Rhodamine B-conjugated encrypted vipericidin nonapeptide is a potent toxin to zebrafish and associated with in vitro cytotoxicity.
BACKGROUND
Animal venoms contain a diverse array of proteins and enzymes that are toxic toward various physiological systems. However, there are also some practical medicinal uses for these toxins including use as anti-bacterial and anti-tumor agents.


METHODS
In this study, we identified a nine-residue cryptic oligopeptide, KRFKKFFKK (EVP50) that is repeatedly encoded in tandem within vipericidin sequences.


RESULTS
EVP50 displayed in vivo potent lethal toxicity to zebrafish larvae (LD50=6 μM) when the peptide's N-terminus was chemically conjugated to rhodamine B (RhoB). In vitro, RhoB-conjugated EVP50 (RhoB-EVP50) exhibited a concentration-dependent cytotoxic effect toward MCF-7 and MDA-MB-231 breast cancer cells. In MCF-7 cells, the RhoB-EVP50 nonapeptide accumulated inside the cells within minutes. In the cytoplasm, the RhoB-EVP50 induced extracellular calcium influx and intracellular calcium release. Membrane budding was also observed after incubation with micromolar concentrations of the fluorescent EVP50 conjugate.


CONCLUSIONS
The conjugate's interference with calcium homeostasis, its intracellular accumulation and its induced membrane dysfunction (budding and vacuolization) seem to act in concert to disrupt the cell circuitry. Contrastively, unconjugated EVP50 peptide did not display neither toxic nor cytotoxic activities in our in vivo and in vitro models.


CONCLUSIONS
The synergic mechanism of toxicity was restricted to the structurally modified encrypted vipericidin nonapeptide.
State Key Laboratory of Quality Research in Chinese Medicine and Institute of Chinese Medical Sciences, University of Macau, Macao, China.
Liang
L
Wang
State Key Laboratory of Quality Research in Chinese Medicine and Institute of Chinese Medical Sciences, University of Macau, Macao, China.
Judy Y W
JY
Chan
Northeast Biotechnology Network (RENORBIO), Post-graduation program in Biotechnology, Federal University of Ceara, Brazil; Laboratory of Biochemistry and Biotechnology, Institute for Marine Sciences, Federal University of Ceara, Brazil.
Juciane V
JV
Rêgo
State Key Laboratory of Quality Research in Chinese Medicine and Institute of Chinese Medical Sciences, University of Macau, Macao, China.
Cheong-Meng
CM
Chong
State Key Laboratory of Quality Research in Chinese Medicine and Institute of Chinese Medical Sciences, University of Macau, Macao, China.
Nana
N
Ai
Laboratory of Biochemistry and Biotechnology, Institute for Marine Sciences, Federal University of Ceara, Brazil.
Cláudio B
CB
Falcão
Laboratory of Biochemistry and Biotechnology, Institute for Marine Sciences, Federal University of Ceara, Brazil. Electronic address: gandhi.radis@ufc.br.
Gandhi
G
Rádis-Baptista
State Key Laboratory of Quality Research in Chinese Medicine and Institute of Chinese Medical Sciences, University of Macau, Macao, China. Electronic address: simonlee@umac.mo.
Simon M Y
SM
Lee
D000595:Amino Acid Sequence; D000818:Animals; D023181:Antimicrobial Cationic Peptides; D000970:Antineoplastic Agents / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology* / Q000633:toxicity; D001943:Breast Neoplasms / Q000378:metabolism / Q000473:pathology*; D020013:Calcium Signaling / Q000187:drug effects; D054804:Cathelicidins / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology* / Q000633:toxicity; D002462:Cell Membrane / Q000187:drug effects / Q000473:pathology; D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D005260:Female; D006801:Humans; D007814:Larva / Q000187:drug effects; D007928:Lethal Dose 50; D061986:MCF-7 Cells; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology* / Q000633:toxicity; D012235:Rhodamines / Q000378:metabolism / Q000494:pharmacology* / Q000633:toxicity; D013997:Time Factors; D014757:Viper Venoms / Q000737:chemistry*; D015027:Zebrafish / Q000196:embryology*
D023181:Antimicrobial Cationic Peptides; D000970:Antineoplastic Agents; D054804:Cathelicidins; D009842:Oligopeptides; D012235:Rhodamines; D014757:Viper Venoms; C029773:rhodamine B
Cell-penetrating peptide; Cytotoxin; Encrypted peptides; Rhodamine B-conjugated peptide; Vipericidins; Zebra-fish model