New branch formation by clefting. (A) Schematics of clefting (or terminal bifurcation) in mouse lung. The clefting tip of developing lung contains a single layer of cells that flattens before clefting. Within the tip, cells toward the center (dashed box in the center panel) divide preferentially parallel to the axis of flattening (black arrow in the center panel). Before clefting, smooth muscle cells differentiate at the future clefting site, which helps to deform the flattened tip to complete clefting. (B) Schematics of clefting in the ureteric bud of mouse kidney. The clefting tip of kidney contains a single layer of cells that also flattens before clefting. For cell proliferation, premitotic cells delaminate from the single-layered epithelium, complete cell division in the lumen, and then reinsert into the epithelium. (C) Schematics of clefting in mouse salivary gland. In these images, the clefting tip (bud) is stratified. The outer tip cells are more columnar and more regularly arranged than the inner tip cells. The outer tip cells also move much faster than the inner tip cells. Shallow clefts form stochastically with ECM invasion into the outer layer of epithelium, and they widen and stabilize to complete clefting. Clefting in all three systems is accompanied by microscopic perforations in the basement membrane toward the tip and accumulation of basement membrane components away from the tip. For each organ, a critically essential growth factor regulator is listed, although others contribute. GDNF, glial cell–derived neurotrophic factor.