Novel Chimeric Peptide Inhibits Protein Kinase C and Induces Apoptosis in Human Immune Cells |
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Authors: | Ana María Perdomo-Arciniegas Manuel Elkin Patarroyo Jean-Paul Vernot |
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Institution: | (1) Cellular and Molecular Physiology, Faculty of Medicine, Universidad Nacional de Colombia, Bogota, Colombia;(2) Fundación Instituto de Inmunología de Colombia, Bogota, D.C., Colombia |
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Abstract: | Protein kinase C (PKC) participates in a myriad of cellular processes. Protein kinase C isoforms play different roles based
on their cellular expression balance and activation. The activity of classical PKC isoforms has been shown to be crucial for
immune cell population homeostasis, playing a positive role in survival and proliferation. Protein kinase C inhibitors have
been used for conditions where up-regulated PKC results in a pathological state. The most commonly investigated PKC inhibitors
are highly effective in inhibiting PKC function but they are relatively unspecific, some of them even inhibiting other kinase
families. Protein kinase C pseudosubstrates are auto-inhibitory domains which have been used to inhibit more specifically
PKC in vitro but they do not freely penetrate cells. This could be resolved by using cell-permeable PKC pseudosubstrates which
would more accurately modulate cellular PKC activity and PKC-related functions in intact cells. Here we show the development
of a chimeric peptide inhibitor of classical PKC isoforms, consisting of a cell permeable sequence and a pseudosubstrate sequence
which was able to translocate into cells, inhibiting PKC kinase activity and PKC T-cell-specific substrate phosphorylation.
We also demonstrate a dramatic reduction in T-cell proliferation at high chimeric peptide concentration; this was attributed
to apoptosis induction, as demonstrated by cell shrinking, phosphatidylserine exposure and DNA fragmentation. As expected,
the control peptide (pseudosubstrate) did not penetrate cells, affect cell proliferation or survival. We also show that a
neoplastic T-cell line which expresses higher levels of PKC is more resistant to chimeric peptide-mediated cell death than
normal cells, corroborating a PKC role in apoptosis resistance. This chimeric peptide could be useful for the specific modulation
of the PKC signalling pathway in pathological conditions. |
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Keywords: | Protein kinase C Apoptosis PKC inhibitor Signal transduction Peptide internalisation Functional cargo |
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