Myeloproliferative Disease Induced by TEL-PDGFRB Displays Dynamic Range Sensitivity to Stat5 Gene Dosage in Mice.
Authors: Cain Jennifer A. , Chen Jing , Xiang Zhifu , O’Neal Julie , Lee Benjamin H. , Kreisel Friederike , Williams Ifor , Colson AnnaLynn , Tomasson Michael H. 
Published: October 2019

Abstract
Expression of the constitutively activated TEL/PDGFβR fusion protein is associated with the t(5;12)(q33;p13) chromosomal translocation found in a subset of patients with chronic myelomonocytic leukemia (CMML). TEL/PDGFβR activates multiple signal transduction pathways in cell culture systems and induces myeloproliferative disease (MPD) in a murine bone marrow transduction/transplantation model of disease. Two TEL-PDGFβR juxtamembrane tyrosines, corresponding to tyrosines activating Stat5 and Src signaling molecules in native PDGFβR, are required for TEL-PDGFRB mediated MPD in mice. We used gene-targeted mice as donors in bone marrow transduction/transplantation experiments to characterize the contribution of Stat and Src genes in the development of TEL-PDGFRB disease. Mice transplanted with cells harboring targeted deletions of both Stat5a and Stat5b genes (TPiGFP→Stat5ab−/−) were protected from rapidly fatal MPD (median survival >125 vs. 25 days in TPiGFP→Stat5ab+/+mice, P<0.0001). In contrast, TPiGFP→Lyn−/−Hck−/−Fgr−/− mice developed rapidly fatal MPD characterized by splenomegaly and leukocytosis. Similarly, TEL-PDGFRB induced MPD in TPiGFP→Stat1−/− mice and TPiGFP→Stat1+/+ strain-matched mice alike (median survival 45 vs. 53 days). These data suggest that Stat5, but not Src family kinases Lyn, Hck and Fgr nor Stat family member Stat1, is functionally relevant in the development of TEL-PDGFRB mediated disease. To assess the relative contribution of the Stat5a gene in TEL-PDGFRB mediated disease, we used mice harboring either homozygous (Stat5a−/−) or heterozygous (Stat5a+/−) deletion as donors in bone marrow transduction/transplantation experiments with TEL-PDGFRB. Surprisingly, survival was significantly prolonged in TPiGFP→Stat5a−/− mice (144 vs. TPiGFP→Stat5a+/+ 31 days, P <0.0001) and TPiGFP→Stat5a+/− mice (145 versus TPiGFP→Stat5a+/+ 31 days, P <0.001), demonstrating significant sensitivity of TEL-PDGFRB-induced MPD to loss of even a single Stat5a allele. TEL-PDGFRB induced severe leukocytosis in TPiGFP→Stat5a+/+ mice (median WBC 547K/μL), but disease was attenuated in TPiGFP→Stat5a+/− mice mice (median WBC 56K/μL) and undetectable in TPiGFP→Stat5a−/−mice (median WBC 5K/μL), suggesting that TEL-PDGFRB disease is exquisitely sensitive to Stat5a gene dosage. On the other hand, TPiGFP→Stat5b−/− mice developed robust MPD with similar latency to wild-type mice. However, 4 of 13 TPiGFP→Stat5b−/− mice failed to develop MPD and were longer-lived, suggesting a role for Stat5b in TEL-PDGFRB disease penetrance.
To address the possibility that Stat5a and Stat5b were distinct in their ability to transmit myeloproliferative signals in TEL-PDGFRB mediated MPD, we designed add-back constructs to co-express TEL-PDGFRB with either Stat5a (TPiStat5a) or Stat5b (TPiStat5b). Addback Stat5 restored MPD with similar disease latency in TPiStat5a→Stat5a−/− and TPiStat5b→Stat5a−/− mice (median survival 96 and 97 days, respectively). TPiStat5a→Stat5a+/− mice developed MPD with shorter disease latency (28 days), again correlating with Stat5a gene dosage. These data provide functional evidence for the importance of the Stat5 pathway in a model of TEL-PDGFRB-induced CMML and suggest that Stat5a and Stat5b play unique roles in the development of disease in this model.