A novel zebrafish jak2aV581F model shared features of human JAK2V617F polycythemia vera
Received 27 June 2009; received in revised form 28 August 2009; accepted 31 August 2009. published online 23 September 2009.
Objective
The Janus kinase 2 (JAK2) is important for embryonic primitive hematopoiesis. A gain-of-function JAK2 (JAK2V617F) mutation in human is pathogenetically linked to polycythemia vera (PV). In this study, we generated a zebrafish ortholog of human JAK2V617F (referred herewith jak2aV581F) by site-directed mutagenesis and examined its relevance as a model of human PV.
Materials and Methods
Zebrafish embryos at one-cell stage were injected with jak2aV581F mRNA (200pg/embryo). In some experiments, the embryos were treated with a specific JAK2 inhibitor, TG101209. The effects of jak2a stimulation on hematopoiesis, jak/stat signaling, and erythropoietin signaling were evaluated at 18-somites.
Results
Injection with jak2aV581F mRNA significantly increased erythropoiesis, as enumerated by flow cytometry based on gfp+ population in dissociated Tg(gata1:gfp) embryos. The response was reduced by stat5.1 morpholino coinjection (control: 4.37% ± 0.08%; jak2aV581F injected: 5.71% ± 0.07%, coinjecting jak2aV581F mRNA and stat5.1 morpholino: 4.66% ± 0.13%; p<0.01). jak2aV581F mRNA also upregulated gata1 (1.83 ± 0.08 fold; p=0.005), embryonic α-hemoglobin (1.61 ± 0.12 fold; p=0.049), and β-hemoglobin gene expression (1.65 ± 0.13–fold; p=0.026) and increased stat5 phosphorylation. These responses were also ameliorated by stat5.1 morpholino coinjection or treatment with a specific JAK2 inhibitor, TG101209. jak2aV581F mRNA significantly reduced erythropoietin gene (0.24 ± 0.03 fold; p=0.006) and protein expression (control: 0.633±0.11; jak2aV581F mRNA: 0.222±0.07 mIU/mL; p=0.019).
Conclusion
The zebrafish jak2aV581F model shared many features with human PV and might provide us with mechanistic insights of this disease.
aDepartment of Medicine, The University of Hong Kong, Hong Kong
bSchool of Medicine, Deakin University, Waurn Ponds, Victoria, Australia
cDepartment of Biology and Chemistry, City University of Hong Kong, Hong Kong
dDepartment of Pathology, The University of Hong Kong, Hong Kong
Offprint requests to: Anskar Y.H. Leung, M.D., Ph.D., Department of Medicine, Queen Mary Hospital, The University of Hong Kong, Pok Fu Lam Road, Room K418, K Block, Hong Kong
ABSTRACT