Fig. 1. Rearrangements by pairs of newly formed hair cells.

(A) In a schematic depiction of a neuromast, four mechanoreceptive hair cells are surrounded by nonsensory supporting, mantle, and periderm cells. The asymmetric position of the kinocilium demarcates each hair cell's axis of planar polarity. (B) An apical view of a neuromast from a cldnB:lynGFP;ET4 doubly transgenic fish shows green fluorescence in the plasma membranes of all cells and in the cytoplasm of only the hair cells. This projection of several sections through the neuromast's apex shows the actin-rich cuticular plates of hair cells (arrow). RFP-tagged centrin (red) labels centrosomes and indicates the orientation of each hair cell by marking the position of its kinociliary basal body (arrowheads). The neuromast's plane of mirror symmetry is designated by the dotted line. In this and all subsequent figures the anterior direction is to the left and dorsal is upward. (C) In an image from a single plane in the same preparation, supporting cells and hair-cell precursors express only membrane-associated GFP and are morphologically indistinguishable. Cytoplasmic GFP demarcates the mature hair cells. (D) A time-lapse series focussed at or above the level of hair-cell nuclei documents the rearrangement of a pair of immature hair cells (arrowhead) in a cldnB:lynGFP;myo6b:mCherry^CAAX larva. Rearrangements are specific to the daughter hair cells arising from a recent division and result in the mirror-symmetrical orientation of the pair. After completing their rearrangement, the maturing cells assume their final shapes (arrows). (E) The red channel of the foregoing time-lapse series visually isolates mature hair cells labeled with membrane-associated mCherry. Note the low level of labeling (arrow) in the newborn pair undergoing rearrangement. (F) The narrow apices of the immature pair of hair cells can first be observed in the third panel (arrows). As the apical membranes expand to encompass hair bundles, the orientations of the newly formed pair (arrowheads) can be determined from the positions of the openings in their cuticular plates. The scale bars represent 10 µm in all panels.