ZFIN ID: ZDB-FIG-140422-44
Holly et al., 2014 - Sfrp1a and Sfrp5 function as positive regulators of Wnt and BMP signaling during early retinal development. Developmental Biology   388(2):192-204 Full text @ Dev. Biol.
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Genes:
Fish:
Condition:
Knockdown Reagents:
Anatomical Terms:
Stage Range: 10-13 somites to Prim-5
PHENOTYPE:
Fish:
Knockdown Reagents:
Observed In:
Stage: Prim-5

Fig. 6

sfrp1a/sfrp5 positively regulate BMP signaling during retinal development. Given the observed alterations in dorsal retina marker gene expression, we sought to investigate the activity of BMP signaling in Sfrp depleted retinas. We measured BMP pathway activity by employing a BMP responsive transgene, Tg(BMPRE-AAV.Mlp:eGFP), which is abbreviated BMPRE:GFP. In situ hybridization for GFP mRNA, or fluorescence microscopy results indicate that compared to WT, sfrp1a/5 MO co-injected embryos display a marked decrease in GFP signal at 28 hpf (A–D). Measurements of mean retinal fluorescence intensity indicate a significant decrease in GFP signal in sfrp1a/5 MO injected embryos (E, NNN, p<0.0001). To assess expression of BMP ligand mRNAs, we conducted in situ hybridization of BMP ligands bmp2b, bmp4 and gdf6a at 28 hpf. Compared to WT, sfrp1a/5 MO injected embryos display an increase in BMP ligand expression (F–G, I–J, L–M). Conversely, inhibition of Wnt signaling by heat shock activation of dkk1b, greatly reduces expression of bmp2b, bmp4 and gdf6a at 28 hpf (H,K,N). qRT-PCR of bmp2b and bmp4 confirmed a 1.38 and 1.47 fold change in expression respectively (O, NNN, p<0.0001). To assess the functions of Sfrp proteins in the BMP-dependent initiation phase of retinal patterning, we conducted in situ hybridization for bambia and tbx5a and ventral marker vax2 during the initiation of dorsal eye patterning at 15 hpf. Compared to WT, sfrp1a/5 MO injected embryos display a decrease in tbx5a and bambia expression levels and domain while vax2 appears unaffected (P–U).

Gene Expression Details
Gene Antibody Fish Conditions Stage Anatomy Assay
bambia WT standard conditions 10-13 somites optic vesicle ISH
WT + MO2-sfrp5 + MO3-sfrp1a standard conditions 10-13 somites optic vesicle ISH
bmp2b WT standard conditions Prim-5 optic cup ISH
WT + MO2-sfrp5 + MO3-sfrp1a standard conditions Prim-5 optic cup ISH
w32Tg heat shock Prim-5 optic cup ISH
bmp4 WT standard conditions Prim-5 optic cup ISH
WT + MO2-sfrp5 + MO3-sfrp1a standard conditions Prim-5 optic cup ISH
w32Tg heat shock Prim-5 optic cup ISH
EGFP mw29Tg standard conditions Prim-5 optic cup ISH
Prim-5 optic cup IFL
mw29Tg + MO2-sfrp5 + MO3-sfrp1a standard conditions Prim-5 optic cup ISH
Prim-5 optic cup IFL
gdf6a WT standard conditions Prim-5 optic cup ISH
WT + MO2-sfrp5 + MO3-sfrp1a standard conditions Prim-5 optic cup ISH
w32Tg heat shock Prim-5 optic cup ISH
tbx5a WT standard conditions 10-13 somites optic vesicle ISH
WT + MO2-sfrp5 + MO3-sfrp1a standard conditions 10-13 somites optic vesicle ISH
vax2 WT standard conditions 10-13 somites optic vesicle ISH
WT + MO2-sfrp5 + MO3-sfrp1a standard conditions 10-13 somites optic vesicle ISH
Antibody Labeling Details No data available
Phenotype Details
Fish Conditions Stage Phenotype
mw29Tg + MO2-sfrp5 + MO3-sfrp1a standard conditions Prim-5 BMP signaling pathway increased process quality, abnormal
Acknowledgments:
ZFIN wishes to thank the journal Developmental Biology for permission to reproduce figures from this article. Please note that this material may be protected by copyright.

Reprinted from Developmental Biology, 388(2), Holly, V.L., Widen, S.A., Famulski, J.K., and Waskiewicz, A.J., Sfrp1a and Sfrp5 function as positive regulators of Wnt and BMP signaling during early retinal development, 192-204, Copyright (2014) with permission from Elsevier. Full text @ Dev. Biol.