Activation of JAK2-V617F by Heterodimeric Receptor Components.

Abstract 2955

Poster Board II-931

Mutational activation of Janus Kinase 2 (JAK2) is an important etiologic factor for the development of myeloproliferative neoplasms (MPNs). JAK2 is mutated in nearly all patients with polycythemia vera and about half of patients with essential thrombocythemia and primary myelofibrosis. The most prevalent mutation of JAK2 is valine 617 mutated to phenylalanine (V617F). The normal function of JAK2 is to interact with and become activated by cytokine receptors following their interaction with ligand. Interestingly, while JAK2-V617F is considered a constitutively activated kinase, there is evidence that it still requires interaction with a cytokine receptor to elicit its transforming signal. When expressed at less than or near endogenous JAK2 levels in hematopoietic cells, JAK2-V617F requires co-expression of a homodimeric receptor in order to become activated, transduce downstream signals, and induce transformation. When expressed at high levels in hematopoietic cells, co-expression of a homodimeric cytokine receptor is not needed. However, a functional cytokine receptor interacting domain is still required, suggesting that even when expressed at high levels JAK2-V617F requires interaction with a receptor. Also, a functional cytokine receptor interacting domain in JAK2-V617F is required for it to induce MPN-like disease in mouse models. The ability of cytokine receptor expression to activate JAK2-V617F has focused on homodimeric receptors. In this study we demonstrate that single components of heterodimeric receptors can also activate JAK2-V617F. Expression of interleukin-27 receptor alpha (IL27Ra), the ligand-binding component of the IL-27 receptor, enhances phosphorylation of JAK2-V617F on tyrosines 1007 and 1008. This activation of JAK2 also leads to tyrosine phosphorylation of signal transducers and activators of transcription-5 (STAT5), a main downstream effector of JAK2 activation. We obtain similar results when we utilize interleukin-12 receptor beta 1 (IL12RB1), a receptor belonging to the same family as IL27Ra. To extend these studies to other heterodimeric cytokine receptor components, we utilized the components of the interleukin-3 receptor, interleukin-3 receptor alpha (IL3Ra) and the beta common chain, which is also utilized in the receptors for interleukin-5 and granulocyte macrophage colony stimulating factor. Expression of each of these receptor subunits activates JAK2-V617F as well as STAT5. Importantly, we demonstrate that expression of IL27Ra can functionally replace the expression of a homodimeric cytokine receptor to support the activation of JAK2-V617F in BaF3 cells as well as promote the transforming activity of JAK2-V617F in these cytokine dependent hematopoietic cells. Interestingly, while IL12RB1 expression activates JAK2-V617F in 293T cells, it is not capable of enhancing the transforming signals of JAK2-V617F in BaF3 cells. Expression of IL3Ra or the beta common chain in BaF3 cells also enhances the ability of JAK2-V617F to transform these cells to cytokine independence. However, this enhancement is not immediate as it only becomes evident at later time points. Together our data demonstrate that in addition to homodimeric receptors, some heterodimeric receptor components may contribute to JAK2-V617F activation. It should also be considered that such receptors may play a role in JAK2-V617F-negative MPNs, perhaps through altered expression or activating receptor mutations, analogous to mutated thrombopoietin receptor proteins that play a role in the development of disease in a fraction of these MPN patients.