Extended Data Fig. 2: Effect of IRE1α-XBP1 inactivation on normal mouse and human bone marrow hematopoietic progenitor cell generation and function.

(a) Xbp1s/Xbp1 total gene expression (±s.e.m., N = 4 mice/group) in purified LSK and GMP cells. (b-e) Representative flow cytometry of Lin^- and LSK cells (b,d) and average (± S.E.M.) proportions (c,e) of BM progenitor cell subsets in adult (3-4 months) mice with HSC-targeted (Vav-iCre) deletion of Ern1 (N = 4 mice/group) (b and c) or Xbp1 (N = 3 mice/group) (d and e) and age-matched littermate controls. Mouse genotypes (a-e): Ern1^wt, Vav-iCre^-Ern1^fl/fl; Ern1^ko, Vav-iCre⁺Ern1^fl/fl; Xbp1^wt, Vav-iCre^-Xbp1^fl/fl; Xbp1^ko, Vav-iCre⁺Xbp1^fl/fl. Each data point represents one mouse. ns, not significant; unpaired Students t-Test. (f-i) Effect of IRE1α inhibition by the small molecule inhibitor MKC8866 (8866) on purified human HSC activity from two healthy donors. For colony-forming unit (CFU) assay, 1 ×10³ FACS-purified HSCs were cultured in MethoCult H4434 (STEMCELL Technologies) supplemented with vehicle (DMSO) or MKC8866 at a final concentration of 10 mm. Data show qRT-PCR-determined XBP1S/XBP1 gene expression ratio (f), representative pictures of CFU assay wells (g), total CFU counts in duplicates wells (h) and enumeration of progenitor cell type (i) after 14 days. Data in f,h,i are average (± S.E.M.) of duplicate wells and the results were reproducible in the two donors.

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