barx1 rabbit polyclonal  (Bioss)

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Developmental stalling and organ-autonomous regulation of morphogenesis

doi: 10.1073/pnas.1112801108

Figure Lengend Snippet: Temporal delay of molar tooth development in Barx1 homozygous mutants. Hematoxylin and eosin stained frontal (A–J) and sagittal (K and L) sections of lower E13.5 incisors (A and B) and first molars (C and D), E14.5 incisors (E and F), and first molars (G and H), E16.5 first molars (I and J), and postnatal day 0 (P0) first molars (K and L). At E13.5, all tooth germs have reached a bud stage both in the Barx1 homozygous mutant (B and D) and control littermate (A and C). Incisors develop normally in all Barx1 homozygous mutants, displaying a characteristic epithelial cap at E14.5 (E and F), whereas the molars of all four quadrants show a developmental delay between E13.5 and E14.5, exhibiting a bud shape instead of a cap (G and H) (n > 10). Arrowheads in E–G indicate the primary enamel knot, visualized as a bulge on the inside of the epithelial cap. At E16.5–P0, Barx1−/− molars are slightly smaller but otherwise normal (I–L). m, Meckel's cartilage; vl, vestibular lamina. (Scale bar, 100 μm in A–J and 200 μm in K and L.)

Article Snippet: C3H10T1/2 cells were sequentially immunostained with anti-Barx1 (H-55, rabbit polyclonal antibody; Santa Cruz Biotechnology) and anti-Msx1 antibody (rabbit polyclonal antibody; Santa Cruz Biotechnology).

Techniques: Staining, Mutagenesis

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Developmental stalling and organ-autonomous regulation of morphogenesis

doi: 10.1073/pnas.1112801108

Figure Lengend Snippet: Barx1 mutant molar teeth exhibit changes in cell proliferation. (A–F) BrdU staining of frontal sections of developing first molar tooth germs at E13.5 (A and B), E14.5 (C and D), and E15.5 (E and F) in a Barx1+/− (A, C, and E) and Barx1−/− (B, D, and F) lower jaw. (G) Graphs comparing the numbers of BrdU-labeled cells in the epithelium and the condensed mesenchyme of developing lower first molars at E13.5, E14.5, and E15.5. Error bars show SD.

Article Snippet: C3H10T1/2 cells were sequentially immunostained with anti-Barx1 (H-55, rabbit polyclonal antibody; Santa Cruz Biotechnology) and anti-Msx1 antibody (rabbit polyclonal antibody; Santa Cruz Biotechnology).

Techniques: Mutagenesis, BrdU Staining, Labeling

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Developmental stalling and organ-autonomous regulation of morphogenesis

doi: 10.1073/pnas.1112801108

Figure Lengend Snippet: BMP4 expression and BMP activity changes in Barx1 mutant tooth development. (A–R) Expression of Bmp4 (A–F), Shh (G–L), and distribution of phospho-Smad1/5/8 (M–R) in lower first molar tooth germs at E13.5 (A, B, G, H, M, and N), E14.5 (C, D, I, J, O, and P), and E15.5 (E, F, K, L, Q, and R) in a WT (A, C, E, G, I, K, M, O, and Q) and Barx1 homozygous mutant (B, D, F, H, J, L, N, P, and R) littermates. (S–X) Expression of Shh in lower incisor tooth germs at E13.5 (S and T), E14.5 (U and V), and E16.5 (W and X). In situ hybridization was carried out on four separate samples for each genotype at each time point and immunostaining on two separate samples for each genotype at each time point. The epithelium of molar and incisor tooth germs is outlined in white.

Article Snippet: C3H10T1/2 cells were sequentially immunostained with anti-Barx1 (H-55, rabbit polyclonal antibody; Santa Cruz Biotechnology) and anti-Msx1 antibody (rabbit polyclonal antibody; Santa Cruz Biotechnology).

