We can all see a variety of ordered cellular patterns consisting of various cell types, throughout nature. It is surprising that these ordered cellular patterns are created reproducibly during development in all individuals. Elucidating their underlying molecular mechanisms has been an interesting research subject for developmental biologists. The essential building blocks in these processes are cell proliferation, cell shape change, cell movement, and apoptosis. These cellular behaviors must be coordinated through cell-cell communication.
Apoptosis is used extensively to refine developing structures, such as in formation of vertebrate digits and sculpting of the insect wing . Apoptosis also contributes to tissue patterning by removing abnormal cells [3-5] and eliminating excess populations of cells .
Difference in adhesiveness between cell types is another important factor in tissue patterning. Differential adhesion mediated by heterophilic adhesion molecules forces cells to rearrange during development. For example, in the oviduct epithelium of the Japanese quail, two distinct types of columnar cells; goblet-type gland cells and ciliated cells are arranged in a checkerboard pattern (Figure 1E) . Preferential adhesion between different cell types rather than between cells of the same type could account for this pattern .
Experiments have shown that spatial and temporal regulation of apoptosis or cell adhesion is indispensable for correct patterning. Inappropriate cells must be removed at the proper time by apoptosis and each living cell must attach properly to its counterparts. How are these processes regulated? In this chapter, we will describe the
Drosophila eye patterning
A striking example of ordered cellular packing is the
The remaining two-thirds of the interommatidial cells, which are in contact with ommatidia, do not undergo apoptosis. Spatial regulation of apoptosis is mediated by epidermal growth factor receptor (EGFR) signaling . Spitz, a ligand of EGFR, is produced in the primary pigment cells and secreted around surrounding cells. This activation of EGFR signaling downregulates the activity of Hid, a proapoptotic protein, which prevents these adjacent cells from undergoing apoptosis [14, 15]. In this fashion, only non-adjacent IPCs lack the EGFR signal and thus undergo apoptosis.
Drosophila wing margin hairs
During this cell rearrangement, a subset of wing margin cells is removed through apoptosis. The dying cells are the cells that have not attached to the hair cells. Blocking apoptosis by expressing the baculovirus caspase inhibitor p35  in wing margin cells using the GAL4/UAS system  inhibits cell rearrangement, indicating that apoptosis is required for this process.
What triggers apoptosis in a precise temporal manner? Ecdysone, an insect steroid hormone, is indispensable for progression in most of the developmental stages of
Then, how is apoptosis in the developing tissue regulated in a precise spatial manner? Vein, a diffusible ligand of EGFR, is expressed specifically in the hair cells. In addition, EGFR activation is observed in cells surrounding hair cells, as revealed by the expression of
4. Coordination of preferential adhesion and secreted survival signaling molecules
In both tissues described above, locally diffusible ligands are used to make neighboring cells survive. Thus, regulation of cell-cell contact is an important factor for controlling the spatial pattern of apoptosis. In both cases,
In the compound eye, immunohistochemical staining reveals that all four molecules accumulate at the interface between ommatidia and IPCs [37, 38]. Hibris and Sns are expressed in the ommatidia, and their binding partners Kirre and Rst are expressed in the IPCs. Computer simulation have shown that preferential adhesion between ommatidia and IPCs contribute to the cell rearrangement .
Similarly, antibody staining of the pupal wing indicate that all these adhesion molecules accumulate the interface between hair cell and their neighboring cells . Enhancer-trap reporters for these genes also show that NEPH1 groups and Nephrin homologs are expressed in an almost complementary pattern (Figure 4). Cell-type-specific knockdown of these molecules results in disruption of the wing margin pattern. For instance, when we knockdown Rst in interhair cells, we observed some hair cells away from interhair cells and tooth cells (unpublished data). Knockdown of each gene results in disruption of the posterior wing margin hairs, indicating that all four of these molecules are required for proper hair patterning.
Therefore, in both the
This seems to be a good strategy for creating ordered repetitive cellular patterns through refinement (Figure 5). It is tempting to speculate that similar mechanisms work in other tissues of other organisms. Lastly, we will discuss the cellular patterning in the cochlea, a mammalian inner ear organ.
5. Patterning of outer hair cells and supporting cells in the organ of Corti
The sensory epithelium of the mammalian cochlea, the organ of Corti, has three rows of outer hair cells and supporting cells, which are aligned in a checkerboard pattern (Figure 6). Outer hair cells (OHCs) are essential for the amplification of sound  and the loss of these cells can lead to hearing loss. Specification of hair cells and supporting cells are mediated by Notch-mediated lateral inhibition [40-42], as is the case with
We have described the apoptosis-dependent cell rearrangement for refining cellular arrangements of the
We would like to thank Dr. Hisao Honda for kindly providing the picture of the oviduct epithelium of the Japanese quail. We are grateful to Daniel Levings and Pui Choi for critical reading of the manuscript.
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