Antimicrobial and host cell-directed activities of Gly/Ser-rich peptides from salmonid cathelicidins.
Cathelicidins, a major family of vertebrate antimicrobial peptides (AMPs), have a recognized role in the first line of defense against infections. They have been identified in several salmonid species, where the putative mature peptides are unusually long and rich in serine and glycine residues, often arranged in short multiple repeats (RLGGGS/RPGGGS) intercalated by hydrophobic motifs. Fragments of 24-40 residues, spanning specific motifs and conserved sequences in grayling or brown, rainbow and brook trout, were chemically synthesized and examined for antimicrobial activity against relevant Gram-positive and Gram-negative salmonid pathogens, as well as laboratory reference strains. They were not active in complete medium, but showed varying potency and activity spectra in diluted media. Bacterial membrane permeabilization also occurred only under these conditions and was indicated by rapid propidium iodide uptake in peptide-treated bacteria. However, circular dichroism analyses indicated that they did not significantly adopt ordered conformations in membrane-like environments. The peptides were not hemolytic or cytotoxic to trout cells, including freshly purified head kidney leukocytes (HKL) and the fibroblastic RTG-2 cell line. Notably, when exposed to them, HKL showed increased metabolic activity, while a growth-promoting effect was observed on RTG-2 cells, suggesting a functional interaction of salmonid cathelicidins with host cells similar to that shown by mammalian ones. The three most active peptides produced a dose-dependent increase in phagocytic uptake by HKL simultaneously stimulated with bacterial particles. The peptide STF(1-37), selected for further analyses, also enhanced phagocytic uptake in the presence of autologous serum, and increased intracellular killing of live E. coli. Furthermore, when tested on HKL in combination with the immunostimulant β-glucan, it synergistically potentiated both phagocytic uptake and the respiratory burst response, activities that play a key role in fish immunity. Collectively, these data point to a role of salmonid cathelicidins as modulators of fish microbicidal mechanisms beyond a salt-sensitive antimicrobial activity, and encourage further studies also in view of potential applications in aquaculture.
Department of Medical and Biological Sciences, Piazzale Kolbe 4, 33100 Udine, Italy.
Francesca
F
D'Este
Department of Life Sciences, University of Trieste, Via Giorgieri 5, 34127 Trieste, Italy.
Monica
M
Benincasa
Department of Life Sciences, University of Trieste, Via Giorgieri 5, 34127 Trieste, Italy.
Giuseppe
G
Cannone
Department of Life Sciences, University of Trieste, Via Giorgieri 5, 34127 Trieste, Italy.
Michela
M
Furlan
Department of Medical and Biological Sciences, Piazzale Kolbe 4, 33100 Udine, Italy.
Michele
M
Scarsini
Department of Agricultural, Food, Environmental and Animal Sciences, Via Sondrio 2/a, 33100 Udine, Italy.
Donatella
D
Volpatti
Department of Life Sciences, University of Trieste, Via Giorgieri 5, 34127 Trieste, Italy.
Renato
R
Gennaro
Department of Life Sciences, University of Trieste, Via Giorgieri 5, 34127 Trieste, Italy.
Alessandro
A
Tossi
Department of Medical and Biological Sciences, Piazzale Kolbe 4, 33100 Udine, Italy.
Barbara
B
Skerlavaj
Department of Life Sciences, University of Trieste, Via Giorgieri 5, 34127 Trieste, Italy. Electronic address: mscocchi@units.it.
Marco
M
Scocchi
D000595:Amino Acid Sequence; D000818:Animals; D000890:Anti-Infective Agents / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D054804:Cathelicidins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology*; D029941:Fish Proteins / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology; D000072417:Protein Domains; D012487:Salmonidae / Q000235:genetics / Q000276:immunology* / Q000382:microbiology; D016415:Sequence Alignment / Q000662:veterinary
D000890:Anti-Infective Agents; D054804:Cathelicidins; D029941:Fish Proteins
Antibacterial activity; Antimicrobial peptide; Head kidney leukocytes; Immunomodulation; Trout cathelicidin