Techniques: Expressing, Activity Assay, Mutagenesis, In Situ Hybridization, Immunostaining

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Developmental stalling and organ-autonomous regulation of morphogenesis

doi: 10.1073/pnas.1112801108

Figure Lengend Snippet: Arrest of molar tooth development associated with a lack of Bmp4 transcription in Barx1/Msx1 compound mutants. Frontal sections through upper (A, C, E, G, I, K, M, O, Q, and S) and lower (B, D, F, H, J, L, N, P, R, and T) developing first molars at E14.5 (A, B, E, F, I, J, M, N, Q, and R) and E16.5 (C, D, G, H, K, L, O, P, S, and T). Hematoxylin and eosin stained sections of Barx1+/−; Msx1+/− (A–D), Barx1+/+; Msx1−/− (E–H), and Barx1−/−; Msx1+/− (I–L). (M–T) Expression of BMP4 in the condensed mesenchyme of first molar tooth germs of Barx1+/−; Msx1+/− (M–P) and Barx1−/−; Msx1+/− (Q–T). Permanent arrest of molar tooth development was observed in three separate Barx1−/−; Msx1+/− animals and was highly penetrant. At E16.5, one molar tooth germ (n = 1/12) was occasionally observed at the cap stage (corresponding to E14.5).

Article Snippet: C3H10T1/2 cells were sequentially immunostained with anti-Barx1 (H-55, rabbit polyclonal antibody; Santa Cruz Biotechnology) and anti-Msx1 antibody (rabbit polyclonal antibody; Santa Cruz Biotechnology).

Techniques: Staining, Expressing

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Developmental stalling and organ-autonomous regulation of morphogenesis

doi: 10.1073/pnas.1112801108

Figure Lengend Snippet: Msx1 interacts with molar tooth-specific transcription factor Barx1. (A) Msx1 interacts with Barx1 in living cells. C3H10T1/2 cells were cotransfected with pIRES2–Barx1–EGFP and either pCMV–FLAG–Msx1 or pCMV–FLAG–Tag2B empty control vector. Cell lysates were subjected to coimmunoprecipitations followed by Western blotting. Barx1 was detected only in the presence of FLAG–Msx1 in the IP sample. IP, immunoprecipitation; IB, immunoblotting. (B–I) Intracellular colocalization of Barx1 and Msx1 in C3H10T1/2 cells. (B–E) Intracellular colocalization of endogenously expressed Barx1 and Msx1. (B) Intracellular localization of Barx1 using anti-Barx1 (green); (C) intracellular localization of Msx1, using anti-Msx1 (red); (D) merged pictures showing intracellular colocalization of Barx1 and Msx1 (yellow); and (E) DNA staining using the fluorescence dye DRAQ5 (blue). (F–I) Intracellular colocalization of exogenously overexpressed Barx1 and Msx1 as EGFP and FLAG-tagged fusion proteins. (F) Intracellular localization of Barx1–EGFP (green); (G) intracellular localization of Msx1 using anti-Msx1 (red); (H) merged pictures show intracellular colocalization of Barx1–EGFP and FLAG–Msx1 (yellow); and (I) DNA staining using the fluorescence dye DRAQ5 (blue).

Article Snippet: C3H10T1/2 cells were sequentially immunostained with anti-Barx1 (H-55, rabbit polyclonal antibody; Santa Cruz Biotechnology) and anti-Msx1 antibody (rabbit polyclonal antibody; Santa Cruz Biotechnology).

