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Fig. S5 Decreased retrograde flow via changes in contractility affects valve morphogenesis. (A–H) Flow pattern at 48 hpf and associated confocal sections of the valve-forming region at 96 hpf in (A) control, (B) cx36.7 (see also Video S6), (C) myh6 (Video S6), (D) ttna (Video S6) knock downs, and (E) in the silent heart (sih) mutants. myh6, ttna, and sih inactivation leads to a dramatic decrease in the RFF and valve defects, whereas cx36.7 knock down has an almost normal RFF and valves compared to the control. (F–H) klf2a expression in (F) control, (G) cx36.7, and (H) myh6 morphants. Absence of klf2a expression was observed in myh6 morphants (41%, n = 36) (H), but normal expression levels were observed in cx36.7 morphants (75%, n = 50) (G). These two populations were significantly different (α = 0.1). (I) Energy expenditure comparison between control, gata1, and myh6 morphants during the retrograde, anterograde, or both flow direction phases. The apparition of valve dysgenesis coincides with a low energy expenditure during phases of retrograde flow rather than a reduction of the overall energy expenditure during phases of anterograde and retrograde flow. (J) Proportionally decreased RFF through treatment with cx36.7, myh6, or ttna MOs leads to an increase in valve defects. The maximal effect is observed in no flow (sih) or no RFF (ttna) conditions.