DNase Activities of a Highly Stable High-Molecular-Mass Multiprotein Complex Isolated from Different Organs of the Sea Cucumber Paracaudina chilensis.

Sea cucumbers exhibit rapid recovery of all organs and the body following various injuries, making them highly promising models for investigating the mechanisms of self-regeneration. This study provides the first comprehensive comparative analysis of DNase activities in both homogenates and highly stable multiprotein complexes isolated from five distinct organs-gut, respiratory trees, body wall, gonads, and coelomic fluid-of the sea cucumber Paracaudina chilensis. Using in situ zymography following SDS-PAGE with copolymerized polymeric DNA, we systematically profiled the molecular weights, optimal pH values, and metal ion dependencies of DNases across these tissues and their corresponding stable complexes (1.2-1.8 MDa). Our results reveal that each organ possesses a unique DNase repertoire, with at least 25 distinct enzymatic species identified on the basis of molecular masses (~19, 23, 30, 35, 50, 100, 130, and 150 kDa) and pH optima ranging from 4.5 to 9.5. Notably, the gut homogenate contained the greatest diversity of DNases, whereas the respiratory tree and coelomic fluid complexes harbored highly active enzymes that were undetectable in the corresponding homogenates, indicating selective enrichment during complex assembly. Most DNases were Mg2+-dependent, with optimal MgCl2 concentrations between 5 and 10 mM; however, two metal-independent DNases (~50 and ~100 kDa) were identified exclusively in respiratory tree complexes, and their activity was strongly enhanced by EDTA, suggesting suppression by endogenous metal ions. Additionally, Ca2+-activated DNases (~19 and ~23 kDa) were found in complexes from body wall, gonads, and coelomic fluid. Several DNases with high relative activity in the complexes were not reliably detected in the corresponding tissue homogenates, whereas some abundant homogenate enzymes were absent from the complexes. This implies that the formation of these supramolecular assemblies is not random but rather involves the specific recruitment of low-abundance enzymes, potentially concentrating them to achieve high local activity for organ-specific functions. The gut, which undergoes complete regeneration after evisceration, contained the most varied set of DNases, hinting at a possible role in DNA metabolism during tissue renewal. Thus, we have shown for the first time that the multiprotein complexes from different organs of the sea cucumber differ not only in molecular weight and protein/peptide composition but also in the number and types of DNases they contain. We propose that in each organ of the sea cucumber, a specific multiprotein complex is formed, which may be necessary to perform additional extended functions of this complex beyond those of canonical individual proteins, peptides, and enzymes.