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?-thymosins and hemocyte homeostasis in a crustacean.

ABSTRACT: Thymosin proteins are well known for their actin-binding activity. Thymosin beta 4 (T?4) has been associated with biological activities in tissue repair and cell migration via interaction with ATP-synthase in vertebrates, while the information of similar thymosin functions in invertebrates is limited. We have shown previously that ATP-synthase is present on the surface of crayfish hematopoietic tissue (HPT) cells, and that astakine 1 (Ast1, an invertebrate cytokine) was found to interact with this ?-subunit of ATP synthase. Here, we identified five different ?-thymosins from Pacifastacus leniusculus, designated Pl-?-thymosin1-5. The two dominant isoforms in brain, HPT and hemocytes, Pl-?-thymosin1 and 2, were chosen for functional studies. Both isoforms could bind to the ?-subunit of ATP-synthase, and Pl-?-thymosin1, but not Pl-?-thymosin2, significantly increased extracellular ATP formation. Moreover, Pl-?-thymosin1 stimulated HPT cell migration in vitro and Ast1 blocked this effect. Pl-?-thymosin2 increased the circulating hemocyte number at an early stage after injection. Additionally, in vivo injection of Pl-?-thymosin1 resulted in significant reduction of reactive oxygen species (ROS) production in crayfish HPT whereas Pl-?-thymosin2 had a similar but transient effect. Both Pl-?-thymosins induced the expression of Ast1 and superoxide dismutase (SOD) transcripts, while silencing of endogenous Pl-?-thymosin 1 and 2 by RNAi resulted in significant reduction of the Ast1 and SOD transcripts. The diverse effects exhibited by Pl-?-thymosin1 and Pl-?-thymosin2 indicates that these proteins are involved in a complex interaction that regulates the hematopoietic stem cell proliferation and differentiation.

SUBMITTER: Saelee N

PROVIDER: S-EPMC3614969 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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β-thymosins and hemocyte homeostasis in a crustacean.

Saelee Netnapa N Noonin Chadanat C Nupan Benjamas B Junkunlo Kingkamon K Phongdara Amornrat A Lin Xionghui X Söderhäll Kenneth K Söderhäll Irene I

PloS one 20130402 4

Thymosin proteins are well known for their actin-binding activity. Thymosin beta 4 (Tβ4) has been associated with biological activities in tissue repair and cell migration via interaction with ATP-synthase in vertebrates, while the information of similar thymosin functions in invertebrates is limited. We have shown previously that ATP-synthase is present on the surface of crayfish hematopoietic tissue (HPT) cells, and that astakine 1 (Ast1, an invertebrate cytokine) was found to interact with th ...[more]

PMID: 23565293

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Project description: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.

Dataset Information

?-thymosins and hemocyte homeostasis in a crustacean.

Publications

β-thymosins and hemocyte homeostasis in a crustacean.

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