Figure 4

Figure 4

Functional analysis identified atp1b3b and colec10 as new players in heartbeat maintenance

(A) Expression of atp1b3b and colec10 within the myocardial subclusters. The highest expression for both genes was observed within Sinoatrial CMs cluster.

(B) Scheme illustrating gene loci targeted by CRISPR/Cas9 strategy. The combination of three sgRNAs were utilized for atp1b3b and colec10 knockout generation. Control embryos were injected with a scrambled sgRNA.

(C) Barplot summarizing observed phenotypes from all replicates for each gene knockout. Numbers within each section indicate the number of embryos associated with a particular phenotype.

(D) Fluorescent images showing heart looping defect in colec10 knockouts as compared to uninjected embryos (WT represents embryos on Tg(myl7:EGFP-Hsa.HRAS)^s883 background at 48 hpf).

(E) Heartbeat rate comparison between appropriate knockouts, uninjected wild-type control, and scrambled-injected embryos (significance values according to Wilcoxon rank-sum test).

(F) Representative image showing morphology of atp1b3b and colec10 knockout embryos.

(G) Whole mount in situ hybridization of atp1b3b.

(H) Hybridization chain reaction in situ for atp1b3b performed on 3 dpf larvae. Note the restricted heart expression within the sinoatrial region (G, H). Detailed single confocal plane images provided in the Figure S4A.