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Notch and MAML-1 complexation do not detectably alter the DNA binding specificity of the transcription factor CSL.

Del Bianco C, Vedenko A, Choi SH, Berger MF, Shokri L, Bulyk ML, Blacklow SC - PLoS ONE (2010)

Bottom Line: Here, we utilized protein-binding microarrays (PBMs) to compare the binding site preferences of isolated CSL with the preferred binding sites of CSL when bound to the CSL-binding domains of all four different human Notch receptors.Our data show no detectable difference in the DNA binding site preferences of CSL before and after loading of Notch and MAML1 proteins.These findings support the conclusion that accrual of Notch and MAML1 promote transcriptional activation without dramatically altering the preferred sites of DNA binding, and illustrate the potential of PBMs to analyze the binding site preferences of multiprotein-DNA complexes.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts, USA.

ABSTRACT

Background: Canonical Notch signaling is initiated when ligand binding induces proteolytic release of the intracellular part of Notch (ICN) from the cell membrane. ICN then travels into the nucleus where it drives the assembly of a transcriptional activation complex containing the DNA-binding transcription factor CSL, ICN, and a specialized co-activator of the Mastermind family. A consensus DNA binding site motif for the CSL protein was previously defined using selection-based methods, but whether subsequent association of Notch and Mastermind-like proteins affects the DNA binding preferences of CSL has not previously been examined.

Principal findings: Here, we utilized protein-binding microarrays (PBMs) to compare the binding site preferences of isolated CSL with the preferred binding sites of CSL when bound to the CSL-binding domains of all four different human Notch receptors. Measurements were taken both in the absence and in the presence of Mastermind-like-1 (MAML1). Our data show no detectable difference in the DNA binding site preferences of CSL before and after loading of Notch and MAML1 proteins.

Conclusions/significance: These findings support the conclusion that accrual of Notch and MAML1 promote transcriptional activation without dramatically altering the preferred sites of DNA binding, and illustrate the potential of PBMs to analyze the binding site preferences of multiprotein-DNA complexes.

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Related in: MedlinePlus

Comparison of sequence preferences for GST-CSL alone compared with GST-CSL in complexes containing Notch1 and MAML-1.Scatter plots compare PBM enrichment scores of individual 8-mers for: A) GST-CSL versus GST-CSL/Notch1 complexes, B) GST-CSL versus GST-CSL/Notch1/MAML-1 complexes. Enrichment scores were determined from the experiments shown in Figure 2.
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pone-0015034-g003: Comparison of sequence preferences for GST-CSL alone compared with GST-CSL in complexes containing Notch1 and MAML-1.Scatter plots compare PBM enrichment scores of individual 8-mers for: A) GST-CSL versus GST-CSL/Notch1 complexes, B) GST-CSL versus GST-CSL/Notch1/MAML-1 complexes. Enrichment scores were determined from the experiments shown in Figure 2.

Mentions: A more comprehensive statistical analysis of the E-scores of all 32,896 ungapped 8-mers (reverse complements are merged), comparing different pairs of conditions (CSL versus Notch1-CSL, and CSL versus MAML-1-Notch1-CSL), shows that the E-scores of the different 8-mers correlate very tightly among the three conditions, providing additional support for the conclusion that the distribution of bound sites is not altered upon loading of Notch1 and MAML-1 (Figure 3).


Notch and MAML-1 complexation do not detectably alter the DNA binding specificity of the transcription factor CSL.

Del Bianco C, Vedenko A, Choi SH, Berger MF, Shokri L, Bulyk ML, Blacklow SC - PLoS ONE (2010)

Comparison of sequence preferences for GST-CSL alone compared with GST-CSL in complexes containing Notch1 and MAML-1.Scatter plots compare PBM enrichment scores of individual 8-mers for: A) GST-CSL versus GST-CSL/Notch1 complexes, B) GST-CSL versus GST-CSL/Notch1/MAML-1 complexes. Enrichment scores were determined from the experiments shown in Figure 2.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2991368&req=5

pone-0015034-g003: Comparison of sequence preferences for GST-CSL alone compared with GST-CSL in complexes containing Notch1 and MAML-1.Scatter plots compare PBM enrichment scores of individual 8-mers for: A) GST-CSL versus GST-CSL/Notch1 complexes, B) GST-CSL versus GST-CSL/Notch1/MAML-1 complexes. Enrichment scores were determined from the experiments shown in Figure 2.
Mentions: A more comprehensive statistical analysis of the E-scores of all 32,896 ungapped 8-mers (reverse complements are merged), comparing different pairs of conditions (CSL versus Notch1-CSL, and CSL versus MAML-1-Notch1-CSL), shows that the E-scores of the different 8-mers correlate very tightly among the three conditions, providing additional support for the conclusion that the distribution of bound sites is not altered upon loading of Notch1 and MAML-1 (Figure 3).

Bottom Line: Here, we utilized protein-binding microarrays (PBMs) to compare the binding site preferences of isolated CSL with the preferred binding sites of CSL when bound to the CSL-binding domains of all four different human Notch receptors.Our data show no detectable difference in the DNA binding site preferences of CSL before and after loading of Notch and MAML1 proteins.These findings support the conclusion that accrual of Notch and MAML1 promote transcriptional activation without dramatically altering the preferred sites of DNA binding, and illustrate the potential of PBMs to analyze the binding site preferences of multiprotein-DNA complexes.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts, USA.

ABSTRACT

Background: Canonical Notch signaling is initiated when ligand binding induces proteolytic release of the intracellular part of Notch (ICN) from the cell membrane. ICN then travels into the nucleus where it drives the assembly of a transcriptional activation complex containing the DNA-binding transcription factor CSL, ICN, and a specialized co-activator of the Mastermind family. A consensus DNA binding site motif for the CSL protein was previously defined using selection-based methods, but whether subsequent association of Notch and Mastermind-like proteins affects the DNA binding preferences of CSL has not previously been examined.

Principal findings: Here, we utilized protein-binding microarrays (PBMs) to compare the binding site preferences of isolated CSL with the preferred binding sites of CSL when bound to the CSL-binding domains of all four different human Notch receptors. Measurements were taken both in the absence and in the presence of Mastermind-like-1 (MAML1). Our data show no detectable difference in the DNA binding site preferences of CSL before and after loading of Notch and MAML1 proteins.

Conclusions/significance: These findings support the conclusion that accrual of Notch and MAML1 promote transcriptional activation without dramatically altering the preferred sites of DNA binding, and illustrate the potential of PBMs to analyze the binding site preferences of multiprotein-DNA complexes.

Show MeSH
Related in: MedlinePlus