Endogenous animal toxin-like human β-defensin 2 inhibits own K+ channels through interaction with channel extracellular pore region

Abstract

Human potassium channels are widely inhibited by peptide toxins from venomous animals. However, no human endogenous peptide inhibitor has been discovered so far. In this study, we demonstrate for the first time using electrophysiological techniques, that endogenous human β-defensin 2 (hBD2) is able to selectively and dose-dependently inhibit the human voltage-gated Kv1.3 channel at picomolar peptide concentration. The co-immunoprecipitation assays further supported the selective binding of hBD2 to Kv1.3 channel. Using mutagenesis experiments, we found that the outer pore domain of Kv1.3 channel was the binding site of hBD2, which is similar to the interacting site of Kv1.3 channel recognized by animal toxin inhibitors. The hBD2 was able to suppress IL-2 production through inhibition of Kv1.3 channel currents in human Jurkat cells, which was further confirmed by the lack of hBD2 activity on IL-2 production after Kv1.3 knockdown in these cells. More interestingly, hBD2 was also found to efficiently inhibit Kv1.3 channel currents and suppress IL-2 production in both human primary CD3(+) T cells and peripheral mononuclear cells from either healthy donors or psoriasis patients. Our findings not only evidenced hBD2 as the first characterized endogenous peptide inhibitor of human potassium channels, but also paved a promising avenue to investigate newly discovered function of hBD2 as Kv1.3 channel inhibitor in the immune system and other fields.

Kv Channel S1-S2 Linker Working as a Binding Site of Human β-Defensin 2 for Channel Activation Modulation

“…We immobilized different FLAG-tagged channels on protein G-Sepharose beads and detected whether the channels could retain hBD2 through Western blotting. Our results showed that the Kv1.3 channel could retain hBD2 and Kv1.2 channels retained much less hBD2 than the Kv1.3 channels, which was consistent with our previous studies (7). When the Kv1.3 channel S1-S2 linker was transferred to the Kv1.2 channel, the binding affinity of hBD2 and the S1-S2 linker channel chimera was higher than the binding affinity for the Kv1.2 channel (Fig.…”

“…recruits T cells and influences the cytokine secretion of T cells through interactions with different receptors (7,11,12). Our recent report showed that hBD2 inhibits Kv1.3 channel currents by interacting with the channel extracellular pore region (7).…”

“…Recently, we identified human ␤-defensin 2 (hBD2) as an animal toxin-like endogenous Kv1.3 channel inhibitor that can interact with extracellular pore region (6, 7). Surprisingly, we found that hBD2, unlike the classic channel pore-blocking toxins that generally do not affect Kv channel kinetics (8,9), works simultaneously as a gating modifier and shifts the conductancevoltage relationship (G-V) curve of the Kv1.3 channel to a more depolarized voltage by nearly 10 mV.…”

Scorpion Potassium Channel-blocking Defensin Highlights a Functional Link with Neurotoxin

“…6A, the basic residues in the potassium channel-interacting interfaces of the scorpion toxins BmKTX196, BmKTX and ChTX were found to be responsible for their affinities (8,22). Similar to the functional importance of basic residues in the classical animal toxins, the basic residues in BmKDfsin4 were also used to recognize the negatively charged vestibule of the potassium channel (19,27) (Figs. 4, 5, and 6A).…”

“…Recently, our group first reported two kinds of wild-type defensins that could directly inhibit potassium channels. One kind of wild-type defensin is from mammals, such as human ␤-defensin 2 (hBD2) (27,28), ␣-defensin 1 (HNP1), and ␣-defensin 5 (HD5) (29), and the other kind of wild-type defensin is from fungi, such as plectasin (30). These findings encouraged us to explore whether venomous animal defensins can block potassium channels.…”