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The transcription factor Sox5 modulates Sox10 function during melanocyte development.

Stolt CC, Lommes P, Hillgärtner S, Wegner M - Nucleic Acids Res. (2008)

Bottom Line: The transcription factor Sox5 has previously been shown in chicken to be expressed in early neural crest cells and neural crest-derived peripheral glia.This modulatory activity involved Sox5 binding and recruitment of CtBP2 and HDAC1 to the regulatory regions of melanocytic Sox10 target genes and direct inhibition of Sox10-dependent promoter activation.Both binding site competition and recruitment of corepressors thus help Sox5 to modulate the activity of Sox10 in the melanocyte lineage.

View Article: PubMed Central - PubMed

Affiliation: Institut für Biochemie, Emil-Fischer-Zentrum, Universität Erlangen, Fahrstrasse 17, D-91054 Erlangen, Germany.

ABSTRACT
The transcription factor Sox5 has previously been shown in chicken to be expressed in early neural crest cells and neural crest-derived peripheral glia. Here, we show in mouse that Sox5 expression also continues after neural crest specification in the melanocyte lineage. Despite its continued expression, Sox5 has little impact on melanocyte development on its own as generation of melanoblasts and melanocytes is unaltered in Sox5-deficient mice. Loss of Sox5, however, partially rescued the strongly reduced melanoblast generation and marker gene expression in Sox10 heterozygous mice arguing that Sox5 functions in the melanocyte lineage by modulating Sox10 activity. This modulatory activity involved Sox5 binding and recruitment of CtBP2 and HDAC1 to the regulatory regions of melanocytic Sox10 target genes and direct inhibition of Sox10-dependent promoter activation. Both binding site competition and recruitment of corepressors thus help Sox5 to modulate the activity of Sox10 in the melanocyte lineage.

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Sox5 counteracts the Sox10-dependent activation of melanocyte-specific promoters. Transient transfections were performed in Neuro2a (A, B, E, F, G and H) and B16 (C and D) cells using reporter plasmids in which the luciferase gene was under control of 1.1 kb of the Dct (A, C, E and G) or 1.5 kb of the Mitf (B, D, F and H) promoter. Expression plasmids for Sox10, the long (L-Sox5) and the short (S-Sox5) isoform of Sox5 were cotransfected as indicated below the bars in amounts of 100 ng each, with exception of Sox5 in panels (E) and (F) where plasmid amounts for L-Sox5 varied between 10, 20, 50, 100 and 200 ng. Transactivation rates for each promoter are presented as fold inductions ± SEM. Luciferase activities were determined in three experiments each performed in duplicates. The inlay in panel (H) shows both L-Sox5 and S-Sox5 detected on a western blot with an anti-Sox5 antiserum.
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Figure 5: Sox5 counteracts the Sox10-dependent activation of melanocyte-specific promoters. Transient transfections were performed in Neuro2a (A, B, E, F, G and H) and B16 (C and D) cells using reporter plasmids in which the luciferase gene was under control of 1.1 kb of the Dct (A, C, E and G) or 1.5 kb of the Mitf (B, D, F and H) promoter. Expression plasmids for Sox10, the long (L-Sox5) and the short (S-Sox5) isoform of Sox5 were cotransfected as indicated below the bars in amounts of 100 ng each, with exception of Sox5 in panels (E) and (F) where plasmid amounts for L-Sox5 varied between 10, 20, 50, 100 and 200 ng. Transactivation rates for each promoter are presented as fold inductions ± SEM. Luciferase activities were determined in three experiments each performed in duplicates. The inlay in panel (H) shows both L-Sox5 and S-Sox5 detected on a western blot with an anti-Sox5 antiserum.

Mentions: We next addressed the ability of both Sox proteins to influence the activities of the Mitf and Dct promoters in luciferase reporter assays. Whereas Sox10 robustly activated both promoters in transiently transfected Neuro2a cells, no such activation was observed for the long Sox5 isoform (L-Sox5) over a broad range of concentrations (Figure 5A and B and data not shown). This agrees with previous findings that all Sox5 isoforms lack a classical transactivation domain (3). When reporter genes were cotransfected simultaneously with both Sox proteins, Sox10 furthermore lost its ability to efficiently activate either the Dct promoter (Figure 5A) or the Mitf promoter (Figure 5B). We thus conclude that the long Sox5 isoform counteracts the transcriptional activity of Sox10 on its melanocytic target gene promoters in heterologous cell lines. Qualitatively similar results were also obtained in transiently transfected B16 melanoma cells (Figure 5C and D), although Sox10-dependent activation rates were lower, likely because of the presence of endogenous Sox10 in these cells (47) (see also Figure10A and B).Figure 5.


