Browsing by Subject "PKC-ALPHA"

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  • Talman, Virpi; Provenzani, Riccardo; af Gennäs, Gustav Boije; Tuominen, Raimo K.; Yli-Kauhaluoma, Jari (2014)
  • Karhu, S. Tuuli; Ruskoaho, Heikki; Talman, Virpi (2021)
    Cardiac fibrosis is characterized by accumulation and activation of fibroblasts and excessive production of extracellular matrix, which results in myocardial stiffening and eventually leads to heart failure. Although previous work suggests that protein kinase C (PKC) isoforms play a role in cardiac fibrosis and remodeling, the results are conflicting. Moreover, the potential of targeting PKC with pharmacological tools to inhibit pathologic fibrosis has not been fully evaluated. Here we investigated the effects of selected PKC agonists and inhibitors on cardiac fibroblast (CF) phenotype, proliferation, and gene expression using primary adult mouse CFs, which spontaneously transdifferentiate into myofibroblasts in culture. A 48-hour exposure to the potent PKC activator phorbol 12-myristate 13-acetate (PMA) at 10 nM concentration reduced the intensity of a-smooth muscle actin staining by 56% and periostin mRNA levels by 60% compared with control. The decreases were inhibited with the pan-PKC inhibitor Gö6983 and the inhibitor of classical PKC isoforms Gö6976, suggesting that classical PKCs regulate CF transdifferentiation. PMA also induced a 33% decrease in 5-bromo-2’-deoxyuridine–positive CFs, which was inhibited with Gö6983 but not with Gö6976, indicating that novel PKC isoforms (nPKCs) regulate CF proliferation. Moreover, PMA downregulated the expression of collagen-encoding genes Col1a1 and Col3a1 nPKC-dependently, showing that PKC activation attenuates matrix synthesis in CFs. The partial PKC agonist isophthalate derivative bis(1-ethylpentyl) 5-(hydroxymethyl)isophthalate induced parallel changes in phenotype, cell cycle activity, and gene expression. In conclusion, our results reveal distinct PKC-dependent regulation of CF transdifferentiation and proliferation and suggest that PKC agonists exhibit potential as an antifibrotic treatment.
  • Provenzani, Riccardo; Tarvainen, Ilari Matti Elias; Brandoli, Giulia; Lempinen, Antti Tapani; Artes, Sanna; Turku, Ainoleena; Jäntti, Maria Helena; Talman, Virpi; Yli-Kauhaluoma, Jari Tapani; Tuominen, Raimo Kalevi; Boije af Gennäs, Per Gustav (2018)
    Protein kinase C (PKC) isoforms play a pivotal role in the regulation of numerous cellular functions, making them extensively studied and highly attractive drug targets. Utilizing the crystal structure of the PKC delta C1B domain, we have developed hydrophobic isophthalic acid derivatives that modify PKC functions by binding to the C1 domain of the enzyme. In the present study, we aimed to improve the drug-like properties of the isophthalic acid derivatives by increasing their solubility and enhancing the binding affinity. Here we describe the design and synthesis of a series of multisubstituted pyrimidines as analogs of C1 domain - targeted isophthalates and characterize their binding affinities to the PKC alpha isoform. In contrast to our computational predictions, the scaffold hopping from phenyl to pyrimidine core diminished the binding affinity. Although the novel pyrimidines did not establish improved binding affinity for PKC alpha compared to our previous isophthalic acid derivatives, the present results provide useful structure-activity relationship data for further development of ligands targeted to the C1 domain of PKC.
  • Chronopoulos, Antonios; Thorpe, Stephen D.; Cortes, Ernesto; Lachowski, Dariusz; Rice, Alistair J.; Mykuliak, Vasyl V.; Rog, Tomasz; Lee, David A.; Hytönen, Vesa P.; Hernandez, Armando E. del Rio (2020)
    A mechanism of cell response to localized tension shows that syndecan-4 synergizes with EGFR to elicit a mechanosignalling cascade that leads to adaptive cell stiffening through PI3K/kindlin-2 mediated integrin activation. Extensive research over the past decades has identified integrins to be the primary transmembrane receptors that enable cells to respond to external mechanical cues. We reveal here a mechanism whereby syndecan-4 tunes cell mechanics in response to localized tension via a coordinated mechanochemical signalling response that involves activation of two other receptors: epidermal growth factor receptor and beta 1 integrin. Tension on syndecan-4 induces cell-wide activation of the kindlin-2/beta 1 integrin/RhoA axis in a PI3K-dependent manner. Furthermore, syndecan-4-mediated tension at the cell-extracellular matrix interface is required for yes-associated protein activation. Extracellular tension on syndecan-4 triggers a conformational change in the cytoplasmic domain, the variable region of which is indispensable for the mechanical adaptation to force, facilitating the assembly of a syndecan-4/alpha-actinin/F-actin molecular scaffold at the bead adhesion. This mechanotransduction pathway for syndecan-4 should have immediate implications for the broader field of mechanobiology.