Fig. 1

Mutations in scospondin Lead to the Absence of the Reissner Fiber and Defects in Body Axis Formation

(A) The Reissner fiber (RF) is localized in posterior ventricles of the brain and spinal central canal. At embryonic stages, SCO-spondin is secreted in the cerebrospinal fluid from the sub-commissural organ (SCO) below the posterior commissure (PC) and from the floor plate (FP) to form the fiber. Top: a scheme based on several immunohistochemistry experiments (below). Bottom: Z projection of a stack of a few lateral optical sections of the brain ventricles and rostral central canal (cc) of a 72 hpf embryo immunostained against the Reissner fiber (arrow). c, caudal; r, rostral. Scale bar represents 100 μm.

(B) 72 hpf scospondin^icm13/icm13 and scospondin^icm15/icm15 larvae showing curled-down phenotypes. Scale bar represents 500 μm.

(C) Proportion of curled-down phenotype over developmental time in both scospondin allele incrosses (mean ± SEM; n = 386 and 248 embryos for scospondin^icm13/icm13 and scospondin^icm15/icm15, respectively, from three independent clutches). The abnormal curvature of the body axis is detected from 30 hpf onward. Gastrulation-defective embryos were negligible (“other,” gray).

(D) Z projection of a stack of lateral optical sections (depth 4–5 μm) of the spinal cord immunostained against the Reissner fiber in control and scospondin mutants at 24 hpf (top) and 48 hpf (bottom). Both mutants are deprived of the Reissner fiber in the central canal from 24 hpf onward but immunoreactivity is detected in the floor plate of scospondin^icm15/icm15 (see insets on right panels highlighting the dotted-box regions) (n = 33; 63 control embryos, n = 7; 19 scospondin^icm15/icm15 embryos, n = 12; 33 scospondin^icm13/icm13 at 24; 48 hpf, respectively). d, dorsal; nc, notochord; v, ventral. Scale bars represent 40 μm.

(E) Z projection of stacks of dorsal optical sections (depth 23–26 μm) of 48 hpf forebrains (FBs) immunostained for acetylated tubulin (gray) and the Reissner fiber (green) show Reissner fiber material in SCO (double arrowheads) of control and scospondin^icm15/icm15 embryos but not scospondin^icm13/icm13. Arrows indicate axonal commissures. POC, post-optic commissure. Scale bars represent 30 μm.

See also Figure S1 and Table S1.