Cathepsin B/X is secreted by Echinometra lucunter sea urchin spines, a structure rich in granular cells and toxins

Abstract

BackgroundEchinometra lucunter is a common American sea urchin responsible for the majority of the marine accidents in Brazil. Although not lethal, these accidents are reported to be extremely painful. Recently, our group described the presence of toxins in its spines that contribute to the pathological reactions. Additionally, we have observed that the E. lucunter spines can regenerate when broken. In the present work we evaluated the enzymatic activities of sea urchin spine extracts in order to identify an enzyme that could contribute not only to the toxicity, but also participate in the spine growth and regeneration.ResultsThe spine aqueous extract was tested for peptidase activity, with synthetic substrates, in the presence and absence of inhibitors and activators. For proper enzyme classification, the FRET-substrate cleavage pattern, pH-dependency activity and Western-blot analyses were performed. The spine extract was able to cleave Z-R-MCA and Abz-GIVRAK(Dnp)-OH following pre-incubation with DTT, and was inhibited by E-64. Furthermore, the double-peaked pH curve (5 and 7) and the cleavage site proportion (4:6, R↓A:A↓K) indicate the presence of both mono and dicarboxypeptidase activities. Moreover, in Western-blot analysis, the spine extract was positive for anti-cathepsin B antibody.ConclusionsE. lucunter spines extracts presented a cysteine peptidase activity that was identified as cathepsin B/X that would participate in the remodeling and growth processes of the spine, as well as in the inflammatory response to the accident.

Initial peptidomic profiling of Brazilian sea urchins: Arbacia lixula, Lytechinus variegatus and Echinometra lucunter

“…Interesting biological activities were found for the matched sequences, such as antibiotic, antitumor, phospholipase A 2 inhibitors or neuroprotective. A similarity was also found with a toxin from scorpion venom, which confirms the presence of toxins in sea urchins, as recently described by our group [ 6 , 7 ].…”

“…Briefly, an inflammatory reaction in mice was described for the spine extract of Echinometra lucunter , confirming the presence of toxins in these structures [ 8 ]. Moreover, our group could identify a cathepsin B/X activity in spines, indicating the presence of enzymes for either chemical defense of the animal or tissue remodeling after injury, a frequent phenomenon observed in our collections [ 7 ].…”

Injuries caused by aquatic animals in Brazil: an analysis of the data present in the information system for notifiable diseases

“…Traumatic wounds by sea urchins are very common, and the permanence of spines in the points of penetration favors secondary infections and the formation of foreign body granulomas (14) . Laboratory assays confi rmed the presence of toxins in some parts of the black sea urchin (Echinometra lucunter) (16) . However, clinical studies conducted in coastal zones of Southeastern Brazil have only found the existence of traumatic injuries and local manifestations in these occurrences (14) .…”

112. Cathepsin B/X is Secreted by Echinometra lucunter Sea Urchin Spines, a Structure Rich in Granular Cells and Toxins

“…This cysteine peptidase cathepsin activity, seemly unique (e.g., due to one single enzyme) was termed as cathepsin B/X, for it presents both carboxi mono-and dipeptidyl peptidase activities, and virtually no endopeptidase activity [25]. Moreover, complementary assays (cleavage pattern of specific substrates, DTT activation, active site E-64 inhibition, pH dependence of the activity, molecular mass evaluation and Ab recognition pattern) corroborate such statement.…”