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

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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.