Extended Data Fig. 3: Functional consequences of Ybx1 deletion in vivo.

a, Schematic representation of the wild-type Ybx1 allele (Genomic Locus), the targeting vector (Targeting Vector), the desired targeted allele (Targeted Allele), the desired conditional allele flanked by LoxP sequences (Conditional Allele) and the null recombined allele (Recombined Allele) after Cre-mediated recombination of the conditional allele. Triangles indicate loxP sequences. b, Excision control by genomic PCR on whole bone marrow cells at week 16 following genetic inactivation of Ybx1 in conditional knockout mice. Representative micrograph of n = 5 animals from a total of n = 9 controls (+/+) and n = 9 knockout (−/−) replicates. c, Schematic as in Fig. 2j. FACS plots showing percentage of Jak2^V617F (CD45.2) cells of Ybx1^+/+ or Ybx1^−/− recipients. d, Histology of liver, spleen and lung of Jak2^V617F-Ybx1^+/+ and Jak2^V617F-Ybx1^−/− recipient mice at week 20 after BMT. Representative micrographs of n = 9 individual mouse replicates. Haematoxylin and eosin stain (H&E) at ×10 magnification. Focal leukocyte infiltration (arrows) and haemorrhage (stars) of liver, spleen and lung, respectively. Scale bar: 200 μm. e, Peripheral blood chimerism of lethally irradiated (12 Gy) recipient mice. FACS plots showing abundance of CD45.2 myeloid cells in Jak2^V617F-Ybx1^+/+ and Jak2^V617F-Ybx1^−/− recipient mice at week 20 after BMT. f, Design for assessment of steady state haematopoiesis. g, White blood count (WBC), Gr-1 positive cells (Gr1⁺), haemoglobin (HGB) and platelets (PLT) following genetic inactivation of Ybx1 (Ybx1^−/− mice, n = 10) compared to Ybx1^+/+ controls (n = 10). Data represented as mean ± s.e.m. h, FACS plots showing comparable percentages of LSK cells and HSCs (SLAM⁺CD34^–L^–S⁺K⁺ cells) following genetic inactivation of Ybx1 in conditional knockout mice (compared to wildtype littermate controls). i, stem- and progenitor cell numbers per 1 × 10⁶ whole bone marrow cells at week 16 after genetic inactivation of Ybx1 (n = 6; mean ± s.d.). j, FACS plot showing comparable abundance of mature myeloid and erythroid cells following genetic deletion of Ybx1. k, Total numbers of mature blood cells of the myeloid (Gr1⁺), erythroid (TER119⁺), B-lymphoid (CD19) and T-lymphoid (CD3) lineages at week 16 after genetic inactivation of Ybx1 (n = 6; mean ± s.d.). l, Experimental protocol for investigation of haematopoietic progenitor cell function. m, Colony numbers of Ybx1^+/+ versus Ybx1^−/− mouse stem/progenitor cells. Colonies were counted at day 8 after plating (each sample plated in duplicate, n = 3 independent experiments, mean ± s.d.). n, Spleen colony numbers counted on day 12 after injection of Ybx1^+/+ or Ybx1^−/− LSK cells into lethally irradiated (12Gy) recipient mice (CFU-S12) (n = 12 Ybx1^+/+; n = 12 Ybx1^−/− independent biological mouse replicates in n = 3 independent cohorts). o, BM chimerism of primary recipient mice (n = 10 individual biological replicates) at week 20 after BMT. Whole bone marrow chimerism (WBMC) and chimerism of myeloid cells (Gr1⁺ BMC) (left panel). HSPC chimerism (LSK) and HSC chimerism (CD34^– LSK) (right panel). Data shown as mean ± s.d. p, BM chimerism of secondary recipient mice (n = 5 individual biological replicates) at week 20 after BMT. Whole BM chimerism (WBMC) and chimerism of myeloid cells (Gr1⁺ BMC) (left panel). HSPC chimerism (LSK) and HSC chimerism (CD34^– LSK) (right panel). Data shown as mean ± s.d.