FIGURE

Fig. 1

ID
ZDB-FIG-231102-39
Publication
Binder et al., 2023 - Microenvironmental control of hematopoietic stem cell fate via CXCL8 and protein kinase C
Other Figures
All Figure Page
Back to All Figure Page
Fig. 1

Transcription of prkcda is repressed in the vascular niche and dysregulated expression of prkcda expands phenotypic HSPCs (A) Representative images of the kdrl:EGFP; lyve1b:DsRed transgenic line. Bar, 100 μm. (B) Volcano plot indicating genes with significantly differentially regulated genes in red; prkcda and selected CHT-specific genes are shown. Positive absolute log fold change (ALFC) indicates upregulation in CHT-derived sinusoidal endothelial cells compared to non-CHT-derived arteriovenous endothelial cells. (C) Differential expression of PKC-family genes in CHT-derived and non-CHT-derived endothelial cells. Values represent average fragments per kilobase of exon per million mapped fragments (FPKM) from three biological replicates. For prkcda, p = 0.00035. (D–F) Whole-mount in situ hybridization for prkcda in 72-hpf zebrafish embryos. (E and F) Flow cytometric analysis of kdrl:EGFP;sele:prkcda-2A-mCherry (E) and Runx1:EGFP;sele:prkcda-2A-mCherry (F) embryos. (G and H) Representative images of kdrl:EGFP zebrafish embryos injected (G) or not (H) with the sele:prkcda-2A-mCherry expression construct. Bar, 100 μm. (I) Dysregulated expression of prkcda in Runx1:EGFP;sele:prkcda-2A-mCherry embryos expands HSPCs. Each PKC family member is compared to clutchmates injected with a vector containing an empty multiple cloning site (control). Data are presented as fold change relative to control for each clutch (two or three clutches analyzed per PKC family member; each point represents a biological replicate). Boxes represent mean ± SEM. Prkcda vs. control: 1.94 ± 0.3-fold increase, p = 0.0092, Welch’s t test. (J) Representative images of CHT colonization in control and prkcda groups. Dotted lines indicate the CHT. Bar, 100 μm. (K and L) Representative images of kdrl:EGFP(+) sele:prkcda-2A-mCherry(+) (K) and kdrl:EGFP(+); sele:prkcda-2A-mCherry(−) (L) transgenics. Bar, 100 μm. (M) Phenotypic HSPCs were quantified in stable Runx1:EGFP;sele:prkcda-2A-mCherry transgenics (+) and Runx1:EGFP clutchmates (−) at 72 hpf. Data are pooled from two independent clutches and are represented as fold change relative to (−). Boxes represent mean ± SEM; each point represents a biological replicate. + vs. −: 1.19 ± 0.08-fold increase, p = 0.014, Wilcoxon rank-sum test. (N) Representative images of CHT colonization for the (−) and (+) animals from Figure 1G. Dotted lines indicate CHT. (O) Phenotypic HSPCs were quantified in Runx1:EGFP transgenics treated with HA-100 or an equivalent volume of DMSO as a vehicle control. Data are pooled from two independent experiments and are shown as fold change relative to control. Box represents mean ± SEM; each point represents a biological replicate. HA-100 vs. control: 0.38 ± 0.06-fold decrease, p = 8.3 × 10−6, Wilcoxon rank-sum test. See also Figure S1 and Table S1.

Expression Data

Expression Detail
Antibody Labeling
Phenotype Data

Phenotype Detail
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 @ Cell Rep.