A model for altered asymmetric nodal gene expression in pkd2-deficient zebrafish embryos. (A-C) RNA in situ hybridization for spaw and lefty1/2 in 20-somite embryos that were flat-mounted in order to see all domains of expression. All panels show lefty1 staining in the midline. White arrows mark the anterior and posterior areas of spaw expression and white arrowheads mark lefty expression in the heart field. (D-F) Schematic interpretations of A-C (spaw in blue, lefty in gray). In a wild-type embryo (A,D), spaw is expressed in a posterior-to-anterior pattern on the LPM from 10 somites through 22 somites. lefty1/2 is activated in the left heartfield, the anterior portion of the LPM, beginning at 18 somites. RNA in situ hybridizations for spaw and lefty1/2 together in cup mutants (B,C) reveal a role for the anterior domain of spaw in the activation of the asymmetric expression of lefty1/2 in the cardiac LPM. In all embryos scored, lefty expression is only apparent in embryos where spaw reaches the anterior LPM and it always correlates with the side of spaw expression (A,B,D,E). cup mutants primarily exhibit bilateral expression of spaw to varying degrees of posterior to anterior extension (B,C,E,F). Bilateral anterior expression of spaw results in randomized lefty expression, where the heartfield will have left, right, bilateral, or absent lefty. Bilateral posterior expression of spaw always correlates with loss of lefty expression in the LPM (C,D). Therefore, because pkd2 zebrafish mutants display a defect in spaw propagation, lefty-expressing cells are largely absent from the LPM. In zebrafish, we propose that pkd2 is acting at Kupffer's vesicle (KV) to restrict spaw to the left side of the embryo and that pkd2 plays an additional role in the proper propagation of spaw anteriorly in the LPM (D). pkd2 could also restrict nodal to the left by acting at Kupffer's vesicle to repress nodal activation and propagation on the right LPM (D).
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