Extended Data Fig. 7: Contractility-induced spatial segregation of cardiomyocytes is necessary and sufficient to trigger Notch reporter expression in CL cardiomyocytes.

a, Representative MIP of 48 hpf heart; Notch reporter expression is observed only in the endocardium (n = 15/15 hearts). b, b’, Hearts of DMSO- (n = 20) or alfacalcidol- (Alfa, n = 27) treated 60 hpf zebrafish embryos; representative mid-sagittal confocal images (b), and quantification (b’). c, c’ Hearts of DMSO- (n = 16) or MEK inhibitor- (n = 20) treated 65 hpf zebrafish embryos; representative mid-sagittal confocal images (c), and quantification (c’). d–d’, Hearts of nrg2a:mRFP;TP1:VenusPest 100 hpf zebrafish larvae injected with CA-MYL9-BFP (d), and quantification (d’, nrg2a^−/−, n = 40; nrg2a^−/− + CA-MYL9, n = 56). e, e’, Hearts of tnnt2a MO-injected 72 hpf zebrafish larvae (n = 22) transplanted with myl7:CA-MYL9-mScarlet blastomeres (n = 27) (e), and quantification (e’). f, f’, Representative mid-sagittal confocal image of 62 hpf hearts (f), and quantification of apical domain length in Notch⁺ CL and Notch⁻ delaminating cardiomyocytes (f’, n = 17). Data are mean ± s.d. Two-tailed Student’s t-test (b’, c’, d’, f’); two-tailed Mann–Whitney U-test (e’). n refers to the number of hearts (b’, c’, d’, e’) or number of cardiomyocytes (f’). Red arrowheads, Notch⁺ cardiomyocytes; yellow arrowheads, Notch⁺ endocardial cells; white arrowheads, CL cardiomyocytes; asterisks, delaminating or TL cardiomyocytes. Scale bars, 50 μm. For more details on statistics and reproducibility, see Methods.

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