与周围组织协调，使轴突和树突在固定的位置发育，在那里它们可以完美地与适当的突触伙伴相互作用。我们的长期目标是 了解外部信号如何改变幼稚神经突起的内在属性，从而确保神经元获得与电路其余部分正确定向的极化形态。为了解决这个问题，我们将研究无长突细胞中树突指定的机制，它调节信息从视网膜外部到内部的流动。无长突细胞发育出单一的初级树突，该树突指向神经纤维层的特定区域，称为内丛状层(IPL)。发育中的无长突细胞在迁移时是双极的，但在接触新生的IPL时变得单极：接触IPL的轴突保留为树突，但细胞相反极的轴突被收回。我们开发了一种延时成像系统，使我们能够在视网膜中的无长突细胞从双极形态转变为单极形态时记录它们，无论是在整体细胞形状水平上还是在细胞骨架水平上。我们发现，这种极性变化的一个关键读数是高尔基体的位置，它移动到新生的初级树枝晶中。在非典型钙粘附素Fat3突变的小鼠中，这种转变不会可靠地发生，导致出现具有两个树突的无长突细胞和一个异位放置的高尔基体。作为一种跨膜受体，Fat3具有保守的胞内区，含有蛋白结合基序，为理解外部线索如何导致神经元形态的内在变化提供了一个强有力的切入点。事实上，Fat3细胞内结构域不仅与已知的肌动蛋白调节因子(即Ena/Vasp家族成员)结合，还与控制微管动力学的蛋白质结合(即CLASP1/2)，这表明Fat3协调对肌动蛋白和微管的双重作用。通过将Fat3及其效应分子的分子和遗传学研究与原位无长突细胞树突状细胞发育的时间推移分析相结合，我们将对支配树突指定的外在和内在机制有新的见解。

 DESCRIPTION (provided by applicant): Neurons exhibit diverse morphologies that influence how information is propagated and modulated through complex networks of connections. One key determinant of circuit function is the number and arrangement of dendrites. For instance, local interneurons extend multiple dendrites symmetrically from the cell body, whereas cerebellar Purkinje neurons elaborate a single huge dendritic arbor that is confined to the molecular layer. Like axons, dendrites develop from totipotent neurites that extend from the cell body of the differentiating neuron. One neurite becomes an axon. The remaining neurites are either retracted or retained to develop as dendrites. In vivo, these events are coordinated with the surrounding tissue, such that axons and dendrites develop in stereotyped locations where they are perfectly positioned to interact with appropriate synaptic partners. Our long term goal is to understand how extrinsic signals alter the intrinsic properties of naïve neurites, thereby ensuring that neurons acquire polarized morphologies that are correctly oriented with the rest of the circuit. To tackle this question, we will investigate mechanisms of dendrite specification in amacrine cells, which modulate the flow of information from the outer to the inner retina. Amacrine cells develop a single primary dendrite that points into a defined region of neuropil called the inner plexiform layer (IPL). Developing amacrine cells are bipolar as they migrate but become unipolar upon contacting the nascent IPL: the neurite that contacts the IPL is retained as a dendrite, but the neurite on the opposite pole of the cell is retracted. We have developed a time‐lapse imaging system that allows us to document amacrine cells in the retina as they transition from a bipolar to unipolar morphology, both at the level of the overall cell shape and a the level of the cytoskeleton. We find that a key readout for this change in polarity is the positin of the Golgi apparatus, which moves into the nascent primary dendrite. In mice mutant for the atypical cadherin Fat3, this transition does not occur reliably, leading to the appearance of amacrine cells with two dendritic arbors and an ectopically placed Golgi apparatus. As a transmembrane receptor with a conserved intracellular domain harboring protein‐binding motifs, Fat3 offers a potent entry point for understanding how extrinsic cues lead to intrinsic changes in neuronal morphology. Indeed, the Fat3 intracellular domain binds not only to known actin regulators (i.e. Ena/VASP family members) but also to proteins that control microtubule dynamics (i.e. CLASP1/2), suggesting that Fat3 coordinates dual effects on actin and microtubules. By pairing molecular and genetic studies of Fat3 and its effectors with time‐lapse analysis of amacrine cell dendrite development in situ, we will gain new insights into the extrinsic and intrinsic mechanisms that govern dendrite specification.