header logo image header logo text
Downloads Login
Research
General Information
ZIRC
ZFIN ID: ZDB-FIG-160308-6
Shen et al., 2016 - The Rag-Ragulator Complex Regulates Lysosome Function and Phagocytic Flux in Microglia. Cell Reports   14(3):547-59 Full text @ Cell Rep.
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Genes:
Antibodies:
Fish:
Anatomical Terms:
Stage Range: Day 4 to Day 6
PHENOTYPE:
Fish:
Observed In:
Stage Range: Day 4 to Day 6

Fig. 5

Accumulation of Undigested Neuronal Material in Microglia of In rragast77/st77 Mutants

(A–D, G, and H) Confocal images show living larvae bearing transgenes that label neurons Tg(nbt:DsRed) and microglia Tg(mpeg1:EGFP).

(A–E) In WT (A and C) and rragast77/st77 mutant (B and D) larvae, arrows indicate microglia that contain neuronal material. In WT larvae, more microglia contain neuronal material at 4 dpf (A and E) than at 6 dpf (C, C′, and E), whereas most microglia in mutants contained neuronal material at both 4 and 6 dpf (B, D, D′, and E). Each point in (E) represents the data from one larva.

(F and G) Frames from time-lapse movies of WT (F) and rragast77/st77 (G) mutant larvae starting at 4 dpf, with time points indicated in minutes.

(H) Quantification of the number of neuronal puncta inside microglia in time-lapse movies of WT and rragast77/st77 mutant larvae. Each line represents a single microglia (WT, n = 4 cells from three larvae; mutant, n = 4 cells from three larvae).

(I) Immunoblot of whole-animal protein lysates at 5 dpf shows normal LC3-I to LC3-II conversion, but an accumulation of p62 in rragast77/st77 mutants. Actin is shown as a loading control.

All larvae shown in (A)–(H) were genotyped by PCR after photography. Larvae analyzed in the immunoblot (I) were genotyped as described in the Experimental Procedures.

Gene Expression Details
Gene Antibody Fish Conditions Stage Anatomy Assay
DsRed gl22Tg ; zf148Tg standard conditions Day 4 brain neuron IFL
Day 6 brain neuron IFL
rragast77/st77 ; zf148Tg ; gl22Tg standard conditions Day 4 brain neuron IFL
Day 6 brain neuron IFL
EGFP gl22Tg ; zf148Tg standard conditions Day 4 brain microglial cell IFL
Day 6 brain microglial cell IFL
rragast77/st77 ; zf148Tg ; gl22Tg standard conditions Day 4 brain microglial cell IFL
Day 6 brain microglial cell IFL
Antibody Labeling Details
Antibody Assay Fish Conditions Stage Qualifier Anatomy
Ab2-map1lc3b WB WT standard conditions Day 5 whole organism
WB rragast77/st77 standard conditions Day 5 whole organism
Ab6-sqstm1 WB WT standard conditions Day 5 Not Detected whole organism
WB rragast77/st77 standard conditions Day 5 whole organism
Phenotype Details
Fish Conditions Stage Phenotype
rragast77/st77 standard conditions Day 5 microglial cell lysosomal protein catabolic process decreased occurrence, abnormal
Day 5 whole organism Ab6-sqstm1 labeling increased amount, abnormal
rragast77/st77 ; zf148Tg ; gl22Tg standard conditions Day 4 brain engulfment of apoptotic cell process quality, normal
Day 6 brain apoptotic cell clearance decreased occurrence, abnormal
Acknowledgments:
ZFIN wishes to thank the journal Cell Reports for permission to reproduce figures from this article. Please note that this material may be protected by copyright. Full text @ Cell Rep.