Figure 1.
Promyelocytic leukemia (PML) regulates epidermal differentiation complex (EDC) genes during embryogenesis. (A) Heat map diagram of genes differentially affected in RNA-seq analysis of extracts from wild type (wt) and PML knockout mouse epidermis. Genome-wide differential expression at stage E13.5 (inner circle), E15.5, E16.5 (middle circles), and E17.5 (outer circle) is depicted. (B) Zoom-in of the region containing the EDC locus.
Figure 1.
Promyelocytic leukemia (PML) regulates epidermal differentiation complex (EDC) genes during embryogenesis. (A) Heat map diagram of genes differentially affected in RNA-seq analysis of extracts from wild type (wt) and PML knockout mouse epidermis. Genome-wide differential expression at stage E13.5 (inner circle), E15.5, E16.5 (middle circles), and E17.5 (outer circle) is depicted. (B) Zoom-in of the region containing the EDC locus.
Figure 2.
PML controls expression of the cornified envelope genes. (A) Graphs depicting differentially expressed cornified envelope genes. Left panel shows data from RNA-seq analysis. Right panel shows a comparison of data obtained from RNA-seq (stapled lines) and RT-qPCR (solid lines), respectively. (B) Box plots showing differential expression of selected genes from the cornified envelope region of the EDC gene locus by RT-qPCR analysis of PML-wt and PML-KO extracts. Differential expression at different time points during embryonic and postnatal development is shown. Each data point (dot) in box plot represents a single embryo. (C) Keratinocytes were isolated from the epidermis of PML-wt and PML-KO mouse embryos (stage E18.5) and subsequently passaged in culture. RNA was extracted from the cultured cells and selected genes from the cornified envelope region of the EDC gene locus were subjected to analysis by qPCR. Box plots show differential gene expression of selected genes. Each data point (dot) in box plot represents a single embryo.
Figure 2.
PML controls expression of the cornified envelope genes. (A) Graphs depicting differentially expressed cornified envelope genes. Left panel shows data from RNA-seq analysis. Right panel shows a comparison of data obtained from RNA-seq (stapled lines) and RT-qPCR (solid lines), respectively. (B) Box plots showing differential expression of selected genes from the cornified envelope region of the EDC gene locus by RT-qPCR analysis of PML-wt and PML-KO extracts. Differential expression at different time points during embryonic and postnatal development is shown. Each data point (dot) in box plot represents a single embryo. (C) Keratinocytes were isolated from the epidermis of PML-wt and PML-KO mouse embryos (stage E18.5) and subsequently passaged in culture. RNA was extracted from the cultured cells and selected genes from the cornified envelope region of the EDC gene locus were subjected to analysis by qPCR. Box plots show differential gene expression of selected genes. Each data point (dot) in box plot represents a single embryo.

Figure 3.
Elevated levels of PML bodies in basal epidermal cells during embryogenesis. (A) Schematic of epidermal cell layers during embryonic and postnatal development. Structural layers of epidermis at E13.5, E15.5, E17.5, and postnatal day (P) 0 is depicted. PML nuclear bodies are predominantly observed in the basal cell layer during embryogenesis. (B) Paraffin embedded sagittal sections obtained from developmental stage E13.5, E15.5, E17.5, and P0 were immunofluorescence (IF) labeled using antibodies against K15 (red) and PML NB (green). 4′,6-diamidino-2-phenylindole (DAPI) is shown in blue. The images are projections of multiple confocal Z-stacks. White rectangles highlight zoom-in of basal cells containing PML. (C) Quantitative assessment of PML bodies in different layers of epidermis obtained from E13.5, E15.5, E17.5, E18.5, and P0 mice. Each data point represents the average of PML bodies per cell from a single embryo.
Figure 3.
Elevated levels of PML bodies in basal epidermal cells during embryogenesis. (A) Schematic of epidermal cell layers during embryonic and postnatal development. Structural layers of epidermis at E13.5, E15.5, E17.5, and postnatal day (P) 0 is depicted. PML nuclear bodies are predominantly observed in the basal cell layer during embryogenesis. (B) Paraffin embedded sagittal sections obtained from developmental stage E13.5, E15.5, E17.5, and P0 were immunofluorescence (IF) labeled using antibodies against K15 (red) and PML NB (green). 4′,6-diamidino-2-phenylindole (DAPI) is shown in blue. The images are projections of multiple confocal Z-stacks. White rectangles highlight zoom-in of basal cells containing PML. (C) Quantitative assessment of PML bodies in different layers of epidermis obtained from E13.5, E15.5, E17.5, E18.5, and P0 mice. Each data point represents the average of PML bodies per cell from a single embryo.
Figure 4.
PML affects epidermal thickness during embryogenesis. (A) Representative sagittal images of epidermis derived from PML-wt and PML-KO E13.5, E15.5, E17.5, and P0 mice. Sections were IF-labeled using antibodies against K15 (red) and K10 (green). DAPI is shown in blue. The images represent projections of multiple confocal Z-stacks. (B) Quantification of epidermal thickness. The plotted lines represent average thickness (±STD) of K15 and K10-positive epidermal layers. Each data point represents measurements from a single microscopic field. More than 50 microscopic fields from 5–7 embryos/newborn mice per developmental stage were analyzed. (C) Quantification of cell density in basal and spinous layer of epidermis at different stages of embryonic development. Data are represented as aforementioned in (B).
Figure 4.
PML affects epidermal thickness during embryogenesis. (A) Representative sagittal images of epidermis derived from PML-wt and PML-KO E13.5, E15.5, E17.5, and P0 mice. Sections were IF-labeled using antibodies against K15 (red) and K10 (green). DAPI is shown in blue. The images represent projections of multiple confocal Z-stacks. (B) Quantification of epidermal thickness. The plotted lines represent average thickness (±STD) of K15 and K10-positive epidermal layers. Each data point represents measurements from a single microscopic field. More than 50 microscopic fields from 5–7 embryos/newborn mice per developmental stage were analyzed. (C) Quantification of cell density in basal and spinous layer of epidermis at different stages of embryonic development. Data are represented as aforementioned in (B).
Figure 5.
PML affects hair follicle size and orientation. (A) Representative sagittal images showing the morphology of hair follicles from PML-wt and PML-KO mice at stage E17.5 and P0. Samples have been IF labeled using antibodies against the basal keratinocyte marker K5 (red). (B) Schematic showing method for calculation of hair follicle size and orientation relative to the skin surface. (C) Polar diagram showing length and relative orientation of hair follicles from Pml+/+ and Pml−/− mice at the P17 and P0 developmental stages of mouse development.
Figure 5.
PML affects hair follicle size and orientation. (A) Representative sagittal images showing the morphology of hair follicles from PML-wt and PML-KO mice at stage E17.5 and P0. Samples have been IF labeled using antibodies against the basal keratinocyte marker K5 (red). (B) Schematic showing method for calculation of hair follicle size and orientation relative to the skin surface. (C) Polar diagram showing length and relative orientation of hair follicles from Pml+/+ and Pml−/− mice at the P17 and P0 developmental stages of mouse development.