header logo image header logo text
Downloads Login
Research
General Information
ZIRC
ZFIN ID: ZDB-FIG-121210-31
Wu et al., 2012 - Chemokine GPCR Signaling Inhibits β-Catenin during Zebrafish Axis Formation. PLoS Biology   10(10):e1001403 Full text @ PLoS Biol.
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Genes:
Antibody:
Fish:
Knockdown Reagent:
Anatomical Terms:
Stage Range: 256-cell to Prim-5
PHENOTYPE:
Fish:
Knockdown Reagent:
Observed In:
Stage: Prim-5

Fig. 2

Depletion of Ccr7 activity partially rescues axis formation in ichabod mutants.

(A) Penetrance of strongly ventralized phenotypes displayed by maternal β-catenin-2, ich, mutant embryos (a, 92%, n = 110) was reduced by injection of MO1-ccr7 (b–h, 10 ng; 40%, n = 98, five experiments). Arrows indicate partial axes and arrowheads indicate rudimentary head structures. (B) myod1 expression in uninjected (a) and MO1-ccr7-injected ich embryos (b,b2) revealed somitic tissue. Arrows indicate partial double axes. (C) In contrast to uninjected ich mutants (a,b,c), in MO1-ccr7-injected ich embryos, the organizer genes gsc (a2, n = 11/12) and chd (b2, n = 8/8) were expressed; while expression of the ventral gene, eve1, was significantly reduced (c2, n = 30/40). Animal views. (D) Lack of β-catenin nuclear accumulation, detected by immunostaining at 256-cell stage, in ich mutants (a), was suppressed in MO1-ccr7-injected ich embryos (b,c; n = 9/10, two experiments). (a2–c2) Higher magnification of boxed areas in a–c. (E) Dorsal domain of nuclear accumulation of β-catenin, detected by immunostaining at 256-cell stage in WT embryos (a), was expanded in ccr7 morphants (b; n = 6/13), while it was diminished in Ccr7 overexpressing embryos (c; n = 8/11). (a2–c2) Higher magnification of boxed areas in a–c. (F) Ectopic β-catenin nuclear accumulation, detected by immunostaining at 256–512-cell stage, in ich mutants injected with β-catenin RNA (a; 25 pg, n = 9/10), was suppressed by co-injecting ccr7 RNA (b; 150 pg, n = 7/10). (a2–b2) Higher magnification of boxed areas in a–b. (G) Ccr7 gain-of-function decreased both levels of endogenous β-catenin and ectopic β-catenin-GFP. Western blotting of β-catenin and GFP protein from uninjected control, β-catenin-GFP RNA (10 pg) injected, or β-catenin-GFP RNA (10 pg)/ccr7 RNA (200 pg) co-injected embryos (all at 3–3.3 hpf). Graphs below show the relative protein level (signal intensity) quantified from three separate immunoblots. * p<0.05.

Gene Expression Details
Gene Antibody Fish Conditions Stage Qualifier Anatomy Assay
chrd ctnnb2p1 standard conditions Shield Not Detected shield ISH
ctnnb2p1 + MO2-ccr7 standard conditions Shield whole organism anatomical region ISH
eve1 ctnnb2p1 standard conditions Shield whole organism ventral region ISH
ctnnb2p1 + MO2-ccr7 standard conditions Shield whole organism ventral region ISH
gsc ctnnb2p1 standard conditions 50%-epiboly Not Detected presumptive shield ISH
ctnnb2p1 + MO2-ccr7 standard conditions 50%-epiboly whole organism anatomical region ISH
myod1 ctnnb2p1 standard conditions Prim-5 somite ISH
ctnnb2p1 + MO2-ccr7 standard conditions Prim-5 somite ISH
Antibody Labeling Details
Antibody Assay Fish Conditions Stage Qualifier Anatomy
Ab1-ctnnb IHC WT standard conditions 256-cell blastomere nucleus
IHC 256-cell whole organism dorsal region
IHC WT + MO2-ccr7 standard conditions 256-cell blastomere nucleus
IHC 256-cell whole organism dorsal region
IHC ctnnb2p1 standard conditions 256-cell Not Detected blastomere nucleus
IHC 256-cell whole organism dorsal region
IHC ctnnb2p1 + MO2-ccr7 standard conditions 256-cell blastomere nucleus
IHC 256-cell whole organism dorsal region
Phenotype Details
Fish Conditions Stage Phenotype
ctnnb2p1 standard conditions Prim-5 caudal fin increased size, abnormal
Prim-5 head absent, abnormal
Prim-5 trunk decreased size, abnormal
Prim-5 whole organism wholly ventralized, abnormal
ctnnb2p1 + MO2-ccr7 standard conditions Prim-5 axis duplicated, abnormal
Prim-5 head malformed, abnormal
Prim-5 trunk malformed, abnormal
Prim-5 whole organism wholly ventralized, abnormal
Acknowledgments:
ZFIN wishes to thank the journal PLoS Biology for permission to reproduce figures from this article. Please note that this material may be protected by copyright. Full text @ PLoS Biol.