header logo image header logo text
Downloads Login
Research
General Information
ZIRC
ZFIN ID: ZDB-FIG-140624-13
Kelly et al., 2014 - A Pak1/Erk Signaling Module Acts through Gata6 to Regulate Cardiovascular Development in Zebrafish. Developmental Cell   29:350-9 Full text @ Dev. Cell
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Genes:
Antibodies:
Fish:
Knockdown Reagent:
Anatomical Terms:
Stage Range: 14-19 somites to Long-pec
PHENOTYPE:
Fish:
Knockdown Reagent:
Observed In:
Stage Range: 14-19 somites to Day 6

Fig. 2

Pak1 Morphants Display Cardiac and Neural Crest Defects

(A) Transcript expression of nkx2.5 shown by in situ hybridization at 20 somites.

(B and C) Pak1 morphant heart does not loop at 48 hpf as shown by (B) Tg(cmlc2:GFP) immunofluorescence and (C) staining with MF20 and S46 antibodies.

(D) Ventral images of 48 hpf Tg(Gata1:DsRed) show a cardiac outflow tract blockage with no blood flow throughout the head vasculature in the pak1 morphants (MO) compared to MM embryos (MM). Arrows point to cardiac outflow tract.

(E) Transverse sections of the cardiac outflow tract in MO- and MM-injected embryos stained with H&E. Arrows point to the cardiac outflow tract.

(F) Eln2 staining of WT and pak1 morphant embryos. Arrow indicates region of Eln positivity. Eln Abs, Eln2 antibodies.

(G) In situ hybridization for sox10 and foxD3 at 16–18 somites (lateral and cranial views). Arrows point to cranial neural crest expression in lateral view. In situ hybridization for crestin at the 16- to 18-somite stage and the 20-somite stage. Note lack of migration of neural crest in cranial view (arrows) and down the tail in the lateral view.

(H) Trunk images of pak1 morphants lacking melanophores at 2 dpf.

(I) In situ hybridization for mitfa.

(J) Quantification of melanophores at 2 dpf. Error bars indicate a significant difference with p < 0.0001. p < 0.0001.

(K) Quantification of melanophore migration in pak1 morphants compared to MM-injected embryos. Error bars indicate a significant difference with p < 0.0001.

(L) Cartilage staining (Alcian blue) at 6 dpf.

(M) Top: WT embryos were injected with either PID (Pak inhibitor) or PID L107F (inactive Pak inhibitor) mRNA. Embryos are shown at 48 hpf. Bottom: WT embryos were injected with expression vectors encoding PID or PID L107F driven by the sox10 promoter or the cmlc promoter, as indicated. Images were taken at 48 hpf.

Gene Expression Details
Gene Antibody Fish Conditions Stage Anatomy Assay
crestin WT control 14-19 somites neural crest cell ISH
20-25 somites neural crest cell ISH
WT + MO2-pak1 standard conditions 14-19 somites neural crest cell ISH
20-25 somites neural crest cell ISH
DsRed sd2Tg control Long-pec blood IFL
sd2Tg + MO2-pak1 standard conditions Long-pec blood IFL
EGFP twu34Tg control Long-pec heart IHC
twu34Tg + MO2-pak1 standard conditions Long-pec heart IHC
foxd3 WT control 14-19 somites neural crest cell ISH
WT + MO2-pak1 standard conditions 14-19 somites neural crest cell ISH
mitfa WT control Long-pec melanocyte ISH
WT + MO2-pak1 standard conditions Long-pec melanocyte ISH
nkx2.5 WT control 20-25 somites heart rudiment ISH
WT + MO2-pak1 standard conditions 20-25 somites heart rudiment ISH
sox10 WT control 14-19 somites neural crest cell ISH
WT + MO2-pak1 standard conditions 14-19 somites neural crest cell ISH
Antibody Labeling Details
Antibody Assay Fish Conditions Stage Qualifier Anatomy
Ab-MF20 IHC WT control Long-pec heart
IHC WT + MO2-pak1 standard conditions Long-pec heart
Ab-S46 IHC WT control Long-pec atrium
IHC WT + MO2-pak1 standard conditions Long-pec atrium
Ab1-elnb IHC WT control Long-pec bulbus arteriosus
IHC WT + MO2-pak1 standard conditions Long-pec Not Detected bulbus arteriosus
Phenotype Details
Fish Conditions Stage Phenotype
WT + MO2-pak1 standard conditions 14-19 somites neural crest cell migration process quality, abnormal
WT + MO2-pak1 standard conditions 20-25 somites neural crest cell migration process quality, abnormal
WT + MO2-pak1 standard conditions Long-pec atrium morphology, abnormal
WT + MO2-pak1 standard conditions Long-pec bulbus arteriosus morphology, abnormal
WT + MO2-pak1 standard conditions Long-pec bulbus arteriosus development process quality, abnormal
WT + MO2-pak1 standard conditions Long-pec heart looping decreased process quality, abnormal
WT + MO2-pak1 standard conditions Long-pec melanocyte decreased amount, abnormal
WT + MO2-pak1 standard conditions Long-pec melanocyte neural crest cell migration process quality, abnormal
WT + MO2-pak1 standard conditions Long-pec pigmentation decreased process quality, abnormal
WT + MO2-pak1 standard conditions Day 6 ceratohyal cartilage morphology, abnormal
WT + MO2-pak1 standard conditions Day 6 Meckel's cartilage morphology, abnormal
WT + MO2-pak1 standard conditions Day 6 pharyngeal arch morphology, abnormal
sd2Tg + MO2-pak1 standard conditions Long-pec blood accumulation heart, abnormal
Long-pec blood circulation decreased process quality, abnormal
twu34Tg + MO2-pak1 standard conditions Long-pec heart looping decreased process quality, abnormal
Acknowledgments:
ZFIN wishes to thank the journal Developmental Cell for permission to reproduce figures from this article. Please note that this material may be protected by copyright.

Reprinted from Developmental Cell, 29, Kelly, M.L., Astsaturov, A., Rhodes, J., Chernoff, J., A Pak1/Erk Signaling Module Acts through Gata6 to Regulate Cardiovascular Development in Zebrafish, 350-9, Copyright (2014) with permission from Elsevier. Full text @ Dev. Cell