Fig. 8. Sulfated GAGs are necessary for transmission of the Nodal signal from the node to the LPM. (A,J) Schematic representation of proteoglycans from normal (A) or chlorate-treated (J) mouse embryos. A serine residue (yellow) of the core protein (orange) is attached to the GAG chain (blue curved line), which is sulfated (blue circles) under normal conditions but not in cells treated with chlorate. (B-D,K-M) Transverse frozen sections of embryos cultured to the six-somite stage in the absence (B-D) or presence (K-M) of 15 mM sodium chlorate were subjected either to immunofluorescence analysis with antibodies to HS (B,K) or to CS (C,L) or to staining with Alcian Blue (D,M). (E-G,N-P) In situ hybridization for Nodal (E,N), GDF1 (F,O) or Cryptic (G,P) mRNAs in embryos cultured in the absence (E-G) or presence (N-P) of chlorate. (H,I,Q,R) Expression vectors for Nodal and GFP were co-injected into the right LPM of embryos at the headfold stage, which were then cultured to the six-somite stage in the absence (H,I) or presence (Q,R) of chlorate. The cultured embryos were examined for GFP fluorescence (H,Q) and then subjected to in situ hybridization for Nodal mRNA (I,R). It should be noted that the region of Nodal expression was much broader than the area expressing GFP (I,R), which is due to a competence of LPM for Nodal auto-activation. (S) The number and percentage of embryos with (blue) or without (white) Nodal expression in LPM after culture in the absence or presence of chlorate. The difference between the two culture conditions was statistically significant (P<0.001) by the two-tailed Fisher's exact probability test. Scale bars: 100 µm in B-D,K-M; 200 µm in E-I,N-R.