Abstract
Protein kinase A (PKA) and the nuclear A-kinase-anchoring protein AKAP95 have previously been shown to localize in separate compartments in interphase but associate at mitosis. We demonstrate here a role for the mitotic AKAP95-PKA complex. In HeLa cells, AKAP95 is associated with the nuclear matrix in interphase and redistributes mostly into a chromatin fraction at mitosis. In a cytosolic extract derived from mitotic cells, AKAP95 recruits the RIIalpha regulatory subunit of PKA onto chromatin. Intranuclear immunoblocking of AKAP95 inhibits chromosome condensation at mitosis and in mitotic extract in a PKA-independent manner. Immunodepletion of AKAP95 from the extract or immunoblocking of AKAP95 at metaphase induces premature chromatin decondensation. Condensation is restored in vitro by a recombinant AKAP95 fragment comprising the 306-carboxy-terminal amino acids of the protein. Maintenance of condensed chromatin requires PKA binding to chromatin-associated AKAP95 and cAMP signaling through PKA. Chromatin-associated AKAP95 interacts with Eg7, the human homologue of Xenopus pEg7, a component of the 13S condensin complex. Moreover, immunoblocking nuclear AKAP95 inhibits the recruitment of Eg7 to chromatin in vitro. We propose that AKAP95 is a multivalent molecule that in addition to anchoring a cAMP/PKA-signaling complex onto chromosomes, plays a role in regulating chromosome structure at mitosis.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Antibodies / pharmacology
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Cell Cycle / drug effects
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Cell Cycle Proteins / metabolism
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Cell Extracts
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Chromatin / chemistry
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Chromatin / genetics
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Chromatin / metabolism*
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Chromosomes, Human / chemistry
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Chromosomes, Human / drug effects
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Chromosomes, Human / genetics
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Chromosomes, Human / metabolism*
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Cyclic AMP / antagonists & inhibitors
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Cyclic AMP / pharmacology
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Cyclic AMP-Dependent Protein Kinases / antagonists & inhibitors
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Cyclic AMP-Dependent Protein Kinases / chemistry
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Cyclic AMP-Dependent Protein Kinases / metabolism*
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DNA-Binding Proteins / antagonists & inhibitors
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DNA-Binding Proteins / chemistry
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / metabolism*
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Egg Proteins / metabolism
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Fluorescent Antibody Technique
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HeLa Cells
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Humans
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Intracellular Signaling Peptides and Proteins
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Mitosis* / drug effects
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Nocodazole / pharmacology
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Nuclear Matrix / drug effects
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Nuclear Matrix / metabolism
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Nuclear Proteins / antagonists & inhibitors
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Nuclear Proteins / chemistry
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Nuclear Proteins / genetics
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Nuclear Proteins / metabolism*
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Peptide Fragments / chemistry
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Peptide Fragments / genetics
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Peptide Fragments / metabolism
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Precipitin Tests
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Protein Binding
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Signal Transduction / drug effects
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Xenopus Proteins*
Substances
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AKAP8L protein, human
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Antibodies
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Cell Cycle Proteins
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Cell Extracts
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Chromatin
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DNA-Binding Proteins
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Eg7 protein, Xenopus
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Egg Proteins
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Intracellular Signaling Peptides and Proteins
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Nuclear Proteins
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Peptide Fragments
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Xenopus Proteins
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Cyclic AMP
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Cyclic AMP-Dependent Protein Kinases
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Nocodazole