Fig. 3

ela Knockout Zebrafish Have Severe Cardiovascular and Endoderm Defects

(A) Null ela^br21 larvae display pericardial edema (white arrowheads) and accumulated erythrocytes (red arrowheads), and have no blood circulation. Variable posterior anomalies are observed, including loss of ventral fin (black arrowheads), tailbud duplications (inset), and extreme tail/trunk truncations (bottom embryo).

(B) Loss of ela causes severe cardiac anomalies ranging from mild heart dysplasia to total heart agenesis, as shown by hematoxylin and eosin staining on sections from the top and bottom embryos in (A).

(C) Null ela^br13 fish have a severe reduction of cmlc1 expression, which marks the developing heart.

(D) Null ela^br13 fish display increased hematopoiesis as judged by the upregulation of scl, a marker of blood precursors.

(E) Classification of null ela^br13 larvae with varying degrees of tail defects. Class I larvae are defined as having pericardial edema and tail blood clot. Class 2 larvae have ventral fin defects or tailbud duplications in addition to class 1 phenotypes. Class 3 larvae have all phenotypes of class 1 combined with mild to severe tail/trunk truncations.

(F) Percentages of ela mutant fertile adults that were obtained from heterozygous intercrosses using all three alleles.

(G) Relative to WT embryos, homozygous null ela^br13 embryos show convergence-extension defects resulting in delayed blastopore closure and thickened notochord (insets) as indicated by altered bra expression.

(H) The loss of ela causes defective migration of mediolateral gata5-expressing cells, which mark mesendoderm cells that guide heart progenitors to the anterior lateral plate mesoderm.

(I) ela mutant embryos show reduced foxa2 expression compared with WT embryos.

(J) ela mutant embryos show a more compact sox17 expression pattern compared with WT embryos, whereas sox17+ forerunners cells are not affected.

(K) ela null embryos have approximately 40% less sox17+ cells than their heterozygous siblings at 75% epiboly.

(L) At 75% epiboly, ela^br21 and ela^br15 mutant embryos exhibit sox17 expression defects similar to those observed for ela^br13 (see also Figure S1).