Extended Data Fig. 1: IDR retained within the leukaemia-related NUP98–HOXA9 chimera forms phase-separated condensates in vitro and is essential for establishing phase-separated chimeric transcription factor assemblies in the nucleus.

a, Schematic showing the domain architecture of normal NUP98 (top), normal HOXA9 (middle) and leukaemic NUP98–HOXA9 chimera (bottom; with either GFP or 3×HA–3×Flag tag fused to C terminus). The GLFG or non-GLFG (xFG) motif contents, which make up IDR, and other important domains are shown. GLEBS represents the GLE2-binding sequence, which directs the NUP98 interaction with GLE2 (also known as RAE1) for mRNA export when NUP98 acts as component of nuclear pore complex⁶⁰. Red arrows indicate the common breakage point of NUP98 and HOXA9. b, Immunoblotting of NUP98–HOXA9, either full-length (WT) or with GLEBS deleted (Δ132-224; see a), as detected by the indicated antibodies after stable transduction into primary mouse HSPCs. For gel source data, see Supplementary Fig. 1. c, Mouse HSPCs stably transduced with wild-type or GLEBS-deleted NUP98–HOXA9 showed similar proliferation in liquid cultures (n = 3 independent cell cultures per group), in agreement to previous reports^17,61. Empty vector (EV)-infected HSPCs served as a control. Data are mean ± s.d. d, Live-cell fluorescence imaging (GFP; with zoomed-in and zoomed-out views shown in the top and bottom panels, respectively) of 293FT cells with stable transduction of GFP-tagged NUP98–HOXA9, wild-type, GLEBS-deleted (also referred to as N-IDR_WT/A9; see Fig. 1a) or carrying a DNA-binding-defective mutation in homeodomain (HD_N51S) or a Phe-to-Ser mutation that substitutes Phe residues within all FG repeats to Ser (IDR_FS, also referred to as N-IDR_FS/A9; see Fig. 1a). The right panel shows immunoblotting of endogenous normal NUP98 in 293FT cells, as well as the stably transduced exogenous NUP98–HOXA9, either wild-type (lane 1) or GLEBS-deleted (lane 2), as detected by antibodies against GLEBS of NUP98¹⁷. For gel source data, see Supplementary Fig. 1. Scale bars, 10 μm. e, Schematic of the indicated N-IDR fusion domains with a varying number of FG repeats. The IDR portion used for in vitro assay in main Fig. 1d is indicated by a red dotted line. f, SDS–PAGE images showing recombinant N-IDR domain protein with the indicated varying number of FG repeats (His6×-tagged; see e), purified with Ni-column and an additional size exclusion column purification step. The protein size is labelled above the recombinant protein. g, Anti-GFP immunoblotting for GFP-tagged NUP98–HOXA9 chimera with the indicated varying number of FG repeats described in e after stable transduction in 293FT cells. For gel source data, see Supplementary Fig. 1. h, Live-cell fluorescence imaging for the N51S-mutated N-IDR/A9 (GFP-tagged) with either wild-type (top) or the Phe-to-Ser mutated IDR (bottom) in 293FT stable expression lines before (left) and after (right) treatment with 10% 1,6-hexanediol for 1 min. The left panels show zoomed-in images of a representative cell from the right panels of zoomed-out cell images. Scale bar, 10 μm.