The transcription factor Sox5 modulates Sox10 function during melanocyte development.

Stolt CC, Lommes P, Hillgärtner S, Wegner M - Nucleic Acids Res. (2008)

Sox5 counteracts the Sox10-dependent activation of melanocyte-specific promoters. Transient transfections were performed in Neuro2a (A, B, E, F, G and H) and B16 (C and D) cells using reporter plasmids in which the luciferase gene was under control of 1.1 kb of the Dct (A, C, E and G) or 1.5 kb of the Mitf (B, D, F and H) promoter. Expression plasmids for Sox10, the long (L-Sox5) and the short (S-Sox5) isoform of Sox5 were cotransfected as indicated below the bars in amounts of 100 ng each, with exception of Sox5 in panels (E) and (F) where plasmid amounts for L-Sox5 varied between 10, 20, 50, 100 and 200 ng. Transactivation rates for each promoter are presented as fold inductions ± SEM. Luciferase activities were determined in three experiments each performed in duplicates. The inlay in panel (H) shows both L-Sox5 and S-Sox5 detected on a western blot with an anti-Sox5 antiserum.
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Figure 5: Sox5 counteracts the Sox10-dependent activation of melanocyte-specific promoters. Transient transfections were performed in Neuro2a (A, B, E, F, G and H) and B16 (C and D) cells using reporter plasmids in which the luciferase gene was under control of 1.1 kb of the Dct (A, C, E and G) or 1.5 kb of the Mitf (B, D, F and H) promoter. Expression plasmids for Sox10, the long (L-Sox5) and the short (S-Sox5) isoform of Sox5 were cotransfected as indicated below the bars in amounts of 100 ng each, with exception of Sox5 in panels (E) and (F) where plasmid amounts for L-Sox5 varied between 10, 20, 50, 100 and 200 ng. Transactivation rates for each promoter are presented as fold inductions ± SEM. Luciferase activities were determined in three experiments each performed in duplicates. The inlay in panel (H) shows both L-Sox5 and S-Sox5 detected on a western blot with an anti-Sox5 antiserum.
Mentions: We next addressed the ability of both Sox proteins to influence the activities of the Mitf and Dct promoters in luciferase reporter assays. Whereas Sox10 robustly activated both promoters in transiently transfected Neuro2a cells, no such activation was observed for the long Sox5 isoform (L-Sox5) over a broad range of concentrations (Figure 5A and B and data not shown). This agrees with previous findings that all Sox5 isoforms lack a classical transactivation domain (3). When reporter genes were cotransfected simultaneously with both Sox proteins, Sox10 furthermore lost its ability to efficiently activate either the Dct promoter (Figure 5A) or the Mitf promoter (Figure 5B). We thus conclude that the long Sox5 isoform counteracts the transcriptional activity of Sox10 on its melanocytic target gene promoters in heterologous cell lines. Qualitatively similar results were also obtained in transiently transfected B16 melanoma cells (Figure 5C and D), although Sox10-dependent activation rates were lower, likely because of the presence of endogenous Sox10 in these cells (47) (see also Figure10A and B).Figure 5.

Bottom Line: The transcription factor Sox5 has previously been shown in chicken to be expressed in early neural crest cells and neural crest-derived peripheral glia.This modulatory activity involved Sox5 binding and recruitment of CtBP2 and HDAC1 to the regulatory regions of melanocytic Sox10 target genes and direct inhibition of Sox10-dependent promoter activation.Both binding site competition and recruitment of corepressors thus help Sox5 to modulate the activity of Sox10 in the melanocyte lineage.

View Article: PubMed Central - PubMed

Affiliation: Institut für Biochemie, Emil-Fischer-Zentrum, Universität Erlangen, Fahrstrasse 17, D-91054 Erlangen, Germany.

ABSTRACT
The transcription factor Sox5 has previously been shown in chicken to be expressed in early neural crest cells and neural crest-derived peripheral glia. Here, we show in mouse that Sox5 expression also continues after neural crest specification in the melanocyte lineage. Despite its continued expression, Sox5 has little impact on melanocyte development on its own as generation of melanoblasts and melanocytes is unaltered in Sox5-deficient mice. Loss of Sox5, however, partially rescued the strongly reduced melanoblast generation and marker gene expression in Sox10 heterozygous mice arguing that Sox5 functions in the melanocyte lineage by modulating Sox10 activity. This modulatory activity involved Sox5 binding and recruitment of CtBP2 and HDAC1 to the regulatory regions of melanocytic Sox10 target genes and direct inhibition of Sox10-dependent promoter activation. Both binding site competition and recruitment of corepressors thus help Sox5 to modulate the activity of Sox10 in the melanocyte lineage.

Show MeSH
Related in: MedlinePlus