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Crystal structure of the human FOXO3a-DBD/DNA complex suggests the effects of post-translational modification.

Tsai KL, Sun YJ, Huang CY, Yang JY, Hung MC, Hsiao CD - Nucleic Acids Res. (2007)

Bottom Line: Because these post-translational modification sites are located within the C-terminal basic region of the FOXO DNA-binding domain (FOXO-DBD), it is possible that these post-translational modifications could alter the DNA-binding characteristics.Based on a unique structural feature in the C-terminal region and results from biochemical and mutational studies, our studies may explain how FOXO-DBD C-terminal phosphorylation by protein kinase B (PKB) or acetylation by cAMP-response element binding protein (CBP) can attenuate the DNA-binding activity and thereby reduce transcriptional activity of FOXO proteins.In addition, we demonstrate that the methyl groups of specific thymine bases within the consensus sequence are important for FOXO3a-DBD recognition of the consensus binding site.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Biology, Academia Sinica, Taipei, Taiwan.

ABSTRACT
FOXO3a is a transcription factor of the FOXO family. The FOXO proteins participate in multiple signaling pathways, and their transcriptional activity is regulated by several post-translational mechanisms, including phosphorylation, acetylation and ubiquitination. Because these post-translational modification sites are located within the C-terminal basic region of the FOXO DNA-binding domain (FOXO-DBD), it is possible that these post-translational modifications could alter the DNA-binding characteristics. To understand how FOXO mediate transcriptional activity, we report here the 2.7 A crystal structure of the DNA-binding domain of FOXO3a (FOXO3a-DBD) bound to a 13-bp DNA duplex containing a FOXO consensus binding sequence (GTAAACA). Based on a unique structural feature in the C-terminal region and results from biochemical and mutational studies, our studies may explain how FOXO-DBD C-terminal phosphorylation by protein kinase B (PKB) or acetylation by cAMP-response element binding protein (CBP) can attenuate the DNA-binding activity and thereby reduce transcriptional activity of FOXO proteins. In addition, we demonstrate that the methyl groups of specific thymine bases within the consensus sequence are important for FOXO3a-DBD recognition of the consensus binding site.

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Detailed contacts of the FOXO3a-DBD/DNA interface. (A) Stereo view of the recognition of the GTAAACA consensus DNA-binding sequence by recognition helix H3 of FOXO3a-DBD. Hydrogen bonding and van der Waals contacts participating in base recognition are represented by dashed gray lines and dotted green lines, respectively. (B) Stereo view of major groove interactions of the C-terminal region of FOXO3a-DBD. Hydrogen bonds are represented by dashed gray lines. The C-terminal coil of FOXO3a-DBD is colored in green. (C) Interactions between DNA and the C-terminal positively charged Lys245 residue. (D) Interactions between wing 1 of FOXO3a-DBD and the minor groove of DNA are shown.
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Figure 3: Detailed contacts of the FOXO3a-DBD/DNA interface. (A) Stereo view of the recognition of the GTAAACA consensus DNA-binding sequence by recognition helix H3 of FOXO3a-DBD. Hydrogen bonding and van der Waals contacts participating in base recognition are represented by dashed gray lines and dotted green lines, respectively. (B) Stereo view of major groove interactions of the C-terminal region of FOXO3a-DBD. Hydrogen bonds are represented by dashed gray lines. The C-terminal coil of FOXO3a-DBD is colored in green. (C) Interactions between DNA and the C-terminal positively charged Lys245 residue. (D) Interactions between wing 1 of FOXO3a-DBD and the minor groove of DNA are shown.

Mentions: In most forkhead proteins, the recognition helices H3 within the DNA-binding domain share a highly conserved amino acid sequence (Figure 1B). As expected, the recognition helix H3 in the FOXO3a-DBD structure docked perpendicular to the major groove and formed extensive contacts with the FOXO consensus binding sequence. The recognition helix H3 centered over the FOXO consensus sequence, with several residues within helix 3 forming direct hydrogen bonds as well as van der Waals contacts with bases of the major groove. Details of the interactions between recognition helix H3 and DNA are shown in Figures 3A and 4. The conserved residues Asn208, Ser215, Arg211 and His212 within H3 were important for DNA recognition. Asn208 bound to the A4 base via two hydrogen bonds. Arg211 recognized T5′ and T7′ through van der Waals contacts and contributed to the specificity for G6′ with a direct hydrogen bond. The side chain of His212 protruded into the major groove and recognized bases T2 and T3′ through a van der Waals contact and a direct hydrogen bond, respectively. Ser215 recognized T4′ through a van der Waals contact. Among these interactions, we note that the methyl groups of thymine bases from the FOXO consensus binding sequence were crucial for FOXO3a-DBD promoter recognition. In addition to base recognition, Ser215 also interacted with the phosphate group of T8′ to further stabilize the complex.Figure 3.


