Fig. 4

Spatio-temporal distribution of the zebrafish PLAGL2 transcripts from 14 somite stage to 48 hpf and comparison with the proliferative marker PCNA. Whole mount in situ hybridization was performed using either PLAGL2, PCNA or Dlx3 antisense probes (except in L,M whole mount immunohistochemistry with anti-phosphohistone H3 antibody). The developmental stage is indicated in each panel. (A) Posterior view of a 14 somite stage embryo. Lateral (B,C,D,F,N,O,Q,R,V,W,X) or dorsal (E,G,H,I,P,S,Y) views of zebrafish embryos with anterior to the left. Transverse sections (5 μm) at the level of the midbrain in J,K,L,M, hindbrain in T or at the trunk level in U. (A,B) At 14 somite stage, PLAGL2 is expressed ubiquitously at low levels and more strongly in the lateral plate mesoderm and in the developing brain and eyes. (C) At 18 somite stage, staining is seen throughout the whole embryo and more intensely in the developing brain and the lateral mesoderm. (D,E) At 20 somite stage, PLAGL2 expression is evident in the anteriomost part of the embryos and more specifically in the ventricular zone. (F,G,H) Staining is evident in the brain, the otic vesicles and the eyes at 24 hpf. (J) A microtome section confirms the preferential expression of PLAGL2 in the apical region of the ventricular zone of the brain. (L) These PLAGL2-labelled cells correspond to a proliferative zone as shown by the high number of mitotically active cells (stained by anti-phospho H3 antibody) at the ventricular border (brown spots in L). (M) H&E-stained section showing the highly cytoplasmic content of the cells lining the brain ventricle. Expression of PLAGL2 at 36 and 48 hpf is evident in the brain ventricles (N,O,P,R), the ganglion cell layer of the retina (S), the pharyngeal region (double white asterisks in O,R,V), the otic vesicles and the roof of the neural tube (black asterisk in Q,R,U). A signal is also observed in the developing fin buds (black arrows in R,S,T) and the developing vasculature at 36 and 48 hpf (Q,R). (W) Expression of Dlx3 in pharyngeal arch primordia, similar to that observed for PLAGL2 at 48 hpf (V). (Q, R) Views of 24 and 48 hpf embryos stained with PCNA antisense probe. Note the striking similarity of the signal observed between PCNA and PLAGL2 at the same stage (compare R and X, G and Y). Double black asterisks indicate the lateral plate mesoderm. White arrows denote the ventricular surface of the brain. Black arrows show the pectoral fin buds. Black asterisk indicates the top of the neural tube. White arrowhead points to the ganglion cell layer of the retina. Double white asterisks denote the developing pharyngeal arches. OV, otic vesicles; AV, axial vasculature.