Techniques: Plasmid Preparation, Western Blot, Immunoprecipitation, Staining, Fluorescence

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Developmental stalling and organ-autonomous regulation of morphogenesis

doi: 10.1073/pnas.1112801108

Figure Lengend Snippet: Barx1 is expressed in all multicusped teeth with expression levels correlating with cusp numbers. (A–C) Supernumerary teeth forming in the diastema of mice homozygous for Tg737orpk display Barx1 expression levels lower than first and second molars. (A and B) Consecutive sagittal sections through the upper jaw of an E18.5 mouse homozygous for Tg737orpk showing from Left to Right a second molar (M2), first molar (M1), and supernumerary tooth (SN), the latter developing mesial to M1 in the normally toothless diastema. (A) Trichrome staining showing the premolar-like shape of the ectopic diastema tooth. (B) Radioactive in situ hybridization for Barx1. (C) Quantification of Barx1 expression level in the dental mesenchyme of Tg737orpk/Tg737orpk second molar (M2), first molar (M1), and supernumerary tooth (SN). (D–I) Level of cpBarx1 expression correlates with cusp number in shrew multicusped teeth. (D and H) Dentition of an adult shrew upper (D) and lower (H) jaw composed of molars (M), premolars (PM), unicusps (U), and incisors (I). (E and F) Radioactive in situ hybridization for cpBarx1 in shrew premolars (E) and molars (F). Developing molars and premolars (outlined in red) have reached a cap stage. (G) Quantification of Barx1 expression levels in the dental mesenchyme of shrew premolar and molar tooth primordia. Gene expression was quantified by analyzing consecutive sections spanning the whole dental papilla of each tooth using ImageJ 1.34s. The number of cusps and crests displayed by each tooth is indicated above their respective Barx1 expression level. (I) 3D reconstructions of micro-CT scans of the upper and lower first molars (M1) and premolars (PM) of a 24-d-old shrew. Teeth are viewed from a lingual side; distal is Right and proximal Left. The number of cusps of each tooth (indicated in G) was carefully assessed by rotating the 3D models.

Article Snippet: C3H10T1/2 cells were sequentially immunostained with anti-Barx1 (H-55, rabbit polyclonal antibody; Santa Cruz Biotechnology) and anti-Msx1 antibody (rabbit polyclonal antibody; Santa Cruz Biotechnology).

Techniques: Expressing, Staining, In Situ Hybridization, Micro-CT

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Developmental stalling and organ-autonomous regulation of morphogenesis

doi: 10.1073/pnas.1112801108

Figure Lengend Snippet: Temporal delay of molar tooth development in Barx1 homozygous mutants. Hematoxylin and eosin stained frontal (A–J) and sagittal (K and L) sections of lower E13.5 incisors (A and B) and first molars (C and D), E14.5 incisors (E and F), and first molars (G and H), E16.5 first molars (I and J), and postnatal day 0 (P0) first molars (K and L). At E13.5, all tooth germs have reached a bud stage both in the Barx1 homozygous mutant (B and D) and control littermate (A and C). Incisors develop normally in all Barx1 homozygous mutants, displaying a characteristic epithelial cap at E14.5 (E and F), whereas the molars of all four quadrants show a developmental delay between E13.5 and E14.5, exhibiting a bud shape instead of a cap (G and H) (n > 10). Arrowheads in E–G indicate the primary enamel knot, visualized as a bulge on the inside of the epithelial cap. At E16.5–P0, Barx1−/− molars are slightly smaller but otherwise normal (I–L). m, Meckel's cartilage; vl, vestibular lamina. (Scale bar, 100 μm in A–J and 200 μm in K and L.)

Article Snippet: For Western blotting of eluted protein, primary antibodies were used with 1:500 dilution of rabbit anti-Barx1 polyclonal antibody (Santa Cruz) or 1:1,000 dilution mouse anti-FLAG M2 monoclonal antibody (Sigma).

Techniques: Staining, Mutagenesis, Control

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Developmental stalling and organ-autonomous regulation of morphogenesis

doi: 10.1073/pnas.1112801108

Figure Lengend Snippet: Barx1 mutant molar teeth exhibit changes in cell proliferation. (A–F) BrdU staining of frontal sections of developing first molar tooth germs at E13.5 (A and B), E14.5 (C and D), and E15.5 (E and F) in a Barx1+/− (A, C, and E) and Barx1−/− (B, D, and F) lower jaw. (G) Graphs comparing the numbers of BrdU-labeled cells in the epithelium and the condensed mesenchyme of developing lower first molars at E13.5, E14.5, and E15.5. Error bars show SD.

