Cyclic AMP-Dependent Protein Kinase Isoenzymes in Human Myeloid Leukemia (HL60) and Breast Tumor (MCF-7) Cells

Taimi, Mohammed; Breitman, Theodore R.; Takahashi, Noriko

doi:10.1006/abbi.2001.2443

Cited by 12 publications

(8 citation statements)

References 41 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…RIIβ is an important physiological inhibitor of the PKA catalytic subunit as demonstrated by the complex phenotypes associated with its disruption 17–21 . To understand the unique features of RIIβ and for eventually designing isoform-specific agonists and antagonists, it is essential to have structures that represent RIIβ in its different conformational states.…”

Section: Discussionmentioning

confidence: 99%

“…The balance of type I and type II activity and expression also plays an important role in cell growth and differentiation 15,16 . Disruption of this balance is implicated in several disease mechanisms, including the autoimmune disease, systemic lupus erythematosus (SLE/Lupus), Carney complex, and breast cancer 17–21 . To understand the role that RIIβ plays in disease and to realize the potential for designing isoform-specific agonists and antagonists, it is important to elucidate the structural and functional features that make RIIβ unique.…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Novel Isoform-Specific Interfaces Revealed by PKA RIIβ Holoenzyme Structures

Brown

Kim

et al. 2009

Journal of Molecular Biology

View full text Add to dashboard Cite

The PKA catalytic (C) subunit is inhibited by two classes of functionally non-redundant regulatory subunits, RI and RII. Unlike RI-subunits, RII-subunits are both substrates and inhibitors. Because RIIβ knockout mice have important disease phenotypes, the RIIβ holoenzyme is a target for developing isoform-specific agonists and/or antagonists. We also know little about the linker region that connects the inhibitor site to the N-terminal dimerization domain although this linker determines the unique globular architecture of the RIIβ holoenzyme. To understand how RIIβ functions as both an inhibitor and substrate and to elucidate the structural role of the linker, we engineered different RIIβ constructs. In the absence of nucleotide RIIβ(108–268) that contains a single cyclic nucleotide binding domain bound C-subunit poorly whereas with AMP-PNP, a non-hydrolyzable ATP analog, the affinity was 11nM. The RIIβ(108–268) holoenzyme structure (1.62Å) with AMP-PNP/Mn2+, showed that we trapped the RIIβ-subunit in an enzyme:substrate complex with the C-subunit in a closed conformation. The enhanced affinity afforded by AMP-PNP/Mn2+ may be a useful strategy for increasing affinity and trapping other protein substrates with their cognate protein kinase. Because mutagenesis predicted that the region N-terminal to the inhibitor site might dock differently to RI and RII, we also engineered RIIβ(102–265) that contained six additional linker residues. The additional linker residues in RIIβ(102–265) increased the affinity to 1.6 nM suggesting that docking to this surface may also enhance catalytic efficiency. In the corresponding holoenzyme structure this linker docks as an extended strand onto the surface of the large lobe. This hydrophobic pocket, formed by the αF-αG loop and conserved in many protein kinases, also provides a docking site for the amphipathic helix of PKI. This novel orientation of the linker peptide provides the first clues as to how this region contributes to the unique organization of the RIIβ holoenzyme.

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Novel Isoform-Specific Interfaces Revealed by PKA RIIβ Holoenzyme Structures

Brown

Kim

et al. 2009

Journal of Molecular Biology

View full text Add to dashboard Cite

show abstract

“…In the absence of retinoids, treatment of NB4 cells with cAMP elevating agents induces only growth inhibition, while HL-60 cells undergo granulocytic maturation in the same experimental conditions [217,218]. In both cell types, co-treatment with ATRA and cAMP analogs, or phosphodiesterase IV inhibitors enhances granulocytic maturation of HL-60 and NB4 cells [38,86,[220][221][222][223][224]. This is accompanied by the appearance of maturation markers of the terminally differentiated granulocyte, such as leukocyte alkaline phosphatase [86,220].…”

Section: Retinoids and The Camp Pathwaymentioning

confidence: 99%

Retinoids as Differentiating Agents in Oncology: A Network of Interactions with Intracellular Pathways as the Basis for Rational Therapeutic Combinations

