Fig. 5: Degree of activation of MPL/JAK2/STAT determines the phenotype.

a The γ2A cells were transfected to express JAK2, mu MPL, hu MPL or an empty vector (PIG) and equal amounts of either the hu CALR wt (black), del52 (dark orange) or ins5 (dark blue) or the chim CALR del52 (light orange) or ins5 (light blue), as indicated. STAT5-dependent transcriptional activity with or without TPO was measured 24 h after transfection by the dual firefly luciferase system with Spi-Luc reporter (STAT5 response elements) and renilla pRL-TK reporter with constitutive expression, as an internal control. Shown are means ± SEM of three independent experiments in triplicate. Significance was assessed using Tukey’s multiple comparison test (**p = 0.026; ***p = 0.0004 (hu CALR ins5), 0.0001 (chim CALR del52), 0.0002 (chim CALR ins5); ns not significant). b BM cellularity and spleen weight of 1 month post-tamoxifen del52/del52 KI mice (red) and wild-type (+/+) littermates (white) treated with romiplostim (+Rom, hatched histograms) or not (∅, solid histograms). Data are means ± SEM (n = 3 different mice) and significance of +Rom to ∅ condition was assessed using a two-sided parametric paired t-test: *p = 0.0198; **p = 0.0017; ns: not significant. Source data are provided as a Source data file. c BM and spleen were examined after vWF immunostainning and RET staining. Images were obtained using a DM2000 Leica microscope and a DFC300FX Leica camera with Leica Application Suite v.2.5,OR1 acquisition software (×25 magnifications). Scale bars represent 50 μm. Similar results were obtained with analyzing three independent mice for each condition.