Figure 1.
Homozygous Dlg3 Mutations Cause Midgestational Embryonic Lethality
(A-D) Lateral views of (A) WT and (B-D) Dlg3 homozygous mutant embryos at E9.0-E9.5. Note the similarity of the phenotype observed with the two different Dlg3 mutant alleles: (B) Dlg3tm1Grnt and (C and D) Dlg3GtP038A02. White arrowheads indicate the tail bud (tb) truncation, absence of embryo turning, unfused allantois (all), and in some embryos forebrain (fb) and midbrain (mb) deletions, whereas the hindbrain (hb) remains intact. At E9.0, the GT β-galactosidase (lacZ) reporter gene is expressed ubiquitously.
(E-K) WT tetraploid embryos expressing a DsRed transgene were aggregated with (E) WT or (F-K) Dlg3GtP038A02/Y mESCs and isolated at E8.75-E9.0. (E-G) Red arrowheads indicate WT tetraploid cells (red) contributing to (E) the yolk sac (ys) or to (F and G) the node (n), midgut (mg), foregut (fg), notochord (nc), and prechordal plate (pcp). (H-K) Chimeras were immunostained with anti-RFP (red) in combination with anti-Foxa2 or anti-T (green) antibodies. DAPI (blue) has been used to mark all nuclei. Optical sections were taken at the level (H) of the foregut, (I) midgut, (J) hindgut (node outlined by broken oval), and (K) notochord. White arrows indicate tetraploid WT cells, expressing (H-J) the endoderm marker Foxa2 or (K) the notochord marker T.
(L-O) Whole-mount immunofluorescence combined with confocal imaging. Ventral views of the hindgut and node region in (L and M) WT and (N and O) Dlg3 homozygous mutant embryos at E8.5. White dashed lines and ovals mark the midline and the node, respectively. Embryos were stained with DAPI (blue) and with Foxa2 (green) and Arl13b (red) antibodies. At the level of the node, some Dlg3 mutants present a lateral expansion of Foxa2 and Arl13b-positive cells compared to the WT (compare i and ii).
Scale bars represent 300 μm in (A)-(G), 50 μm in (H)-(K), and 75 μm in (L)-(O).
See also Figures S1 and S2.