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Figure 1. Bilateral Xnr1 in Somitic Gastrocoel Roof Plate Cells Acts Downstream of Flow(Aa–Bb) Xnr1 expression during flow (stage 17) in 1–3 rows on the lateral margins of the gastrocoel roof plate (GRP) in prospective somitic cells. In situ hybridization was performed with probes specific for Xnr1 (Aa) and MyoD (Ba and Bb). Histological sections at comparable levels (indicated in Aa) identify Xnr1 (Ab and Ac) cells as MyoD-positive somitic cells (Ba and Bb). For assignment of boundaries (broken lines), please see Figure S1A.(Ca–Cc) Xnr1 cells harbor unpolarized monocilia. The image shows an overlay of in situ hybridization signal and scanning electron micrograph of same specimen (Ca). Higher magnification in (Cb) reveals central cilia (yellow) on Xnr1-positive cells, as opposed to polarized cilia (white) at the center of the GRP. Evaluation of the entire width of GRP (Cc) demonstrates the unpolarized nature of cilia on Xnr1 cells on either side (green dot represents a cilium with unclear polarization).(Da) Wild-type (WT) leftward flow in morphants injected bilaterally with Xnr1-MO (cf. Movie S1). Particle movement in a representative specimen is visualized by gradient time trails (GTTs; cf. Experimental Procedures for details), in which the color gradient from green to red represents 25 s (cf. color bar). Targeted area is indicated by red lines (corresponding to coinjected lineage tracer rhodamine-B dextran).(Db) Quantification of results from 8 embryos, representing 2103 particles. Morphants display a robust leftward flow (3.2 ± 1.9 μm/s), as demonstrated by the distribution of mean particle directionality (wind rose) and WT p = 0.8 [38].The following abbreviations are used: a, anterior; bp, blastopore; d, dorsal; e, endoderm; hyGRP, prospective hypochordal cells; l, left; LECs, lateral endodermal crest cells; no, notochord; p, posterior; r, right; s, presomitic mesoderm; SEM, scanning electron micrograph; sGRP, prospective somitic cells, v, ventral. The color gradient in (Da) represents 25 s. Boxes in (Ab), (Ba), (Ca), and (Cb) indicate areas of higher magnification in (Ac), (Bb), (Cb), and (Cc). Scale bars in (Ca), (Cb), and (Da) represent 100 μm, 10 μm, and 50 μm, respectively. |
