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ZFIN ID: ZDB-FIG-090113-93
Huang et al., 2009 - The embryonic expression patterns and the knockdown phenotypes of zebrafish ADP-ribosylation factor-like 6 interacting protein gene. Developmental dynamics : an official publication of the American Association of Anatomists   238(1):232-240 Full text @ Dev. Dyn.
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Genes:
Antibodies:
Fish:
Condition:
Knockdown Reagent:
Anatomical Terms:
Stage Range: Long-pec to Day 4
PHENOTYPE:
Fish:
Condition:
Knockdown Reagent:
Observed In:
Stage Range: Long-pec to Day 4

Fig. 3 A-D: The histological examination of zebrafish retinae. Histological section and hematoxylin-eosin staining of retinae were performed on wild-type embryos and arl6ip-morphants at 72 and 96 hpf. Unlike the wild-type embryos (A,C), the neuronal differentiation is severely delayed, resulting in no formation of retinal layers in the arl6ip-morphants, both at 72 (B) and 96 hpf (D). In addition, retina development of arl6ip-morphants was markedly retarded. GCL, ganglion cell layer; INL, inner nuclear layer; L, lens; MO, arl6ip-morphant; ONL, outer nuclear layer; ON, optic nerve; OPL, outer plexiform layer; IPL, inner plexiform layer; PCL, pigment cell layer; WT, wild-type. The scale bar = 100 μm. E,F: Knockdown of Arl6ip causes the defective formation of the retinal axons in zebrafish embryos. By using the confocal fluorescence microscope, we observed that Zn8 protein was detected in the retinal ganglion cell axons of wild-type (WT) embryos at 48 hpf (E). However, the protein level of Zn8 presented a very weak signal near the retinal ventral (F, arrow) in the embryos injected with 4 ng/embryo of arl6ip-MO1 (MO). GCL, ganglion cell layer; ON, optic nerve. G-J: The retinal cells stayed at S-phase in retinal developmental period in zebrafish embryos when Arl6ip was lost. BrdU (red) and DAPI (blue) were used to label the S-phase of cell cycle and the nuclear, respectively. There was no BrdU signal detected in the outer nuclear layer (onl), inner nuclear layer (inl), and ganglion cell layer (gcl) (G,I) of wild-type embryos (WT) at 72 hpf, except the ciliary marginal zone (cmz, arrowhead). However, at the same stage, in embryos injected with 4 ng/embryo of arl6ip-MO (MO), we found that there was a large number of BrdU signal in retina (H,J), indicating that the retina cells of arl6ip-morphant were maintained at S-phase. The images were taken by confocal microscopy. The scale bars = 75 μm (G,I) and 50 μm (H,J). K: Detection of the transcripts of rhodopsin and red opsin mRNA of zebrafish embryos using RT-PCR. After the total RNA was extracted from wild-type embryos and arl6ip-morphants at 48, 72, and 96 hpf, RT-PCR was used to detect the existence of the transcripts of rhodopsin, which are expressed exclusively in rods and red cones, respectively. The expression of tubulin served as positive control. Results showed that neither rhodopsin nor red opsin transcripts were detected in either the wild-type embryos or arl6ip-morphants at 48 hpf as a result of the fact that the opsin gene is not transcribed until 50 hpf in the retina. After starting transcription of rhodopsin mRNA and red opsin mRNA at, for example, 72 and 96 hpf, the transcripts of rhodopsin and red opsin were present in the wild-type embryos. However, both rhodopsin and red opsin mRNA were totally absent in the arl6ip-morphants. WT, wild-type; MO, arl6ip-morphants.

Gene Expression Details
Gene Antibody Fish Conditions Stage Qualifier Anatomy Assay
opn1lw2 AB standard conditions Long-pec Not Detected whole organism RTPCR
Protruding-mouth to Day 4 whole organism RTPCR
AB + MO1-arl6ip1 standard conditions Long-pec Not Detected whole organism RTPCR
Protruding-mouth to Day 4 Not Detected whole organism RTPCR
rho AB standard conditions Long-pec Not Detected whole organism RTPCR
Protruding-mouth to Day 4 whole organism RTPCR
AB + MO1-arl6ip1 standard conditions Long-pec Not Detected whole organism RTPCR
Protruding-mouth to Day 4 Not Detected whole organism RTPCR
Antibody Labeling Details
Antibody Assay Fish Conditions Stage Qualifier Anatomy
Ab6-BrdU IHC AB chemical treatment: 5-bromo-2'-deoxyuridine Protruding-mouth ciliary marginal zone
IHC Protruding-mouth Not Detected retinal ganglion cell layer
IHC Protruding-mouth Not Detected retinal inner nuclear layer
IHC Protruding-mouth Not Detected retinal outer nuclear layer
IHC AB + MO1-arl6ip1 chemical treatment: 5-bromo-2'-deoxyuridine Protruding-mouth retinal ganglion cell layer
IHC Protruding-mouth retinal inner nuclear layer
IHC Protruding-mouth retinal outer nuclear layer
zn-8 IHC AB standard conditions Long-pec retinal ganglion cell axon
IHC AB + MO1-arl6ip1 standard conditions Long-pec retinal ganglion cell axon
Phenotype Details
Fish Conditions Stage Phenotype
AB + MO1-arl6ip1 chemical treatment: 5-bromo-2'-deoxyuridine Protruding-mouth camera-type eye photoreceptor cell differentiation disrupted, abnormal
Protruding-mouth cell cycle disrupted, abnormal
Protruding-mouth retinal bipolar neuron differentiation disrupted, abnormal
AB + MO1-arl6ip1 standard conditions Long-pec retina axon malformed, abnormal
Long-pec retina morphogenesis in camera-type eye disrupted, abnormal
Protruding-mouth retina decreased size, abnormal
Protruding-mouth retina development in camera-type eye disrupted, abnormal
Protruding-mouth retina layer formation disrupted, abnormal
Protruding-mouth retinal bipolar neuron differentiation delayed, abnormal
Day 4 retina decreased size, abnormal
Day 4 retina development in camera-type eye disrupted, abnormal
Day 4 retina layer formation disrupted, abnormal
Day 4 retinal bipolar neuron differentiation delayed, abnormal
Acknowledgments:
ZFIN wishes to thank the journal Developmental dynamics : an official publication of the American Association of Anatomists for permission to reproduce figures from this article. Please note that this material may be protected by copyright. Full text @ Dev. Dyn.