Fig. 2: CUL5 knockout (KO) promotes sustained proliferation and enhances effector functions later after stimulation. | Nature Communications

Fig. 2: CUL5 knockout (KO) promotes sustained proliferation and enhances effector functions later after stimulation.

From: Cullin-5 deficiency promotes chimeric antigen receptor T cell effector functions potentially via the modulation of JAK/STAT signaling pathway

Fig. 2: CUL5 knockout (KO) promotes sustained proliferation and enhances effector functions later after stimulation.The alternative text for this image may have been generated using AI.

a–d Changes in the percentage of CUL5 KO-CD19 CAR T cells generated by sgRNA#4690 from Days 10–20 (a) and 20–30 (b) or by sgRNA#8377 from Days 10–20 (c) and 20–30 (d). (n = 4 in a and b; n = 3 in c and d) (e) Representative flow plots of CellTrace Violet (CTV)-labeled control (Ctrl) or CUL5 KO-CD19 CAR T-cell division assays. GFP− fractions are unmodified CAR T cells, whereas GFP+ fractions are CUL5 KO-CAR T cells. f, g CTV mean fluorescence intensity (MFI) after stimulation at either earlier (f, Days 0–3, n = 3) or later time points (g, Days 4–7, n = 5) in Ctrl or CUL5 KO-CD19 CAR T cells. h, i Ki-67+ cell percentage after stimulation during earlier (h, Days 0–3, n = 4) or later (i, Days 4–7, n = 5) culture periods in Ctrl or CUL5 KO-CD19 CAR T cells. j Intracellular cytokine staining of Ctrl or CUL5 KO-CD19 CAR T cells stimulated with K562 or Raji cells. Representative flow plots of interferon-gamma (IFN-γ) and interleukin (IL)-2 are shown. k, l Intracellular cytokine staining for IFN-γ (k) and IL-2 (l). (n = 5) (m) CD25 MFI after Raji stimulation in Ctrl or CUL5 KO-CD19CAR T cells. Data were expressed as mean ± SEM. (n = 3) (n) Phenotype analysis of Ctrl and CUL5 KO-CD19 CAR T cells for CD45RA and C-C chemokine receptor 7 (CCR7) on Day 30. o, p Percentages of CD45RA( − )-CCR7(−) effector memory T cells and CD45RA( − )-CCR7(+) central memory T cells after CD3/CD28 beads stimulation. q, r Percentages of CD45RA( − )-CCR(−) effector memory T cells and CD45RA( − )-CCR7(+) central memory T cells after Raji cell stimulation. s CCR7 MFI after Raji stimulation. (n = 5 in (o–s) Data from three to five different donors are acquired and each dot represents data from each donor. The paired two-sided Student’s t-test was used for (a–d, f–i, and o–s). The two-way ANOVA with the Šidák multiple comparisons test was used for (k and l). The multiple unpaired t-test was used for (m). Source data are provided as a Source Data file.

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