Emergence of functional polarity in a tubular epithelium: a mechanistic study

我们体内的许多组织都是围绕着复杂的细胞管阵列构建起来的，这些细胞管允许维持生命的分子（如氧气、葡萄糖和水）进出和运输。但这些管道不仅仅是简单地来回输送材料，它们还在修饰通过它们的材料方面发挥着至关重要的作用。上皮管的功能差异是我们身体各器官的普遍特征；这些差异的起源是由区域细胞分化支撑的。组成肾脏的微观单位——肾元——就是一个很好的例子。一端专门用来过滤血液，产生初级尿液。接收尿液的管状系统有不同的部分，细胞的运输特性和对水、盐和其他化合物的渗透性不同。正是沿着肾元近端到远端（P-D）轴的这些活动序列——我们将其定义为“功能极性”——允许肾脏产生和浓缩尿液。对于大多数器官来说，协调功能极性出现的机制还没有得到很好的理解。本研究的重点是如何沿着果蝇肾脏系统中简单细胞小管的P-D轴建立功能极性。昆虫的肾脏系统或马氏小管（MpT）是排泄、清除废物和毒素的主要器官，它控制着身体的盐和水的预算。mpt是简单的细胞管，包含少量不同类型的细胞，总的细胞数量很少（果蝇约120个）。然而，这种简单掩盖了几项截然不同且至关重要的活动。这些功能包括（但不限于）尿液的分泌、尿液的再吸收（以回收有价值的物质、调节液体和盐）和控制身体的钙水平。这些不同的活动是在小管空间受限的部分进行的。它们的执行顺序强调了器官功能（例如，在发生修饰之前产生尿液显然很重要）。MpT的简单性、实验的可追溯性和测量功能的能力，使它成为解决这里概述的基本问题的理想学科。我们将研究可能指定P-D轴的三种替代机制：(a)重叠的化学信号起作用，以建立叠加在细胞平面上的楔形和同心圆的差异基因表达的“飞镖样图案”。当小管从薄片上伸缩出来形成三维结构时，靶心、中环和外环细胞就会根据这些基因活动的同心圆形成不同的身份。(b)不对称化学信号(s)从发育中的小管的基部(P)和尖端(D)发出。细胞根据其沿P-D轴的位置经历不同浓度的信号，并利用这一信号来指导它们的分化。(c)细胞在小管中的增殖受到严格控制，并在不同的阶段依次发生。这种出生顺序控制分化：早、中、晚出生的细胞发育成不同的细胞类型。我们还将研究一组已确定的“控制基因”，即转录因子（tf），它们具有片段特异性活性。我们认为这些tf是由上述发育机制激活的，而这些机制反过来又激活编码通道和运输蛋白的基因，最终执行分泌、重吸收和钙处理活动。使用各种遗传方法来操纵和标记不同的细胞组，我们将确定基因之间的网络和连接，这些网络和连接绘制了沿着mpt的P-D轴出现的功能极性。我们期望我们的工作将导致更好地了解我们自己身体管状器官的发育和功能。

Many of the tissues in our bodies are built up around complex arrays of cellular tubes, which permit the entry, exit, and transport of life-sustaining molecules such as oxygen, glucose, and water. But these tubes do more than simple convey materials to and fro - they also carry out vital roles in modifying the material passing through them. Functional differences along epithelial tubes are a prevalent feature in the organs in our bodies; the origin of these differences is underpinned by regional cell differentiation. The microscopic unit that makes up our kidney - the nephron - is a good example. One end is specialised to filter the blood, producing the primary urine. The tubular system that receives this urine has distinct segments with cells differing in their transport properties and permeabilities to water, salts and other compounds. It is these sequences of activity along the proximal-to-distal (P-D) axis of the nephron - a property we define as 'functional polarity' - that allows the kidney to produce and concentrate urine. The mechanisms that orchestrate the emergence of functional polarity are not well understood for most organs. This proposal focuses on how functional polarity is established along the P-D axis of the simple cellular tubule in the fruit fly (Drosophila) renal system.The insect renal system or Malpighian tubule (MpT) is the major organ for excretion, removal of wastes and toxins, and it controls the body's salt and water budget. MpTs are simple cellular tubes that contain a handful of different cell types, and a small number of cells overall (~120 in Drosophila). Yet this simplicity belies several distinct and vitally important activities. These include (but are not limited to) secretion of urine, reabsorption from the urine (to retrieve valuable materials, and to regulate fluid and salt) and control of the body's calcium levels. These different activities are carried out in spatially restricted segments of the tubule. Their order of execution underscores organ function (e.g. it is clearly important to produce urine before modification can take place). The simplicity of the MpT, its experimental tractability, and an ability to measure function, makes it an ideal subject to tackle the fundamental question outlined here.We will investigate three alternative mechanisms that might specify the P-D axis: (a) Overlapping chemical signals act to establish a 'dartboard-like pattern' of wedges and concentric rings of differential gene expression superimposed onto a flat sheet of cells. When the tubule telescopes-out from the sheet as a 3-D structure, the bullseye, mid ring- and outer ring-cells develop alternative identities based on these concentric rings of gene activity. (b) Asymmetric chemical signal(s) are issued from the base (P) and tip (D) of the developing tubule. Cells experience differing concentrations of the signal(s) depending on their position along the P-D axis, and use this to guide their differentiation. (c) Cell proliferation in the tubule is tightly controlled and occurs sequentially in distinct phases. This birth order acts to controls differentiation: with early-, mid- and late-born cells developing into alternative cell-types. We will also investigate an identified set of 'control genes', known as transcription factors (TFs), which have segment specific activities. We suggest these TFs are activated by the developmental mechanism(s) outlined above, and these in turn, activate genes encoding the channels and transport proteins that ultimately execute the activities of secretion, reabsorption and calcium handling. Using a variety of genetic methods to manipulate and mark different sets of cells, we will identify the networks and connections between genes that map out the emergence of functional polarity along the P-D axis of MpTs. We anticipate our work will lead to a better understanding of the development and function of the tubular organs of our own bodies.