An ENU-induced recessive mutation in Mpl

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• 作者：E. Ricky Chan ; Heather Lavender ; Geqiang Li ; Peter Haviernik ; Kevin D. Bunting ; Mark D. Adams
• 刊名：Experimental Hematology
• 出版年：2009
• 出版时间：February 2009
• 年：2009
• 卷：37
• 期：2
• 页码：276-284
• 全文大小：626 K

文摘

Objective

The aim of this study was to identify and characterize the causative mutation in the thrombocytopenic mouse strain HLB219 that was generated at the Jackson Laboratory as part of a large-scale N-ethyl-N-nitrosourea mutagenesis screen.

Materials and Methods

The HLB219 mutation was identified by interval mapping of F2 mice generated from intercross breeding of HLB219 to both BALB/cByJ (BALB) and 129/SvImJ (129/Sv). Mpl was identified as a candidate gene and sequenced. The mutation was characterized in vivo in mouse hematopoietic stem/progenitor cell assays and in cell culture by expression in Ba/F3 cells.

Results

A novel mutation in the thrombopoietin (TPO) receptor Mpl in HLB219 mice caused a Cys→Arg substitution at codon 40 in the extracellular region of the receptor. Mice homozygous for the Mpl^hlb219 mutation had an 80 % decrease in the number of platelets in comparison to the wild-type C57BL/6J strain, low numbers of bone marrow megakaryocytes, high TPO levels, and decreased competitive repopulating ability, consistent with a loss-of-function mutation in the receptor. Mice heterozygous for Mpl^hlb219 however, showed an overdominance effect with a significant increase in platelet number. Functional analysis in vitro demonstrated that Ba/F3 cells expressing the mutant MPL^hlb219 protein failed to activate extracellular signal-regulated kinase and signal transducers and activators of transcription 5, but proliferated in the absence of TPO and required constitutive phosphorylation of RAC-alpha serine/threonine protein kinase (AKT) for cytokine-independent growth.

Conclusion

Thrombocytopenia in HLB219 mice is caused by a recessive mutation in Mpl that abrogates mitogen-activated protein kinase–extracellular signal regulated kinase and janus kinase–signal transducers and activators of transcription signaling.