Crystal structure of the human FOXO3a-DBD/DNA complex suggests the effects of post-translational modification.

Tsai KL, Sun YJ, Huang CY, Yang JY, Hung MC, Hsiao CD - Nucleic Acids Res. (2007)

Detailed contacts of the FOXO3a-DBD/DNA interface. (A) Stereo view of the recognition of the GTAAACA consensus DNA-binding sequence by recognition helix H3 of FOXO3a-DBD. Hydrogen bonding and van der Waals contacts participating in base recognition are represented by dashed gray lines and dotted green lines, respectively. (B) Stereo view of major groove interactions of the C-terminal region of FOXO3a-DBD. Hydrogen bonds are represented by dashed gray lines. The C-terminal coil of FOXO3a-DBD is colored in green. (C) Interactions between DNA and the C-terminal positively charged Lys245 residue. (D) Interactions between wing 1 of FOXO3a-DBD and the minor groove of DNA are shown.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2175300&req=5

Figure 3: Detailed contacts of the FOXO3a-DBD/DNA interface. (A) Stereo view of the recognition of the GTAAACA consensus DNA-binding sequence by recognition helix H3 of FOXO3a-DBD. Hydrogen bonding and van der Waals contacts participating in base recognition are represented by dashed gray lines and dotted green lines, respectively. (B) Stereo view of major groove interactions of the C-terminal region of FOXO3a-DBD. Hydrogen bonds are represented by dashed gray lines. The C-terminal coil of FOXO3a-DBD is colored in green. (C) Interactions between DNA and the C-terminal positively charged Lys245 residue. (D) Interactions between wing 1 of FOXO3a-DBD and the minor groove of DNA are shown.
Mentions: In most forkhead proteins, the recognition helices H3 within the DNA-binding domain share a highly conserved amino acid sequence (Figure 1B). As expected, the recognition helix H3 in the FOXO3a-DBD structure docked perpendicular to the major groove and formed extensive contacts with the FOXO consensus binding sequence. The recognition helix H3 centered over the FOXO consensus sequence, with several residues within helix 3 forming direct hydrogen bonds as well as van der Waals contacts with bases of the major groove. Details of the interactions between recognition helix H3 and DNA are shown in Figures 3A and 4. The conserved residues Asn208, Ser215, Arg211 and His212 within H3 were important for DNA recognition. Asn208 bound to the A4 base via two hydrogen bonds. Arg211 recognized T5′ and T7′ through van der Waals contacts and contributed to the specificity for G6′ with a direct hydrogen bond. The side chain of His212 protruded into the major groove and recognized bases T2 and T3′ through a van der Waals contact and a direct hydrogen bond, respectively. Ser215 recognized T4′ through a van der Waals contact. Among these interactions, we note that the methyl groups of thymine bases from the FOXO consensus binding sequence were crucial for FOXO3a-DBD promoter recognition. In addition to base recognition, Ser215 also interacted with the phosphate group of T8′ to further stabilize the complex.Figure 3.

Bottom Line: Because these post-translational modification sites are located within the C-terminal basic region of the FOXO DNA-binding domain (FOXO-DBD), it is possible that these post-translational modifications could alter the DNA-binding characteristics.Based on a unique structural feature in the C-terminal region and results from biochemical and mutational studies, our studies may explain how FOXO-DBD C-terminal phosphorylation by protein kinase B (PKB) or acetylation by cAMP-response element binding protein (CBP) can attenuate the DNA-binding activity and thereby reduce transcriptional activity of FOXO proteins.In addition, we demonstrate that the methyl groups of specific thymine bases within the consensus sequence are important for FOXO3a-DBD recognition of the consensus binding site.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Biology, Academia Sinica, Taipei, Taiwan.

ABSTRACT
FOXO3a is a transcription factor of the FOXO family. The FOXO proteins participate in multiple signaling pathways, and their transcriptional activity is regulated by several post-translational mechanisms, including phosphorylation, acetylation and ubiquitination. Because these post-translational modification sites are located within the C-terminal basic region of the FOXO DNA-binding domain (FOXO-DBD), it is possible that these post-translational modifications could alter the DNA-binding characteristics. To understand how FOXO mediate transcriptional activity, we report here the 2.7 A crystal structure of the DNA-binding domain of FOXO3a (FOXO3a-DBD) bound to a 13-bp DNA duplex containing a FOXO consensus binding sequence (GTAAACA). Based on a unique structural feature in the C-terminal region and results from biochemical and mutational studies, our studies may explain how FOXO-DBD C-terminal phosphorylation by protein kinase B (PKB) or acetylation by cAMP-response element binding protein (CBP) can attenuate the DNA-binding activity and thereby reduce transcriptional activity of FOXO proteins. In addition, we demonstrate that the methyl groups of specific thymine bases within the consensus sequence are important for FOXO3a-DBD recognition of the consensus binding site.

Show MeSH
Related in: MedlinePlus