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ZFIN ID: ZDB-FIG-180817-27
Hofsteen et al., 2018 - ALPK2 Promotes Cardiogenesis in Zebrafish and Human Pluripotent Stem Cells. iScience   2:88-100 Full text @ iScience
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Genes:
Fish:
Knockdown Reagents:
Anatomical Term:
Stage: Prim-5
PHENOTYPE:
Fish:
Knockdown Reagents:
Observed In:
Stage Range: Prim-5 to Day 4

Fig. 2

Alpk2 Is Essential for Zebrafish Cardiac Development

(A) Zebrafish Alpk2 locus and targeted region by splice-blocking morpholinooligonucleotides (MO). * denotes stop codon.

(B–D) RT-PCR and Sanger sequencing demonstrating splice-blocking MO caused alternative splicing resulting in a 131-base pair deletion and premature stop codon. * denotes stop codon.

(E and F) Zebrafish injected with negative control MO (E) or Alpk2 MO (F) at 24 hr post fertilization (hpf).

(G and H) Transcript analysis (G) and heart beating rate (H) of control and Alpk2 morphants at 24 and 48 hpf, respectively (N = 5, 15–22 pooled embryos per N).

(I–O) Representative images of control and Alpk2 MO-injected hearts carrying a transgene for cmlc2:DsRed-nuc to quantify cardiomyocyte nuclei (K) at 51 hpf (N = 7–9). Red denotes cmlc2:DsRed-nuc and magenta is antibody staining for activated leukocyte cell adhesion molecule (ALCAM). Representative bright-field images of control MO (L) and Alpk2 MO (M) injected zebrafish at approximately 72 hpf. Representative images of control MO (N) and Alpk2 MO (O) injected zebrafish hearts carrying a transgene (cmlc2:DsRed-nuc) to denote cardiac morphology at approximately 72 hpf.

(P and Q) Confocal images of control and Alpk2 morphant fish carrying myl7:GFP and tcf21:DsRed at 96 hpf. White arrowheads denote tcf21+ epicardial cells on the heart ventricle. N = 3; independent biological replicates with 45–76 pooled animals per N unless otherwise noted.

Data are mean ± SEM; * denotes p ≤ 0.05.

Gene Expression Details
Gene Antibody Fish Conditions Stage Anatomy Assay
atp2a2a AB + MO1-alpk2 + MO2-alpk2 standard conditions Prim-5 whole organism RTPCR
cacna1c AB + MO1-alpk2 + MO2-alpk2 standard conditions Prim-5 whole organism RTPCR
cx43 AB + MO1-alpk2 + MO2-alpk2 standard conditions Prim-5 whole organism RTPCR
gata4 AB + MO1-alpk2 + MO2-alpk2 standard conditions Prim-5 whole organism RTPCR
mef2ca AB + MO1-alpk2 + MO2-alpk2 standard conditions Prim-5 whole organism RTPCR
mef2cb AB + MO1-alpk2 + MO2-alpk2 standard conditions Prim-5 whole organism RTPCR
myl7 AB + MO1-alpk2 + MO2-alpk2 standard conditions Prim-5 whole organism RTPCR
tnnt2a AB + MO1-alpk2 + MO2-alpk2 standard conditions Prim-5 whole organism RTPCR
Antibody Labeling Details No data available
Phenotype Details
Fish Conditions Stage Phenotype
AB + MO1-alpk2 + MO2-alpk2 standard conditions Prim-5 heart tube morphology, normal
Prim-5 whole organism mef2ca expression decreased amount, abnormal
Prim-5 whole organism cx43 expression decreased amount, abnormal
Prim-5 whole organism mef2cb expression decreased amount, abnormal
Prim-5 whole organism tnnt2a expression decreased amount, abnormal
Prim-5 whole organism gata4 expression decreased amount, abnormal
Prim-5 whole organism myl7 expression decreased amount, abnormal
Prim-5 whole organism atp2a2a expression decreased amount, abnormal
Prim-5 whole organism cacna1c expression decreased amount, abnormal
Long-pec cardiac muscle cell decreased amount, abnormal
Long-pec heart contraction decreased rate, abnormal
Protruding-mouth heart malformed, abnormal
Protruding-mouth pericardium edematous, abnormal
Protruding-mouth yolk syncytial layer edematous, abnormal
pd37Tg ; f1Tg + MO1-alpk2 + MO2-alpk2 standard conditions Day 4 ventricular epicardium absent, abnormal
Acknowledgments:
ZFIN wishes to thank the journal iScience for permission to reproduce figures from this article. Please note that this material may be protected by copyright. Full text @ iScience