A new multigene HCIQ subfamily from the sea anemone Heteractis crispa encodes Kunitz-peptides exhibiting neuroprotective activity against 6-hydroxydopamine

Abstract

the Kunitz/Bpti-type peptides are ubiquitous in numerous organisms including marine venomous animals. the peptides demonstrate various biological activities and therefore they are the subject of a number of investigations. We have discovered a new HciQ subfamily belonging to recently described multigene HcGS family of Heteractis crispa Kunitz-peptides. the uniqueness of this subfamily is that the HciQ precursors contain a propeptide terminating in Lys-Arg (endopeptidase cleavage site) the same as in the neuro-and cytotoxin ones. Moreover, the HCIQ genes contain two introns in contrast to HCGS genes with one intron. As a result of Sanger and amplicon deep sequencings, 24 HCIQ isoforms were revealed. The recombinant peptides for the most prevalent isoform (HCIQ2c1) and for the isoform with the rare substitution Gly17Glu (HCIQ4c7) were obtained. They can inhibit trypsin with K i 5.2 × 10 −8 M and K i 1.9 × 10 −7 M, respectively, and interact with some serine proteinases including inflammatory ones according to the SPR method. For the first time, Kunitz-peptides have shown to significantly increase neuroblastoma cell viability in an in vitro 6-OHDA-induced neurotoxicity model being a consequence of an effective decrease of ROS level in the cells. Kunitz-type proteinase inhibitors are present in various living organisms and in viruses. They are widely distributed and well-characterized in animals including marine invertebrates, snakes, spiders, ticks, flies and mammals 1. In spider, snake, scorpion, cone snail and sea anemone venoms Kunitz-peptides may exist in multiple isoforms possessing conserved BPTI-like fold but exhibit different biological activities 2-9. This phenomenon is associated with gene duplication and their diversification throughout adaptive evolution leading to the formation of families of evolutionarily related but functionally distinct genes 10. Among sea anemone transcriptomes, such multigene families have been discovered in Anemonia viridis 11 , Stichodactyla haddoni 12 , and Heteractis crispa 5. HCGS multigene family of H. crispa has been found to be divided in four distinct subfamilies (GS, RG, GG, and GN) forming the combinatory library of Kunitz/BPTI peptides 5. The group of HCGN peptides was presented by one sequence, different from other sequences that are characterized by a propeptide insertion containing the cleavage site Lys-Arg, and additional residues Ile-Gln at the N-terminus of a mature peptide. Its full-length homolog HMIQ3c1, containing a mature peptide with the same residues at N-terminus, was derived from the cDNA of the sea anemone Heteractis magnifica 13. The most abundant HCGS and HCRG peptides are being actively studied nowadays 14-17. The main targets of Kunitz-peptides are serine proteinases, such as trypsin and α-chymotrypsin.

A Review of Toxins from Cnidaria

“…The Kunitz-type IQ-peptide HMIQ3c1, extracted from the venom of Heteractis magnifica , has shown neuroprotective activity, which may find application in the treatment of neurodegenerative diseases, such as Alzheimer’s [ 138 ]. Another Kunitz-type proteinase inhibitor that may support the treatment of Parkinson disease is HCIQ2c1 extracted from Heteractis crispa [ 121 ]. This study shows for the first time that Kunitz-peptides significantly increase neuroblastoma cell viability in an in vitro 6- hydroxydopamine-induced neurotoxicity model of Parkinson’s disease [ 121 ].…”

“…Another Kunitz-type proteinase inhibitor that may support the treatment of Parkinson disease is HCIQ2c1 extracted from Heteractis crispa [ 121 ]. This study shows for the first time that Kunitz-peptides significantly increase neuroblastoma cell viability in an in vitro 6- hydroxydopamine-induced neurotoxicity model of Parkinson’s disease [ 121 ].…”

Kunitz-Type Peptides from the Sea Anemone Heteractis crispa Demonstrate Potassium Channel Blocking and Anti-Inflammatory Activities

“…Moreover, these peptides contain conserved amino acids at the N-and C-termini. Besides trypsin, Kunitz-type peptides from sea anemone H. crispa are also able to interact with other serine proteases (chymotrypsin, elastase, kallikrein) [19,39,53], modulate or block TRPV1 channel [22][23][24], revealing different kind of biological effects such as analgesic [22,[54][55][56], anti-inflammatory [9,41,54,57], antihistamine [41,58], as well as neuroprotective activity [59,60]. However, despite their high degree of homology with the known bifunctional peptides, like kalicludines, SHTXIII, APEKTx1, and ShPI-1 [16,18,19], none of them has shown potassium channels blocking activity.…”

Cnidarian toxins: recent evidences for potential therapeutic uses

“…Further, a novel Kunitz-like peptide (PcKuz3) has been identified from Palythoa caribaeorum, as a neuroprotective agent providing an opportunity to develop a treatment for neurodegenerative diseases (Liao et al 2019). According to a latest research, another type of kunitz peptide was reported that exhibit neuroprotective activity against 6-hydroxydopamine (Kvetkina et al 2020).…”