Article Snippet: For Western blotting of eluted protein, primary antibodies were used with 1:500 dilution of rabbit anti-Barx1 polyclonal antibody (Santa Cruz) or 1:1,000 dilution mouse anti-FLAG M2 monoclonal antibody (Sigma).

Techniques: Mutagenesis, BrdU Staining, Labeling

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Developmental stalling and organ-autonomous regulation of morphogenesis

doi: 10.1073/pnas.1112801108

Figure Lengend Snippet: BMP4 expression and BMP activity changes in Barx1 mutant tooth development. (A–R) Expression of Bmp4 (A–F), Shh (G–L), and distribution of phospho-Smad1/5/8 (M–R) in lower first molar tooth germs at E13.5 (A, B, G, H, M, and N), E14.5 (C, D, I, J, O, and P), and E15.5 (E, F, K, L, Q, and R) in a WT (A, C, E, G, I, K, M, O, and Q) and Barx1 homozygous mutant (B, D, F, H, J, L, N, P, and R) littermates. (S–X) Expression of Shh in lower incisor tooth germs at E13.5 (S and T), E14.5 (U and V), and E16.5 (W and X). In situ hybridization was carried out on four separate samples for each genotype at each time point and immunostaining on two separate samples for each genotype at each time point. The epithelium of molar and incisor tooth germs is outlined in white.

Article Snippet: For Western blotting of eluted protein, primary antibodies were used with 1:500 dilution of rabbit anti-Barx1 polyclonal antibody (Santa Cruz) or 1:1,000 dilution mouse anti-FLAG M2 monoclonal antibody (Sigma).

Techniques: Expressing, Activity Assay, Mutagenesis, In Situ Hybridization, Immunostaining

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Developmental stalling and organ-autonomous regulation of morphogenesis

doi: 10.1073/pnas.1112801108

Figure Lengend Snippet: Arrest of molar tooth development associated with a lack of Bmp4 transcription in Barx1/Msx1 compound mutants. Frontal sections through upper (A, C, E, G, I, K, M, O, Q, and S) and lower (B, D, F, H, J, L, N, P, R, and T) developing first molars at E14.5 (A, B, E, F, I, J, M, N, Q, and R) and E16.5 (C, D, G, H, K, L, O, P, S, and T). Hematoxylin and eosin stained sections of Barx1+/−; Msx1+/− (A–D), Barx1+/+; Msx1−/− (E–H), and Barx1−/−; Msx1+/− (I–L). (M–T) Expression of BMP4 in the condensed mesenchyme of first molar tooth germs of Barx1+/−; Msx1+/− (M–P) and Barx1−/−; Msx1+/− (Q–T). Permanent arrest of molar tooth development was observed in three separate Barx1−/−; Msx1+/− animals and was highly penetrant. At E16.5, one molar tooth germ (n = 1/12) was occasionally observed at the cap stage (corresponding to E14.5).

Article Snippet: For Western blotting of eluted protein, primary antibodies were used with 1:500 dilution of rabbit anti-Barx1 polyclonal antibody (Santa Cruz) or 1:1,000 dilution mouse anti-FLAG M2 monoclonal antibody (Sigma).