Garattini¹,

Giannı̀²,

Terao

2007

CPD

View full text Add to dashboard Cite

Retinoic acid and natural as well as synthetic derivatives (retinoids) are promising anti-neoplastic agents endowed with both therapeutic and chemopreventive potential. Although the treatment of acute promyelocic leukemia with all-trans retinoic acid is an outstanding example, the full potential of retinoids in oncology has not yet been exploited and a more generalized use of these compounds is not yet a reality. This may be the result of issues such as natural and induced resistance as well as local and systemic toxicity. One way to enhance the therapeutic and chemopreventive activity of retinoic acid and derivatives is to identify rational combinations between these compounds and other pharmacological agents. This is now possible given the wealth of information available on the biochemical and molecular mechanisms underlying the biological activity of retinoids. At the cellular level, the anti-leukemia and anti-cancer activity of retinoids is the result of three main actions, cell-differentiation, growth inhibition and apoptosis. At the molecular level, retinoids act through the activation of nuclear-retinoic-acid-receptor-dependent and-independent pathways. The cellular pathways and molecular networks relevant for retinoid activity are modulated by a panoply of other intra-cellular and extra-cellular pathways that may be targeted by known drugs and other experimental therapeutics. The review article aims to summarize and critically discuss the available knowledge in the field and provide a rational framework that may be useful for the design of effective drug combinations with the potential to enhance the therapeutic index of retinoids.

show abstract

“…The heat-stable protein kinase inhibitors (PKIs) are a family of small proteins that inhibit PKA by binding to the substrate-binding site of the catalytic subunit with subna-nomolar affinity. 32 The high-affinity binding motif is found at residues 5-24, and the respective 20-mer peptide spanning this range (named PKI (5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24) or IP20) binds to the C-subunit with high affinity, with a K i of 2.3 nM. 33 Thus, IP20 is the ideal probe for this novel method.…”

Section: Resultsmentioning

confidence: 99%

“…There are four isoforms of the PKA regulatory subunit (RIR, RIβ, RIIR, RIIβ) that diversify PKA-mediated signaling by differing in their abundance, affinity for the C-subunit, sensitivity to cAMP, and specificity for different AKAPs. 1,2 PKA is implicated in several cancers (e.g., carney complex, 3 pituitary, 4 and breast 5,6 ) and is also suggested to be a therapeutic target for diseases of the immune system (e.g., SLE and HIV). 7 However, given the ubiquitous distribution of this protein kinase, it is unwise to target the ATP-binding site of the catalytic subunit, since an ATP-competitive inhibitor would just as likely kill healthy cells.…”

mentioning

confidence: 99%

Assay Principle for Modulators of Protein−Protein Interactions and Its Application to Non-ATP-Competitive Ligands Targeting Protein Kinase A

et al. 2006

View full text Add to dashboard Cite

Targeting sites that modulate protein-protein interactions represents an ongoing challenge for drug discovery. We have devised an assay principle, named Ligand-Regulated Competition (LiReC), in an effort to find non-ATP competitive small molecule regulators for type Iα cAMP-dependent Protein Kinase A (PKA-Iα), a protein complex that is implicated in disease. Our assay based on the LiReC principle utilizes a competitive fluorescent peptide probe to assess the integrity of the PKA-Iα complex upon introduction of an allosteric ligand. The developed fluorescence polarization method screens for small molecules that specifically protect (antagonists) or conversely activate (agonists) this protein complex. In high throughput format, various cyclic nucleotide-derived agonists and antagonists are successfully detected with high precision. Furthermore, assay performance (Z'-factors above 0.7) far exceeds the minimum requirement for small molecule screening. To identify compounds that operate through novel modes of action, our method shields the ATP binding site and purposely excludes ATP-competitive ligands. These proof-of-principle experiments highlight the potential of the LiReC technique and suggest its application to other protein complexes, thereby providing a novel approach to identify and characterize modulators (small molecules, proteins, peptides, or nucleic acids) of protein-protein systems.

show abstract

Cyclic AMP-Dependent Protein Kinase Isoenzymes in Human Myeloid Leukemia (HL60) and Breast Tumor (MCF-7) Cells

Cited by 12 publications

References 41 publications

Novel Isoform-Specific Interfaces Revealed by PKA RIIβ Holoenzyme Structures

Novel Isoform-Specific Interfaces Revealed by PKA RIIβ Holoenzyme Structures

Retinoids as Differentiating Agents in Oncology: A Network of Interactions with Intracellular Pathways as the Basis for Rational Therapeutic Combinations

Assay Principle for Modulators of Protein−Protein Interactions and Its Application to Non-ATP-Competitive Ligands Targeting Protein Kinase A

Contact Info

Product

Resources

About