Fig. 4. PSIP1 acts as a dependency factor in a murine-derived T-ALL cell line and T-ALL maintenance in vivo.

(A) Western blot analysis (left) and quantification (right) of PSIP1 and ACTIN (loading control) in a murine Pten^fl/fl;Lck-Cre^tg/+ (Pten) cell line that was transduced with a scrambled control hairpin or hairpins targeting murine Psip1. The PSIP1 levels normalized against house-keeping gene β-ACTIN. (B) Loss of Psip1 expression also had a negative effect on the cell proliferation rate of a murine Pten cell line (n = 3, two-way ANOVA). (C) Schematic representation of the conducted in vivo experiment. Immunocompromised NXG mice were engrafted with one million of Jurkat cells that were either transduced with a doxycycline-inducible control hairpin (shCtrl) or a hairpin targeting PSIP1 (shPSIP1 2). After 5 days, mice were switched to doxycycline-containing food or maintained on control food. Thirty days after the start of doxycycline treatment, the mice were euthanized, and the percentage of hCD45⁺ leukemic cells was analyzed (created in BioRender). (D) Doxycycline-inducible knockdown of PSIP1 (shPSIP1 2) led to lower levels of hCD45-positive cells in the bone marrow (BM) compared to control mice (shCtrl) (n = 5, Kruskal-Wallis test with Dunn’s multiple comparisons test). *P < 0.05 and ***P < 0.001.