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ZFIN ID: ZDB-FIG-160824-20
Huang et al., 2016 - The circadian clock regulates autophagy directly through the nuclear hormone receptor Nr1d1/Rev-erbα and indirectly via Cebpb/(C/ebpβ) in zebrafish. Autophagy   12(8):1292-309 Full text @ Autophagy
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Genes:
Fish:
Condition:
Anatomical Term:
Stage Range: Day 4 to Day 5
PHENOTYPE:
Fish:
Condition:
Observed In:
Stage Range: Long-pec to Adult

Fig. 4

Disrupted autophagy activities and altered expression of autophagy genes in nr1d1 mutant zebrafish. (A) Zebrafish larval autolysosomes were estimated by expression of mCherry-Map1lc3b at 56 hpf. The scale bar in the lower left of the wild-type panel represents 100 microns. The right upper panels show high magnification highlighting a hindbrain region in both wild-type and nr1d1 mutant larva labeled with mCherry-Map1lc3b. (B) Transmission electron micrograph of wild-type and nr1d1 mutant liver sections at ZT 0. The scale bar in the lower left of the wild-type panel represents one micron. (C) Quantification of autolysosomes in (A), calculated manually. (D) Quantification of autophagosome abundance in (B) with software ImageJ. The figure represents one of the 3 independent sets of samples. Data represent mean ± s.d.(**, Pd0 .01). The Student t test was conducted, (**, P ≤ 0 .01). (E) RT-PCR analysis of autophagy genes in nr1d1 (7bp or +124,-2bp) mutant zebrafish at different time points under DD condition. Zebrafish larvae were raised under light/dark (14 h/10 h) condition for the first 3 dpf, and then placed under constant darkness condition. Approximately 50 zebrafish larvae were pooled for each time point. The mRNA expression levels were analyzed by the JTK-CYCLE method. ADJ.P for adjusted minimal p-values (*, P ≤ 0 .05; ***, P ≤ 0 .001), AMP for amplitude. Two-way ANOVA with the Tukey post hoc test was conducted (*, P ≤ 0 .05; **, P ≤ 0 .01).

Gene Expression Details
Gene Antibody Fish Conditions Stage Anatomy Assay
atg4a nr1d1sus010/sus010 constant dark Day 4 to Day 5 whole organism RTPCR
nr1d1sus012/sus012 constant dark Day 4 whole organism RTPCR
Day 5 whole organism RTPCR
atg4da nr1d1sus010/sus010 constant dark Day 4 to Day 5 whole organism RTPCR
atp6v1d nr1d1sus010/sus010 constant dark Day 4 to Day 5 whole organism RTPCR
becn1 nr1d1sus010/sus010 constant dark Day 4 to Day 5 whole organism RTPCR
bnip3 nr1d1sus010/sus010 constant dark Day 4 to Day 5 whole organism RTPCR
gabarapa nr1d1sus010/sus010 constant dark Day 4 to Day 5 whole organism RTPCR
map1lc3b nr1d1sus010/sus010 constant dark Day 4 to Day 5 whole organism RTPCR
nr1d1sus012/sus012 constant dark Day 4 whole organism RTPCR
Day 5 whole organism RTPCR
ulk1a nr1d1sus010/sus010 constant dark Day 4 to Day 5 whole organism RTPCR
nr1d1sus012/sus012 constant dark Day 4 whole organism RTPCR
Day 5 whole organism RTPCR
Antibody Labeling Details No data available
Phenotype Details
Fish Conditions Stage Phenotype
nr1d1sus010/sus010 control Long-pec hindbrain autolysosome increased amount, abnormal
Adult liver autophagosome increased amount, abnormal
nr1d1sus010/sus010 constant dark Day 4 - Day 5 whole organism becn1 expression increased amount, abnormal
Day 4 - Day 5 whole organism map1lc3b expression increased amount, abnormal
Day 4 - Day 5 whole organism atg4a expression increased amount, abnormal
Day 4 - Day 5 whole organism atg4da expression increased amount, abnormal
Day 4 - Day 5 whole organism ulk1a expression increased amount, abnormal
Day 4 - Day 5 whole organism atp6v1d expression increased amount, abnormal
Day 4 - Day 5 whole organism gabarapa expression increased amount, abnormal
Day 4 - Day 5 whole organism bnip3 expression increased amount, abnormal
nr1d1sus012/sus012 constant dark Day 4 whole organism atg4a expression increased amount, abnormal
Day 4 whole organism ulk1a expression increased amount, abnormal
Day 4 whole organism map1lc3b expression increased amount, abnormal
Day 5 whole organism map1lc3b expression increased amount, abnormal
Day 5 whole organism ulk1a expression increased amount, abnormal
Day 5 whole organism atg4a expression increased amount, abnormal
Acknowledgments:
ZFIN wishes to thank the journal Autophagy for permission to reproduce figures from this article. Please note that this material may be protected by copyright. Full text @ Autophagy