Caspase-1-Like Regulation of the proPO-System and Role of ppA and Caspase-1-Like Cleaved Peptides from proPO in Innate Immunity

Abstract

Invertebrates rely on innate immunity to respond to the entry of foreign microorganisms. One of the important innate immune responses in arthropods is the activation of prophenoloxidase (proPO) by a proteolytic cascade finalized by the proPO-activating enzyme (ppA), which leads to melanization and the elimination of pathogens. Proteolytic cascades play a crucial role in innate immune reactions because they can be triggered more quickly than immune responses that require altered gene expression. Caspases are intracellular proteases involved in tightly regulated limited proteolysis of downstream processes and are also involved in inflammatory responses to infections for example by activation of interleukin 1ß. Here we show for the first time a link between caspase cleavage of proPO and release of this protein and the biological function of these fragments in response to bacterial infection in crayfish. Different fragments from the cleavage of proPO were studied to determine their roles in bacterial clearance and antimicrobial activity. These fragments include proPO-ppA, the N-terminal part of proPO cleaved by ppA, and proPO-casp1 and proPO-casp2, the fragments from the N-terminus after cleavage by caspase-1. The recombinant proteins corresponding to all three of these peptide fragments exhibited bacterial clearance activity in vivo, and proPO-ppA had antimicrobial activity, as evidenced by a drastic decrease in the number of Escherichia coli in vitro. The bacteria incubated with the proPO-ppA fragment were agglutinated and their cell morphology was altered. Our findings show an evolutionary conserved role for caspase cleavage in inflammation, and for the first time show a link between caspase induced inflammation and melanization. Further we give a more detailed understanding of how the proPO system is regulated in time and place and a role for the peptide generated by activation of proPO as well as for the peptides resulting from Caspase 1 proteolysis.

An updated molecular basis for mussel immunity

“…There are no previous reports of inflammasomes in invertebrates [120] and the absence of pyrin domains (characterizing both NALPs and the key adaptor protein PYCARD) in invertebrates, as well the absence of a caspase-1 homologous sequence in mussel implies that a similar system for the activation of the inflammatory response, if existing, is highly divergent in molluscs. Furthermore, NLRs seem to have a fundamental role in the activation of the autophagic machinery (specifically via the ATG16L1 protein) at the site of bacterial entry, thus promoting the elimination of bacterial pathogens through xenophagy [121].…”

The clotting system in decapod crustaceans: History, current knowledge and what we need to know beyond the models

“…But before go any further, it is important to know that conversely to insects, where the relationship of the proPO system and coagulation is evident [11,100], this relationship is unclear in crustaceans. In addition, the proPO zymogen is a cytoplasmic leaderless protein that is released by an unknown UPS mechanism, probably involving caspase-1 activity [101], or another pathway that remains to be elucidated. Other humoral components of the proPO system are stored in secretory granules and are released by exocytosis [101,102].…”

“…In addition, the proPO zymogen is a cytoplasmic leaderless protein that is released by an unknown UPS mechanism, probably involving caspase-1 activity [101], or another pathway that remains to be elucidated. Other humoral components of the proPO system are stored in secretory granules and are released by exocytosis [101,102]. Both protein releasing processes should coincide temporarily and might involve similar signaling mechanisms/molecules.…”

Adult Neurogenesis: Lessons from Crayfish and the Elephant in the Room

“…Pacifasticus leniusculus and Procambarus clarkii) live for up to 20 years and possess distinct hematopoietic tissues that produce hemocytes throughout the animal's lifetime. Innate immunity has been intensively studied in P. leniusculus , including one of the most important mechanisms, the prophenoloxidase-activating (proPO) system, which leads to the activation of the Toll pathway and production of antimicrobial peptides [Noonin et al, 2012;Jearaphunt et al, 2014]. Astakines, crustacean cytokines belonging to the prokineticin family, are synthesized in hematopoietic tissues and are required for the proliferation and release of hemocytes into the circulation [Söderhäll et al, 2005;Lin et al, 2010;.…”