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The Nuclear Protein Chromodomain Helicase Binding Protein 8 is a Novel A‐kinase Anchoring Protein
Author(s) -
O'Donnell Maureen E,
Russell Mary,
Lund Linda,
Zhang Yinghua,
Bloch Robert,
Bond Meredith
Publication year - 2010
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.24.1_supplement.855.4
Subject(s) - chromodomain , arrestin , protein kinase a , helicase , biology , binding domain , microbiology and biotechnology , zinc finger , protein subunit , kinase , binding protein , binding site , biochemistry , transcription factor , signal transduction , gene , rna , g protein coupled receptor
A‐kinase anchoring proteins (AKAPs) anchor the regulatory type II (RII) subunit of protein kinase A (PKA) via an amphipathic a‐helix. Using phage display, we isolated peptides from a chromatin remodeling protein, chromodomain helicase binding protein 8 (Chd8). Our objective was to characterize Chd8 as an AKAP. We analyzed isolated Chd8 peptides and found a candidate site in the amino terminus. We evaluated RII binding with far Western blotting, in which blots of a peptide encompassing this domain were incubated with purified RII. A probe for RII highlighted a band of the same molecular weight as the Chd8 peptide. Mutation of an isoleucine (I464P) within the putative Chd8 RII binding site, and addition of Ht31, a peptide that disrupts PKA:AKAP binding, eliminated this band. RII coimmunoprecipitated with the amino terminus of Chd8, but not the I464P mutant. This demonstrates a novel AKAP domain in Chd8. Using qPCR of embryonic, post‐natal, and adult heart, we demonstrated Chd8 mRNA decreases in development, but is present in adult tissue. Immunoblots and immunocytochemistry identified nuclear Chd8 in cultured cardiac myocytes. We predict that altered PKA binding to Chd8 in response to beta‐adrenergic stimulation regulates interaction of Chd8 with its binding partners and alters activation of gene transcription. This work supported by NIH/NHLBI T32 HL072751, NIH/NIA F31AG032162‐01, NIH R01 AG16613 and RO1 HL75016.