Techniques: Staining, Expressing

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Developmental stalling and organ-autonomous regulation of morphogenesis

doi: 10.1073/pnas.1112801108

Figure Lengend Snippet: Msx1 interacts with molar tooth-specific transcription factor Barx1. (A) Msx1 interacts with Barx1 in living cells. C3H10T1/2 cells were cotransfected with pIRES2–Barx1–EGFP and either pCMV–FLAG–Msx1 or pCMV–FLAG–Tag2B empty control vector. Cell lysates were subjected to coimmunoprecipitations followed by Western blotting. Barx1 was detected only in the presence of FLAG–Msx1 in the IP sample. IP, immunoprecipitation; IB, immunoblotting. (B–I) Intracellular colocalization of Barx1 and Msx1 in C3H10T1/2 cells. (B–E) Intracellular colocalization of endogenously expressed Barx1 and Msx1. (B) Intracellular localization of Barx1 using anti-Barx1 (green); (C) intracellular localization of Msx1, using anti-Msx1 (red); (D) merged pictures showing intracellular colocalization of Barx1 and Msx1 (yellow); and (E) DNA staining using the fluorescence dye DRAQ5 (blue). (F–I) Intracellular colocalization of exogenously overexpressed Barx1 and Msx1 as EGFP and FLAG-tagged fusion proteins. (F) Intracellular localization of Barx1–EGFP (green); (G) intracellular localization of Msx1 using anti-Msx1 (red); (H) merged pictures show intracellular colocalization of Barx1–EGFP and FLAG–Msx1 (yellow); and (I) DNA staining using the fluorescence dye DRAQ5 (blue).

Article Snippet: For Western blotting of eluted protein, primary antibodies were used with 1:500 dilution of rabbit anti-Barx1 polyclonal antibody (Santa Cruz) or 1:1,000 dilution mouse anti-FLAG M2 monoclonal antibody (Sigma).

Techniques: Control, Plasmid Preparation, Western Blot, Immunoprecipitation, Staining, Fluorescence

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Developmental stalling and organ-autonomous regulation of morphogenesis

doi: 10.1073/pnas.1112801108

Figure Lengend Snippet: Barx1 is expressed in all multicusped teeth with expression levels correlating with cusp numbers. (A–C) Supernumerary teeth forming in the diastema of mice homozygous for Tg737orpk display Barx1 expression levels lower than first and second molars. (A and B) Consecutive sagittal sections through the upper jaw of an E18.5 mouse homozygous for Tg737orpk showing from Left to Right a second molar (M2), first molar (M1), and supernumerary tooth (SN), the latter developing mesial to M1 in the normally toothless diastema. (A) Trichrome staining showing the premolar-like shape of the ectopic diastema tooth. (B) Radioactive in situ hybridization for Barx1. (C) Quantification of Barx1 expression level in the dental mesenchyme of Tg737orpk/Tg737orpk second molar (M2), first molar (M1), and supernumerary tooth (SN). (D–I) Level of cpBarx1 expression correlates with cusp number in shrew multicusped teeth. (D and H) Dentition of an adult shrew upper (D) and lower (H) jaw composed of molars (M), premolars (PM), unicusps (U), and incisors (I). (E and F) Radioactive in situ hybridization for cpBarx1 in shrew premolars (E) and molars (F). Developing molars and premolars (outlined in red) have reached a cap stage. (G) Quantification of Barx1 expression levels in the dental mesenchyme of shrew premolar and molar tooth primordia. Gene expression was quantified by analyzing consecutive sections spanning the whole dental papilla of each tooth using ImageJ 1.34s. The number of cusps and crests displayed by each tooth is indicated above their respective Barx1 expression level. (I) 3D reconstructions of micro-CT scans of the upper and lower first molars (M1) and premolars (PM) of a 24-d-old shrew. Teeth are viewed from a lingual side; distal is Right and proximal Left. The number of cusps of each tooth (indicated in G) was carefully assessed by rotating the 3D models.

Article Snippet: For Western blotting of eluted protein, primary antibodies were used with 1:500 dilution of rabbit anti-Barx1 polyclonal antibody (Santa Cruz) or 1:1,000 dilution mouse anti-FLAG M2 monoclonal antibody (Sigma).

Techniques: Expressing, Staining, In Situ Hybridization, Gene Expression, Micro-CT