Extended Data Fig. 6: YBX1 phospho-null mutants display impaired nuclear localization and increased sensitivity to JAK inhibitor.

a, Western blot showing the expression of YBX1 phosphomutants in Ba/F3 JAK2VF cells as indicated. b, Confocal images of YBX1 localization (Red) in YBX1 phosphomutants expressing Ba/F3 JAK2VF cells. Cells were counterstained with DAPI and GFP confirms the phosphomutant expression. a b, Representative images from n = 3 biological experiments. c, Bar plot showing quantification of nuclear YBX1 expression in YBX1 phosphomutants expressing Ba/F3 JAK2VF cells; P values were determined by two-tailed t-test. Control (n = 67): min = 15.39; max = 33.01; whisker = [15.39–33.01]; median = [24.74], S1(n = 70): min = 4.14; max = 9.75; whisker = [4.14–9.75]; median = [7.0053], S2(n = 64): min = 5.32; max = 14.11; whisker = [5.32–14.11]; median = [7.9], S3(n = 57):min = 4.44; max = 17.6; whisker = [4.44–17.6]; median = [11.4], S4(n = 72): min = 13.32; max = 23.82; whisker = [13.32–23.82]; median = [19.3], S5(n = 61): min = 13.08; max = 27.97; whisker = [13.08–27.97]; median = [22.05], S6(n = 57):min = 18.89; max = 30.04; whisker = [18.89–30.04]; median = [22.92], S7(n = 60): min = 16.69; max = 32.53; whisker = [16.69–32.53]; median = [24.65]. d, Cell growth curve of YBX1 phosphomutants expressing Ba/F3 JAK2VF cells following treatment with increase doses of JAK inhibitor (1 nM–10 μM RUX) measured by MTS assay. n = 4 independent experiments each with 8 technical replicates. e, FACS plots showing induction of apoptosis in Ba/F3 JAK2VF cells expressing YBX1 phosphomutants following JAK inhibitor treatment (RUX, 0.5 μM) compared to untreated YBX1 wild-type Ba/F3 JAK2VF cells. f, Bar plot shows quantification of the percentages of apoptotic (annexin V- and 7-AAD-positive) cells. n = 6 independent experiments since YBX1 phosphomutants endogenously express GFP, n = 3 for annexin V–APC staining and n = 3 for 7-AAD–APC staining (error bars represent mean ± s.d.).