Leukemia Associated Mutants of SHP2 Protein Tyrosine Phosphatase Inhibit Nf1-Expression by Decreasing ICSBP and IRF2 Phosphorylation.

Neurofibromin 1 (Nf1) is a ubiquitous protein with Ras-GAP activity. In myeloid cells, Nf1 down regulates Ras which has been activated by hematopoietic cytokines. Congenital absence of Nf1 is associated with the myeloproliferative/dysplastic disorder Juvenile Chronic Myelomonocytic Leukemia (JMMoL). JMMoL is characterized by hypersensitivity to cytokines such as GM-CSF, SCF and M-CSF. In previous studies, we found that cytokine stimulation of myeloid progenitor cells increases transcription of the NF1-gene, thereby increasing Nf1-expression. We identified a cis element in the NF1-promoter which is activated by the interferon consensus sequence binding protein (ICSBP). This is of interest because ICSBP-deficiency confers hypersensitivity to the same cytokines as does Nf1-deficiency. In the current studies, we find that interferon regulatory factor 2 (IRF2) cooperates with ICSBP to activate the NF1-cis element in differentiating myeloid cells. This interaction requires specific phosphorylation of conserved tyrosine residues in the IRF-domains of IRF2 and ICSBP. Phosphorylation of both proteins occurs in differentiating myeloid progenitor cells and myeloid cell lines. In myeloid progenitor cells, ICSBP and IRF2 are maintained in a non-phosphorylated state by the protein tyrosine phosphatases SHP1 and SHP2. During myeloid differentiation, activity of these PTPs decreases, permitting increased phosphorylation of ICSBP and IRF2. Leukemia associated, activating mutants of SHP2-PTP have been recently described in JMMoL, MDS and AML. Such mutations induce conformational changes in SHP2, resulting in constitutive activation. When introduced into myeloid progenitor cells, such SHP2-mutants induced a profile of cytokine hypersensitivity which is similar to those seen in Nf1-deficiency or ICSBP-deficiency. We find that expression of one of these leukemia associated SHP2 mutants blocks cytokine-induced phosphorylation of ICSBP and IRF2, impairing Nf1-expression in differentiating myeloid cells. This results in hypersensitivity and sustained proliferation in response to differentiating cytokines. Therefore, our studies mechanistically associate three molecular defects that have been associated with myeloproliferation and cytokine hypersensitivity; ICSBP-deficiency, Nf1-deficiency and expression of activated SHP2 mutants. Our results suggest that hematopoietic cytokine signaling induces tyrosine phosphorylation of IRF2 and ICSBP. These phospho-proteins activate transcription of the NF1-gene. Increased Nf1-expression down regulates cytokine-activated Ras. However, expression of constitutively active forms of SHP2-PTP blocks this process by impairing cytokine induced phosphorylation of IRF2 and ICSBP, decreasing Nf1-expression, and resulting in unopposed Ras activation. These results may be important to targeting this pathway in myeloid disorders characterized by cytokine hypersensitivity and myeloproliferation.