ZFIN ID: ZDB-FIG-130801-32
Wang et al., 2013 - miR-34b regulates multiciliogenesis during organ formation in zebrafish. Development (Cambridge, England)   140(13):2755-2764 Full text @ Development
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Genes:
Antibody:
Fish:
Condition:
Knockdown Reagents:
Anatomical Terms:
Stage Range: Prim-25 to Protruding-mouth
PHENOTYPE:
Fish:
Knockdown Reagents:
Observed In:
Stage Range: Prim-25 to Protruding-mouth

Fig. 5 miR-34b regulates multiciliogenesis and kidney morphogenesis through cmyb. (A) WISH analysis of cmyb expression at 36 hpf. The arrow and dashed line show the region of MCCs. (B) QRT-PCR analysis of cmyb expression in MCCs that were isolated from 3-dpf Tg(cmyb:EGFP) embryos by flow cytometry. The expression level was normalized to that of β-actin 1. (C) A 2-dpf Tg(cmyb:EGFP) embryo shows enhanced fluorescence in MCCs (arrow) after miR-34b knockdown. (D) Statistical analysis of A and C showing normal versus high expression of cmyb. (E) The percentage of cmyb:EGFP+ cells with cilia bundles was calculated for each embryo and analyzed statistically at 36 hpf. (F,G) A magnified lateral view of 3-dpf embryos after in situ hybridization with rfx2 or pax2 probe shows the distribution of MCCs and the state of proximal tubule convolution. Arrows (F) indicate rfx2+ MCCs. The DT angle was measured. (H) The percentage of cmyb:EGFP+ cells with cilia bundles was calculated for each Tg(hsp70:cmyb) embryo and analyzed statistically at 36 hpf (I) A magnified lateral view of 2-dpf Tg(hsp70:cmyb) embryos after in situ hybridization with rfx2 probe shows the distribution of MCCs. Arrows indicate rfx2+ MCCs. The percentage of embryos with accumulated rfx2+ MCCs (Accumulated) in F and I was calculated. (J) WISH analysis of cmyb expression at 2 dpf in the olfactory placode (arrows). A representative image of a 3-dpf Tg(cmyb:EGFP) embryo shows enhanced fluorescence of the olfactory placode (arrows) after miR-34b knockdown. The number of embryos with normal (top pictures) or high expression level (bottom pictures) of cmyb among all analyzed embryos is indicated (bottom right). (K) The ciliary (anti-acetylated-tubulin staining, red) morphology of the olfactory placode at 3 dpf in normal embryos, miR-34b morphants and embryos injected with both miR-34b MO and cmyb MO. Cilia length was analyzed at three positions (arrows) for each embryo, and the average cilia length is listed. Each circle/triangle/cross represents an embryo. Mean ± s.d. ***P<0.0001 (unpaired t-test). Scale bar: 10 μm.

Gene Expression Details
Gene Antibody Fish Conditions Stage Anatomy Assay
EGFP zf169Tg standard conditions Long-pec pronephros multi-ciliated epithelial cell IFL
Protruding-mouth peripheral olfactory organ IFL
zf169Tg + MO1-mir34b standard conditions Long-pec pronephros multi-ciliated epithelial cell IFL
Protruding-mouth peripheral olfactory organ IFL
myb WT standard conditions Prim-25 pronephros multi-ciliated epithelial cell ISH
Long-pec peripheral olfactory organ ISH
WT + MO1-mir34b standard conditions Prim-25 pronephros multi-ciliated epithelial cell ISH
Long-pec peripheral olfactory organ ISH
pax2a WT standard conditions Protruding-mouth pronephros epithelium ISH
WT + MO1-mir34b standard conditions Protruding-mouth pronephros epithelium ISH
WT + MO1-mir34b + MO2-myb standard conditions Protruding-mouth pronephros epithelium ISH
rfx2 WT standard conditions Protruding-mouth pronephros multi-ciliated epithelial cell ISH
WT + MO1-mir34b standard conditions Protruding-mouth pronephros multi-ciliated epithelial cell ISH
WT + MO1-mir34b + MO2-myb standard conditions Protruding-mouth pronephros multi-ciliated epithelial cell ISH
hkz03tTg standard conditions Long-pec pronephros multi-ciliated epithelial cell ISH
hkz03tTg heat shock Long-pec pronephros multi-ciliated epithelial cell ISH
Antibody Labeling Details
Antibody Assay Fish Conditions Stage Anatomy
Ab1-tuba IHC WT standard conditions Protruding-mouth peripheral olfactory organ cilium
IHC WT + MO1-mir34b standard conditions Protruding-mouth peripheral olfactory organ cilium
IHC WT + MO1-mir34b + MO2-myb standard conditions Protruding-mouth peripheral olfactory organ cilium
Phenotype Details
Fish Conditions Stage Phenotype
WT + MO1-mir34b standard conditions Prim-25 multi-ciliated epithelial cell cilium physical object quality, abnormal
Protruding-mouth multi-ciliated epithelial cell accumulation pronephric duct, abnormal
Protruding-mouth olfactory epithelium cilium shortened, abnormal
Protruding-mouth pronephric proximal convoluted tubule closed, abnormal
WT + MO1-mir34b + MO2-myb standard conditions Prim-25 multi-ciliated epithelial cell cilium normal, normal
Protruding-mouth multi-ciliated epithelial cell accumulation pronephric duct, normal
Protruding-mouth olfactory epithelium cilium shortened, normal
Protruding-mouth pronephric proximal convoluted tubule closed, normal
hkz03tTg standard conditions Prim-25 multi-ciliated epithelial cell cilium physical object quality, abnormal
Long-pec multi-ciliated epithelial cell accumulation pronephric duct, abnormal
Acknowledgments:
This image is the copyrighted work of the attributed author or publisher, and ZFIN has permission only to display this image to its users. Additional permissions should be obtained from the applicable author or publisher of the image. Full text @ Development
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