Fig 1.
Three different processes in chicken embryo skin development based on morphogenesis.
Three different processes in chicken embryo skin development were analyzed: Micro-patterning (E6–E8), intra-bud morphogenesis (E9–E10) and follicle morphogenesis (After E11). Histological sections of three stages of chicken skin during embryonic development (first column: photograph; the second column: Magnified 10×; the third column: Magnified 20×).
Fig 2.
The gene expression pattern in skin during chicken embryo development.
(A) Hierarchical clustering analysis of differentially expressed genes. Upregulated genes are displayed as red, whereas downregulated genes are displayed as green. Abscissa: days, ordinate: genes. (B) Hierarchical clustering of 16 time-course transcriptomes performed by pvclust. Values at the branches are Approximately Unbiased(AU) p-values (left), Bootstrap Probability(BP) values (right), and cluster labels (bottom). Clusters with AU ≥ 95 are indicated by rectangles.
Table 1.
KEGG pathway analysis clustering of highly expressed for four subdivisions.
Fig 3.
During micro-patterning, the skin expressed genes related to the rearrangement of the cytoskeleton.
(A) Cluster day 6 displayed a skin development pattern in micro-patterning (E6–E8), showing a sharp decline in expression from E6. (B) Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showing the global gene expression patterns from Cluster day 6. (C) The expression of these differentially expressed genes (DEGs) encode core regulatory transcription factors for basal metabolism and amino acid metabolism, declined during the period of micro-patterning (E6–E8). (D) Cluster day 8 displayed skin development patterns that increased sharply during the period of micro-patterning (E6–E8) and peaked at E8. (E) KEGG analysis showing the global gene expression patterns from Cluster day 8. (F) Expression of members of the collagen family began at E6 but was downregulated from E7. (G) Expression of genes associated with the rearrangement of the cytoskeleton began at E6 and continued to increase over the course of time (E6–E8).
Fig 4.
Dynamics of Wnt signaling pathways in intra-bud morphogenesis (E9–E10) during the development of skin in chicken embryos.
(A) Expression levels of Wnt family members (WNT5A, WNT11, WNT10A) were upregulated at E10 compared with E6. (B) CTNNB1 expression was dramatically increased by 20-fold during E9–E10. (C) The expression levels of FZD10 and FZD6, encoding the receptors for Wnt proteins, were significantly upregulated and downregulated, by 2.4- and 2.3-fold, respectively, over the course of time. (D) The expression of LEF, which encodes a transcription factor involved in the Wnt signaling pathway, was increased by 44-fold at E10 compared with E6. (E) Several genes encoding negative regulators of the Wnt/beta-catenin canonical signaling pathway, such as DKK1, NKD1, TCF1, were consistently upregulated during intra-bud morphogenesis (E9–E10).
Fig 5.
Representative time-course profile clusters in feather follicle morphogenesis and the results of pathway analysis.
We clustered the profiles into coherent groups using weighted gene co-expression network analysis (WGCNA) and captured the two representative time-course profile clusters that included Cluster day12 (peaked at E12) (A), Cluster day14 (peaked at E14) (C), and performed Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis for the set of genes in the two clusters (B, D).
Fig 6.
The expression trends of two Wnt signaling pathways in feather follicle morphology.
(A) Schematic diagram of the expression of the canonical Wnt/β-catenin signaling pathway. (B) Schematic diagram of the expression of inhibitory factors of the canonical Wnt/β-catenin signaling pathway.
Table 2.
Gene expression changes in Wnt signaling pathway in feather follicle morphogenesis.
Fig 7.
qPCR validation of gene expression including that of CTNNB1 and NKD1 from day6 to day14 of embryonic development.
Fig 8.
The dynamic regulation of the two kinds of Wnt pathways.