Extended Data Fig. 10: Induction of ER stress responses and progressive TAGLN2 loss in adoptively transferred CER T cells.
From: Transgelin 2 guards T cell lipid metabolism and antitumour function
a, FSH-CER retroviral construct. b, Expression of the follicle-stimulating hormone receptor (FSHR) by PPNM cancer cells determined by immunoblot analysis. β-actin was used as loading control. c,d, CD45.1+CER T cells were isolated from the indicated tumour sites seven days after adoptive transfer into WT female mice developing PPNM HGSC. Experimental scheme (c). Xbp1s, Sec61a1, ERdj4, and Tagln2 expression in pre- or post-infusion CER T cells was determined via qRT-PCR. Data were normalized to Actb (n = 12) (d). e-h, FACS histograms and quantitative analysis of TAGLN2 and surface FABP5 expression in the indicated T cells isolated from peritoneal lavage (e) or omentum (f) at different days after adoptive transfer into WT female mice developing PPNM HGSC (D3: n = 6 per group, D7 and D14: n = 7 per group). Dark blue bar graph represents the expression of TAGLN2 and surface FABP5 in pre-infusion CER T cells (n = 1). FACS plots and quantitative analysis of CD44+IFN-γ+ (g) and CD44+GZMB+ (h) frequencies in the indicated CER T cells isolated from the same mice described in e and f (n = 6-7 per group). i,j, CD8+GFP+ sorted CER T cells were stimulated with recombinant chorionic gonadotropin alpha (CGα) in the absence or presence of TM (i). Xbp1s, Sec61a1, and Tagln2 expression was determined via qRT-PCR. Data were normalized to Actb (n = 3 per condition) (j). k, Experimental scheme for CER T cell killing assays. T cells were washed to remove TM and then cocultured with PPNM cancer cells at a 1:1 ratio. l, FACS plots and quantitative analysis of cancer cell death assessed by Annexin V and PI staining 18 h after incubation with mock-transduced CD8+ T cells (n = 3-4 per condition). Data are presented as mean ± s.e.m. d, Two-tailed paired Student’s t-test. e-h, One-way ANOVA with Tukey’s multiple comparisons test. j, Two-tailed unpaired Student’s t-test. P < 0.05 is considered statistically significant and exact P-values are shown. The ‘n’ values represent biologically independent samples. gMFI, Geometric mean fluorescence intensity. The images in a, c, i and k were created using BioRender (https://biorender.com).
