ZFIN ID: ZDB-FIG-150629-1
Korolnek et al., 2014 - Control of Metazoan Heme Homeostasis by a Conserved Multidrug Resistance Protein. Cell Metabolism   19(6):1008-19 Full text @ Cell Metab.
EXPRESSION / LABELING:
Genes:
Fish:
Knockdown Reagent:
Anatomical Terms:
Stage Range: Prim-5 to Day 4
PHENOTYPE:
Fish:
Knockdown Reagent:
Observed In:
Stage Range: Prim-5 to Protruding-mouth

Fig. 4

mrp5 Is Required for Zebrafish Erythropoiesis

(A) Phylogenetic analysis of MRP5/ABCC5 clade in C. elegans (orange), zebrafish (red), mice (green), and humans (blue). Sequences were aligned using ClustalW, and a phylogenetic tree was generated using the Neighbor-Joining method in MEGA5. The tree is drawn to scale, with branch lengths in the same units as those of the evolutionary distances used to infer the phylogenetic tree. The evolutionary distances are in the units of the number of amino acid substitutions per site.

(B) Lateral view of zebrafish mrp5 expression by whole-mount in situ hybridization using antisense probe, 4 days postfertilization. Anterior is to the left. Sense probe image is shown to indicate background staining. B, brain, N, neural tube.

(C) Knockdown of zebrafish mrp5 using morpholinos (MOmrp5) results in severe anemia, as indicated by reduced staining of o-dianisidine-positive red cells, as indicated by black arrowheads.

(D) Knockdown of zebrafish mrp-5 using MOmrp5 results in reduced red cell formation. Transgenic embryos expressing GFP from the globin locus control region (LCR-GFP) were injected with control MO or MOmrp5. Left: on day 2 postfertilization, percent GFP-positive RBCs were analyzed by FACS. x and y axes measure GFP and forward scatter, respectively; boxed area indicates gate for RBCs. Right: quantification of morphants shown at left; error bars represent mean ± SEM. For MOmrp5 injection, n = 4. p < 0.01 for MOmrp5 morphants compared to control morphants under identical conditions (one-way ANOVA, Bonferroni post test).

(E) Lateral view of zebrafish gata1 expression in wild-type, control MO, and MOmrp5 morphants by whole-mount in situ hybridization using antisense probe 24 hpf. Anterior is to the left.

(F) Quantification of anemia rescue in zebrafish coinjected with mrp5 cRNA. p < 0.001 for MOmrp5 morphants compared to control morphants under identical conditions; n = 4. p < 0.01 for mrp5 morphants coinjected with rescue cRNA when compared to mrp5 morphants with no rescue cRNA under identical conditions; n = 3 (one-way ANOVA, Bonferroni post test).

See also Figure S4 and Table S1.

Gene Expression Details
Gene Antibody Fish Conditions Stage Qualifier Anatomy Assay
abcc5 WT standard conditions Day 4 central nervous system ISH
gata1a WT control Prim-5 intermediate cell mass of mesoderm ISH
WT standard conditions Prim-5 intermediate cell mass of mesoderm ISH
WT + MO1-abcc5 standard conditions Prim-5 Not Detected intermediate cell mass of mesoderm ISH
Antibody Labeling Details No data available
Phenotype Details
Fish Conditions Stage Phenotype
WT + MO1-abcc5 standard conditions Prim-5 erythrocyte differentiation decreased occurrence, abnormal
Protruding-mouth erythrocyte differentiation decreased occurrence, abnormal
Protruding-mouth hematopoietic system has fewer parts of type nucleate erythrocyte, abnormal
cz3325Tg + MO1-abcc5 standard conditions Long-pec erythrocyte differentiation decreased occurrence, abnormal
Long-pec hematopoietic system has fewer parts of type nucleate erythrocyte, abnormal
Acknowledgments:
ZFIN wishes to thank the journal Cell Metabolism for permission to reproduce figures from this article. Please note that this material may be protected by copyright.

Reprinted from Cell Metabolism, 19(6), Korolnek, T., Zhang, J., Beardsley, S., Scheffer, G.L., Hamza, I., Control of Metazoan Heme Homeostasis by a Conserved Multidrug Resistance Protein, 1008-19, Copyright (2014) with permission from Elsevier. Full text @ Cell Metab.