Transcriptional activity of the islet β cell factor Pdx1 Is augmented by lysine methylation catalyzed by the methyltransferase Set7/9

Aarthi V. Maganti, Bernhard Maier, Sarah A. Tersey, Megan L. Sampley, Amber Mosley, Sabire Özcan, Boobalan Pachaiyappan, Patrick M. Woster, Chad S. Hunter, Roland Stein, Raghu Mirmira

Research output: Contribution to journalArticle

18 Citations (Scopus)

Abstract

The transcription factor Pdx1 is crucial to islet β cell function and regulates target genes in part through interaction with coregulatory factors. Set7/9 is a Lys methyltransferase that interacts with Pdx1. Here we tested the hypothesis that Lys methylation of Pdx1 by Set7/9 augments Pdx1 transcriptional activity. Using mass spectrometry and mutational analysis of purified proteins, we found that Set7/9 methylates the N-terminal residues Lys-123 and Lys-131 of Pdx1. Methylation of these residues occurred only in the context of intact, full-length Pdx1, suggesting a specific requirement of secondary and/or tertiary structural elements for catalysis by Set7/9. Immunoprecipitation assays and mass spectrometric analysis using β cells verified Lys methylation of endogenous Pdx1. Cell-based luciferase reporter assays using wild-type and mutant transgenes revealed a requirement of Pdx1 residue Lys-131, but not Lys-123, for transcriptional augmentation by Set7/9. Lys-131 was not required for high-affinity interactions with DNA in vitro, suggesting that its methylation likely enhances post-DNA binding events. To define the role of Set7/9 in β cell function, we generated mutant mice in which the gene encoding Set7/9 was conditionally deleted in β cells (Set Δβ). SetΔβ mice exhibited glucose intolerance similar to Pdx1-deficient mice, and their isolated islets showed impaired glucose-stimulated insulin secretion with reductions in expression of Pdx1 target genes. Our results suggest a previously unappreciated role for Set7/9-mediated methylation in the maintenance of Pdx1 activity and β cell function.

Original languageEnglish
Pages (from-to)9812-9822
Number of pages11
JournalJournal of Biological Chemistry
Volume290
Issue number15
DOIs
StatePublished - Apr 10 2015

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Methylation
Methyltransferases
Islets of Langerhans
Lysine
Assays
Genes
Glucose
Gene encoding
Glucose Intolerance
DNA
Luciferases
Catalysis
Transgenes
Immunoprecipitation
Mass spectrometry
Mass Spectrometry
Transcription Factors
Maintenance
Insulin
Proteins

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

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Transcriptional activity of the islet β cell factor Pdx1 Is augmented by lysine methylation catalyzed by the methyltransferase Set7/9. / Maganti, Aarthi V.; Maier, Bernhard; Tersey, Sarah A.; Sampley, Megan L.; Mosley, Amber; Özcan, Sabire; Pachaiyappan, Boobalan; Woster, Patrick M.; Hunter, Chad S.; Stein, Roland; Mirmira, Raghu.

In: Journal of Biological Chemistry, Vol. 290, No. 15, 10.04.2015, p. 9812-9822.

Research output: Contribution to journalArticle

Maganti, Aarthi V. ; Maier, Bernhard ; Tersey, Sarah A. ; Sampley, Megan L. ; Mosley, Amber ; Özcan, Sabire ; Pachaiyappan, Boobalan ; Woster, Patrick M. ; Hunter, Chad S. ; Stein, Roland ; Mirmira, Raghu. / Transcriptional activity of the islet β cell factor Pdx1 Is augmented by lysine methylation catalyzed by the methyltransferase Set7/9. In: Journal of Biological Chemistry. 2015 ; Vol. 290, No. 15. pp. 9812-9822.
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AU - Maganti, Aarthi V.

AU - Maier, Bernhard

AU - Tersey, Sarah A.

AU - Sampley, Megan L.

AU - Mosley, Amber

AU - Özcan, Sabire

AU - Pachaiyappan, Boobalan

AU - Woster, Patrick M.

AU - Hunter, Chad S.

AU - Stein, Roland

AU - Mirmira, Raghu

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N2 - The transcription factor Pdx1 is crucial to islet β cell function and regulates target genes in part through interaction with coregulatory factors. Set7/9 is a Lys methyltransferase that interacts with Pdx1. Here we tested the hypothesis that Lys methylation of Pdx1 by Set7/9 augments Pdx1 transcriptional activity. Using mass spectrometry and mutational analysis of purified proteins, we found that Set7/9 methylates the N-terminal residues Lys-123 and Lys-131 of Pdx1. Methylation of these residues occurred only in the context of intact, full-length Pdx1, suggesting a specific requirement of secondary and/or tertiary structural elements for catalysis by Set7/9. Immunoprecipitation assays and mass spectrometric analysis using β cells verified Lys methylation of endogenous Pdx1. Cell-based luciferase reporter assays using wild-type and mutant transgenes revealed a requirement of Pdx1 residue Lys-131, but not Lys-123, for transcriptional augmentation by Set7/9. Lys-131 was not required for high-affinity interactions with DNA in vitro, suggesting that its methylation likely enhances post-DNA binding events. To define the role of Set7/9 in β cell function, we generated mutant mice in which the gene encoding Set7/9 was conditionally deleted in β cells (Set Δβ). SetΔβ mice exhibited glucose intolerance similar to Pdx1-deficient mice, and their isolated islets showed impaired glucose-stimulated insulin secretion with reductions in expression of Pdx1 target genes. Our results suggest a previously unappreciated role for Set7/9-mediated methylation in the maintenance of Pdx1 activity and β cell function.

AB - The transcription factor Pdx1 is crucial to islet β cell function and regulates target genes in part through interaction with coregulatory factors. Set7/9 is a Lys methyltransferase that interacts with Pdx1. Here we tested the hypothesis that Lys methylation of Pdx1 by Set7/9 augments Pdx1 transcriptional activity. Using mass spectrometry and mutational analysis of purified proteins, we found that Set7/9 methylates the N-terminal residues Lys-123 and Lys-131 of Pdx1. Methylation of these residues occurred only in the context of intact, full-length Pdx1, suggesting a specific requirement of secondary and/or tertiary structural elements for catalysis by Set7/9. Immunoprecipitation assays and mass spectrometric analysis using β cells verified Lys methylation of endogenous Pdx1. Cell-based luciferase reporter assays using wild-type and mutant transgenes revealed a requirement of Pdx1 residue Lys-131, but not Lys-123, for transcriptional augmentation by Set7/9. Lys-131 was not required for high-affinity interactions with DNA in vitro, suggesting that its methylation likely enhances post-DNA binding events. To define the role of Set7/9 in β cell function, we generated mutant mice in which the gene encoding Set7/9 was conditionally deleted in β cells (Set Δβ). SetΔβ mice exhibited glucose intolerance similar to Pdx1-deficient mice, and their isolated islets showed impaired glucose-stimulated insulin secretion with reductions in expression of Pdx1 target genes. Our results suggest a previously unappreciated role for Set7/9-mediated methylation in the maintenance of Pdx1 activity and β cell function.

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