Abstract
BackgroundTransglutaminases (TGases) form a group of enzymes that have many different substrates and among the most well known are fibrin for Factor XIIIa and the clotting protein in crustaceans. We also found that TGase is an abundant protein in the hematopoietic tissue (Hpt) cells of crayfish and hence we have studied the possible function of this enzyme in hematopoiesis.ResultsTGase is one of the most abundant proteins in the Hpt and its mRNA expression as well as enzyme activity is very high in the Hpt cells, lesser in the semi-granular hemocytes and very low in the granular cells. In cultured hematopoietic tissues, high activity was present in cells in the centre of the tissue, whereas cells migrating out of the tissue had very low TGase activity. RNAi experiments using dsRNA for TGase completely knocked down the transcript and as a result the cell morphology was changed and the cells started to spread intensely. If astakine, a cytokine directly involved in hematopoiesis, was added the cells started to spread and adopt a morphology similar to that observed after RNAi of TGase. Astakine had no effect on TGase expression, but after a prolonged incubation for one week with this invertebrate cytokine, TGase activity inside and outside the cells was completely lost. Thus it seems as if astakine addition to the Hpt cells and RNAi of TGase in the cell culture will lead to the same results, i.e. loss of TGase activity in the cells and they start to differentiate and spread.ConclusionThe results of this study suggest that TGase is important for keeping the Hpt cells in an undifferentiated stage inside the hematopoietic tissue and if expression of TGase mRNA is blocked the cells start to differentiate and spread.This shows a new function for transglutaminase in preventing hematopoietic stem cells from starting to differentiate and migrate into the hemolymph, whereas their proliferation is unaffected. Astakine is also important for the hematopoiesis, since it induces hemocyte synthesis in the Hpt but now we also show that it in some unknown way participates in the differentiation of the Hpt cells.
Highlights
Transglutaminases (TGases) form a group of enzymes that have many different substrates and among the most well known are fibrin for Factor XIIIa and the clotting protein in crustaceans
Astakine is important for the hematopoiesis, since it induces hemocyte synthesis in the hematopoietic tissue (Hpt) but we show that it in some unknown way participates in the differentiation of the Hpt cells
TGase is abundant in Hpt cells TGase has earlier been shown to be an important enzyme in the clotting reaction of crayfish and other crustaceans [4], and here we report that TGase
Summary
Transglutaminases (TGases) form a group of enzymes that have many different substrates and among the most well known are fibrin for Factor XIIIa and the clotting protein in crustaceans. Transglutaminases (TGases) form a family of Ca2+dependent enzymes catalyzing post-translational remodelling of proteins such as, cross-linking in homeostasis, keratinocyte cornified envelope formation and semen coagulation [1,2,3]. These enzymes catalyze a R-glutaminylpeptide:amine-γ-glutamyl-transferase reaction leading to cross-linking within or between polypeptide chains, or covalent incorporation of polyamines into protein substrates. The horseshoe crab TGase was recently found to participate in the host defence in the cuticle by cross-linking Caraxin-1, a component of cuticle, into a stable mesh, which promotes wound healing and sclerotization at injured sites of the cuticle [12]
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