Genetic and biophysical analysis of morphogen gradient formation

主要研究者：Schlissel，Gavin 项目摘要 动物的发育和生理学需要嵌入组织中的细胞类型之间的定期交流。 细胞通信通常依赖于信号蛋白，信号蛋白可以在细胞之间的空间传递， 在从纳米到米的广泛空间尺度上传递信息。信号的正确控制 范围在动物发育过程中是严格必要的，信号范围的失调可导致 一系列胚胎致死性疾病或发育障碍。 信号蛋白形成的图案梯度反映了信号蛋白的旅行能力 通过细胞外基质，这是一种蛋白质，糖和脂质的混合物，在自然状态下组织细胞 组织中虽然信号蛋白被认为是从它们的来源扩散到它们的靶点，但许多蛋白质违反了它们的分子结构。 自由扩散的假设，而是在它们的信令范围中示出了上下文相关的差异。为 例如，Sonic Hedgehog家族发育形态发生素在睾丸中形成超过约10µm的信号梯度， 在发育中的神经管或成年毛囊中约50µm，在发育中的长骨中约300µm。值得注意的是， Sonic Hedgehog形成较长信号梯度的组织倾向于表达Scube家族细胞外 基质蛋白和Scube家族蛋白可以显著延长细胞内的Sonic Hedgehog信号梯度， 文化 我怀疑，信号传导范围的组织特异性差异可能反映了Sonic的直接调节。 Hedgehog的扩散率，以及Hedgehog的受管制扩散可能反映了一种广泛使用的策略， 控制动物体内信号梯度的大小。为了了解形态发生素和细胞外基质 相互作用产生适当大小的信号梯度，我将测量蛋白质扩散的变化， 不同的信号蛋白之间，不同的细胞外环境之间。我将运用这一机制 了解发现信号蛋白和细胞外基质的生化特征如何导致 信号梯度之间的尺寸变化以及解剖特征之间的形态变化， 他们的模式。为此，我提出以下具体目标： 1)了解Scube家族蛋白如何通过跟踪声波的单个粒子来修改刺猬扩散 刺猬扩散通过细胞外基质。 2)发现信号蛋白的哪些生化特征会影响其在细胞中的扩散速率。 细胞外基质通过发展合成形态发生素，其中扩散可以从 下游信号转导。 3)确定有助于组织或生物体特异性的蛋白质扩散的细胞外基质修饰剂 通过遗传模拟组织特异性细胞外基质的信号梯度大小差异 变化 K99/R 00奖学金申请 二〇二二年十月

Principal Investigator: Schlissel, Gavin Project summary Animal development and physiology require regular communication among cell types embedded in tissues. Cellular communication commonly relies on signaling proteins, which can transit the space between cells and relay information across a wide range of spatial scales from nanometers to meters. Proper control of signaling range is strictly necessary during animal development, and dysregulation of signaling range can result in a spectrum of embryonic lethal conditions or developmental disorders. The patterning gradient formed by a signaling protein reflects the signaling protein’s ability to travel through the extracellular matrix, which is an amalgam of protein, sugar and lipids that organize cells in natural tissues. Although signaling proteins are thought to diffuse from their source to their target, many proteins violate the assumptions of free diffusion and instead show context-dependent differences in their signaling range. For example, Sonic Hedgehog family developmental morphogens form signaling gradients over ~10µm in the testes, ~50µm in the developing neural tube or in adult hair follicles, and ~300µm in developing long bones. Notably, tissues in which Sonic Hedgehog forms longer signaling gradients tend to express Scube family extracellular matrix proteins, and Scube family proteins can dramatically extend Sonic Hedgehog signaling gradients in cell culture. I suspect that tissue-specific differences in signaling range might reflect direct regulation of Sonic Hedgehog’s diffusion rate, and that regulated diffusion of Hedgehog might reflect a broadly used strategy to control the size of signaling gradients in animals. To understand how morphogens and the extracellular matrix interact to generate appropriately sized signaling gradients, I will measure variation in protein diffusion both between diverse signaling proteins, and between distinct extracellular environments. I will apply this mechanistic understanding to discover how biochemical features of signaling proteins and the extracellular matrix result in size variation among signaling gradients as well as morphological variation among the anatomical features that they pattern. To that end, I propose the following specific aims: 1) Understand how Scube family proteins modify hedgehog diffusion by tracking single particles of sonic hedgehog diffusing through the extracellular matrix. 2) Discover which biochemical features of a signaling protein affect its diffusion rate through the extracellular matrix by developing synthetic morphogens, in which diffusion can be uncoupled from downstream signal transduction. 3) Identify extracellular matrix modifiers of protein diffusion that contribute to tissue- or organism-specific signaling gradient size discrepancies by genetically simulating tissue-specific extracellular matrix variation K99/R00 Fellowship Application October 2022