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. 2021 Dec 23;10:e74336. doi: 10.7554/eLife.74336

Figure 5. POU-IV is necessary for hair cell differentiation in the sea anemone.

(A-I) Confocal sections of oral tentacles of wildtype (F2 pou-iv +/+, A-C), heterozygous (F2 pou-iv +/-, D-F), and homozygous pou-iv mutant (F2 pou-iv -/-, G-I) Nematostella vectensis polyps. Filamentous actin is labeled with phalloidin (Pha), and nuclei are labeled with DAPI (dapi). In all panels, the distal end of the tentacle is to the top. (A, D, G) Sections through the tentacle. (B, C, E, F, H, I) Sections through hair bundles/ciliary cones at the tip of tentacles. Ciliary cones occur on the epithelial surface of the tentacle regardless of the genotype (arrowheads in A, D, G). However, stereovilli (st) with rootlets (rl) characteristic of mechanosensory hair cells are observed in wildtype (B) and heterozygous (E) siblings, but not in homozygous pou-iv mutants whose ciliary cones contain microvilli without prominent actin rootlets (mv in H). Arrowheads in C and F indicate spaces occupied by cnidocysts in wildtype and heterozygous siblings, respectively, which are absent in pou-iv homozygous mutants (I; Figure 5—figure supplement 1). (J–L) Electron microscopic sections of a hair cell of a F2 pou-iv +/+ polyp (J) and an epithelial cell with hair-cell-like morphologies in an F2 pou-iv -/- polyp (K, L). In all panels, apical cell surfaces face up. K and L are sections of the same cell at different levels. The hair-cell-like epithelial cell of the mutant has a central apical cilium surrounded by a collar of rootlet-less microvilli (mv in K, L), which are encircled by microvilli of the adjacent support cells (mvs in L), forming a ciliary cone. It also has numerous clear vesicles (ve in L) in the cytoplasm, characteristic of hair cells (ve in J; Figure 1G). Support cells of mutants are morphologically indistinguishable from those of wildtype animals, having characteristic large electron-dense vacuoles (va in L) in addition to apical microvilli (mvs in L) that contribute to the ciliary cone/hair bundle. Consistent with light microscopy data (A–C, G–I), stereovillar rootlets (rlst) are absent in the F2 pou-iv -/- polyp, but are present in hair cells of their wildtype siblings (J). In K and L, regions of apical cytoplasm where stereovillar rootlets would normally be observed are boxed with dotted purple lines. Abbreviations: ms muscle fibers; ec ectoderm; st stereovilli; ci cilium; rlst stereovillar rootlets. Scale bar: 10 µm (A-I); 2 µm (insets in B, E, H); 500 nm (J–L).

Figure 5.

Figure 5—figure supplement 1. pou-iv mutants lack mature cnidocytes.

Figure 5—figure supplement 1.

Electron microscopic sections of tentacular ectoderm of an F2 pou-iv +/+ polyp (A) and F2 pou-iv -/- polyp (B). Note the occurrence of numerous cnidae – nematocysts (nc) and spirocysts (sp) – in A that are absent in B. Abbreviations: nu, nucleus; ec, ectoderm; en, endoderm; ms, muscle fibers; mg, mesoglea. Scale bar: 2 µm.
Figure 5—figure supplement 2. F2 pou-iv wildtype siblings develop hair cells with stereovillar rootlets.

Figure 5—figure supplement 2.

Electron microscopic sections of hair cells of an F2 pou-iv +/+ polyp. In all panels, apical cell surfaces face up. Note the presence of stereovillar rootlets (arrowheads in A and B). Abbreviations: st, stereovilli; ci, cilium. Scale bar: 500 nm.
Figure 5—figure supplement 3. Hair-cell-like cells of pou-iv mutants have ciliary rootlets.

Figure 5—figure supplement 3.

Electron microscopic sections of a hair-cell-like cell of an F2 pou-iv -/- polyp, characterized by ciliary-cone-associated microvilli (mv). The apical cell surface faces up. Note the presence of a ciliary rootlet (rlci). Abbreviations: ve, vesicles. Scale bar: 500 nm.