Epidermoid carcinoma-derived antimicrobial peptide (ECAP) inhibits phosphorylation by protein kinases in vitro.
Animal peptide antibiotics are thought to mediate their cytotoxic and growth inhibitory action on bacteria, fungi, and cancer cells through a membrane-targeted mechanism. Although the membrane interactions of the peptide antibiotics and their penetration through the membranes have been studied in several models, the precise chain of events leading to cell death or growth arrest is not established yet. In this study we used in vitro kinase assays followed by imaging analyses to examine the effect of human cationic antimicrobial peptide ECAP on the activity of the protein kinases. We report that HPLC-grade ECAP is responsible for inhibition of EGFR autophosphorylation in plasma membrane fractions obtained from A-431 cells. The activity of ECAP is concentration dependent with a half-inhibitory concentration in the range of 0.1-0.2 microM. Marked decrease in autophosphorylation of immunoprecipitated non-receptor protein kinases belonging to different families, namely PKCmu, Lyn and Syk, is observed in the presence of as little as 0.2 microM of the peptide. Among the examined non-receptor protein kinases PKCmu was the most sensitive to the inhibitory action of ECAP, whereas Syk was inhibited least of all. ECAP exerted no detectable cytotoxicity on non-nucleate animal cells at concentrations up to 3 microM. The capability of ECAP to inhibit protein kinases at concentrations, that are at least 10 fold lower than antibacterial and cytotoxic ones, suggests that the protein kinases are possible intracellular targets for antimicrobial peptides. We suppose that inhibition of the protein kinases may provide a mechanism for the action of cationic antimicrobial peptides on host cells including tumour cells.
Hobta
A
A
R. E. Kavetsky Institute of Experimental Pathology, Oncology, and Radiobiology, NAS of Ukraine, 45 Vasylkivska, 03022 Kyiv, Ukraine. hobta@yahoo.com
Lisovskiy
I
I
Mikhalap
S
S
Kolybo
D
D
Romanyuk
S
S
Soldatkina
M
M
Markeyeva
N
N
Garmanchouk
L
L
Sidorenko
S P
SP
Pogrebnoy
P V
PV
D000818:Animals; D023181:Antimicrobial Cationic Peptides / Q000302:isolation & purification / Q000494:pharmacology*; D002294:Carcinoma, Squamous Cell / Q000737:chemistry*; D002458:Cell Fractionation; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D066246:ErbB Receptors / Q000378:metabolism*; D004912:Erythrocytes / Q000187:drug effects; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D010766:Phosphorylation; D011233:Precipitin Tests; D011494:Protein Kinases / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D023181:Antimicrobial Cationic Peptides; D011494:Protein Kinases; D066246:ErbB Receptors