Abstract
Tribbles homolog 2 (Trib2) is a member of Tribbles protein pseudokinases and involves in apoptosis, autoimmunity, cancer, leukemia and erythropoiesis, however, the physiological function of Trib2 in hematopoietic system remains to be elucidated. Here, we report that Trib2 knockout (KO) mice manifest macrocytic anemia and increase of T lymphocytes. Although Trib2 deficient RBCs have similar half-life as the control RBCs, Trib2 KO mice are highly vulnerable to oxidant-induced hemolysis. Endogenous Trib2 mRNA is expressed in early hematopoietic progenitors, erythroid precursors, and lymphoid lineages, but not in mature RBCs, myeloid progenitors and granulocytes. Consistently, flow cytometric analysis and in vitro colony forming assay revealed that deletion of Trib2 mainly affected erythroid lineage development, and had no effect on either granulocyte or megakaryocyte lineages in bone marrow. Furthermore, a genetic approach using double knockout of Trib2 and C/ebpα genes in mice suggested that Trib2 promotes erythropoiesis independent of C/ebpα proteins in vivo. Finally, ectopic expression of human Trib2 in zebrafish embryos resulted in increased expression of erythropoiesis-related genes and of hemoglobin. Taking all data together, our results suggest that Trib2 positively promotes early erythrocyte differentiation and is essential for tolerance to hemolysis.
Highlights
In the peripheral blood were measured by Abbott Cell Dyn 3700 at the indicated time points during recovery from anemia (n = 4 for the control group; n = 11~12 for PHZ-treated group)
Complete blood count analysis of peripheral blood from tribbles homolog 2 (Trib2)+/+ and Trib2−/− mice revealed that Trib2−/− mice had statistically significant decreased levels of RBC numbers, hemoglobin and hematocrit compared to Trib2+/+ mice (p < 0.005, Fig. 1A), but numbers of platelets and neutrophils were similar (Fig. 1A)
The increased mean corpuscular volume was consistent with the RBC volume, as assessed by flow cytometry (Fig. 1B), suggesting a phenotype of macrocytic anemia (Fig. 1A)
Summary
Trib[2] is shown to be selectively expressed in the premegakaryocyte erythroid progenitors (preMegEs)[6], and in a chromatin immunoprecipitation assay its promoter sequences were bound by Gata-2/Fog-1 in a population of sorted preMegEs6. These results prompted us to investigate the physiological role of Trib[2] in hematopoietic lineage commitment and development. We observed an obvious decrease in erythroid progenitors, but not granulocytes or megakaryocytes, in Trib2−/− mice Both kinase domain and COP1-binding domain, that are essential to degrade C/ebpα, of Trib[2] are required for its ability to promote erythropoiesis in the Trib2-/- bone marrow cells. Promotion of hemoglobin production and increased expression of various globin mRNAs in zebrafish further confirmed the pro-erythropoietic function of Trib[2]
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