Journal: PLoS Biology

Article Title: Cryptic Patterning of Avian Skin Confers a Developmental Facility for Loss of Neck Feathering

doi: 10.1371/journal.pbio.1001028

Figure Lengend Snippet: (A) Adult Na / Na . Feathers are absent on the neck and head, excepting the crown. (B) E8.5 embryos hybridized with a β - catenin probe to mark the patterning field and feather primordia. Punctate expression of β - catenin in feather placodes is seen on the body but not the neck of the mutant. WT, wild type; Na/Na , Naked neck. (C) E12.5 embryos showing limited lateral tract expansion (arrows) in Na / Na , reducing body feather coverage. (D) Quantitative RT-PCR determination of BMP12 expression in body and neck skin of E7.5 and E8.5 wild type and Na / Na embryos. (E,F) In situ hybridization detecting BMP12 in wild type and Na/Na embryos at (E) E7.5 and (F) E8.5. Wild type and mutant embryos were hybridized and photographed together. Na/Na embryos have elevated and diffuse expression of BMP12 in the skin. (G) Sequence traces of PCR products from E8.5 Na/+ . Genomic DNA PCR products display double peaks following a TA indel polymorphism in the BMP12 3′UTR. RT-PCR products from neck and body skin show a single trace throughout, indicating predominant expression of the Naked neck BMP12 allele, while both alleles are detected in RT-PCR products from other tissues. (H) Schematic showing insertion of chromosome 1 sequences into chromosome 3 at the Naked neck locus. Chromosome coordinates, the Naked neck identical by descent segment, gene names, exons, untranslated regions, and non-coding elements conserved between chicken and human genomes, based on the ENSEMBL genome viewer, are indicated.

Article Snippet: Recombinant human BMP4 and mouse BMP12 (R&D Systems) were used.

Techniques: Expressing, Mutagenesis, Quantitative RT-PCR, In Situ Hybridization, Sequencing, Reverse Transcription Polymerase Chain Reaction

Journal: PLoS Biology

Article Title: Cryptic Patterning of Avian Skin Confers a Developmental Facility for Loss of Neck Feathering

doi: 10.1371/journal.pbio.1001028

Figure Lengend Snippet: (A) Application of recombinant BMP12 to cultured skin for 15 h leads to elevation of SOSTDC1 expression, determined by quantitative RT-PCR. (B–E) Detection of SOSTDC1 expression by in situ hybridization. (B) At E7.5 wild type embryos have two rows of feather placodes running up the neck. SOSTDC1 is expressed at the periphery of the placodes and is not detected in the medial region between the lateral rows of placodes. (C) By E8.5 the medial region of the neck is populated by feather placodes. (D) E7.5 Na/Na embryos have placodes on the dorsum, but widespread SOSTDC1 expression on the neck, including the medial region. (E) At E8.5 the Naked neck skin maintains a high level of widespread SOSTDC1 expression, with peri-placode expression visible on the body. (F) Ex vivo rescue of the Naked neck phenotype by suppression of BMP signaling. E7.0 Na/Na skin was cultured in the presence of dorsomorphin (DM, used at 8 µM) and SB203580 (SB, used at 5 µM), pharmacological inhibitors of BMP signal transduction, for 48 h. This permitted feather development across most of the mutant neck skin.

Article Snippet: Recombinant human BMP4 and mouse BMP12 (R&D Systems) were used.

Techniques: Recombinant, Cell Culture, Expressing, Quantitative RT-PCR, In Situ Hybridization, Ex Vivo, Transduction, Mutagenesis

Journal: PLoS Biology

Article Title: Cryptic Patterning of Avian Skin Confers a Developmental Facility for Loss of Neck Feathering

doi: 10.1371/journal.pbio.1001028

Figure Lengend Snippet: (A,B) β - catenin in situ hybridization revealing the effects of recombinant BMP12 application on feather periodicity and regional distribution in wild type skin after 48 h. (C,D) Dose effects of BMP12 on the number of feather placode rows on the spinal tract of the body. Feather primordia are visualized by β - catenin in situ hybridization. (E) SOSTDC1 expression on control and 80 ng/ml BMP12 treated skin explants. Feather placodes express SOSTDC1 at their periphery on both body and neck. Upon application of BMP12, the non-placode skin of the neck expresses a higher level of SOSTDC1 than does the body (compare signal intensity in the red boxed area to that of the blue boxed area). (F) Schematic of reaction-diffusion regulatory interactions. Adjacent numbering refers to mathematical terms in the supporting methods. C I represents the constitutive, ubiquitous production of the Inhibitor. (G) Quantification of periodicity of Activator foci in simulated neck and body with differential sensitivities to Inhibitor. C I increases along the x -axis. (H) Pattern outcomes from reaction-diffusion dynamics in a field with graded sensitivity to the Inhibitor. Abolition of Activator foci in the more sensitive part of the field is achieved with little effect on periodic spacing in the remainder of the field, producing a macropatttern that matches the effects of BMP12 treatment on cultured skin. Colors denote local Activator concentrations, with black representing the highest and white the lowest Activator levels. Areas with high Activator concentration represent placodes.

Article Snippet: Recombinant human BMP4 and mouse BMP12 (R&D Systems) were used.

Techniques: In Situ Hybridization, Recombinant, Expressing, Control, Diffusion-based Assay, Cell Culture, Concentration Assay

Journal: PLoS Biology

Article Title: Cryptic Patterning of Avian Skin Confers a Developmental Facility for Loss of Neck Feathering

doi: 10.1371/journal.pbio.1001028

Figure Lengend Snippet: (A) RA administration reduces the density of placodes, which are detected by β - catenin in situ hybridization, completely inhibiting placode formation at high doses. Suppression of BMP signaling with 4 µM dorsomorphin and 5 µM SB203580 rescues placode formation in the presence of RA. (B) Quantification of placode density on neck and body upon RA treatment. With increasing doses of RA the feather density on body and neck converges and ultimately all feather placode formation is suppressed. (C) RA sensitizes body skin to BMP-driven inhibition of feather development. The application of 0.1 µM RA has little effect on the placode pattern and application of 40 ng/ml BMP12 permits placode formation on the body. Co-treatment with RA and BMP12 has a synergistic effect, completely suppressing feather development on the body. Conversely, treatment of skin with the RA synthesis inhibitor Citral renders the neck resistant to suppression of placode formation by BMPs.

Article Snippet: Recombinant human BMP4 and mouse BMP12 (R&D Systems) were used.

Techniques: In Situ Hybridization, Inhibition