Fig 2: Subclustering basal stem cells point to the BAS-I cluster as essential to epidermal homeostasis.a Subclustering basal clusters (BAS-I – BAS-IV) using SoptSC. b Heatmap showing top 100 differentially expressed genes per cluster. c Correlation between overlapping differentially expressed genes from basal clusters and subclustering of basal clusters (kBAS) in A. d Pseudotime inference of kBAS keratinocytes. e RNA velocity of kBAS keratinocytes using 1000 vectors. Cells color-coded as in A. f Relative gene expression of KRT14 and KRT10 across the kBAS subclusters. g Differentiation score of kBAS subclusters using average gene expression of GRN genes. h Violin plot of relative gene expression of PTTG1 and RRM2 across kBAS subclusters. i Relative mRNA expression of indicated genes along Ca2+-induced differentiation of primary human keratinocytes. n = 3 experiments each time point (n = 2 for PCNA). j Coimmunostaining of PTTG1 (red) and RRM2 (green) in human neonatal skin. Green arrowheads highlight RRM2-only expressing cells; yellow arrowheads point to PTTG1-RRM2 double-positive cells. White dotted line denotes basement membrane. Scale bar 50 µm. n = 3. k Knock-down (KD) of PTTG1 and RRM2, with a scramble control, in human skin equivalent organoids grown for 16 days. Hematoxylin and eosin (H&E) staining shown. Dotted lines denote basement membrane. Scale bar 100 µm. n = 4 experiments each condition. l Quantification of total cells per squared area (*p = 0.0104; ***p < 0.0001; n = 12 areas each [PTTG1 and RRM2 KD]), (m) KI67 + cells per 200 µm area along the basement membrane (*p = 0.0123; ***p < 0.0001; n = 26 areas [PTTG1] or n = 25 areas [RRM2]), and (n) qPCR of the respective transcripts for each KD condition (PTTG1: p = 0.0056, 0.0018; RRM2: p = 0.0485, 0.025; ASS1: p = 0.0025; GJB2: p = 0.0037; PCNA: p = 0.0339, 0.0295; n = 3 experiments each [PTTG1 and RRM2 KD]). For whisker and box plots: box represents 25th to 75th percentiles, whiskers represent minimum and maximum data points, and bar represents mean. For bar graph: error bar represents SEM. For scatter plot: data are presented as mean values ± SEM. Significance was determined by unpaired two-tailed t test.

Fig 3: Quantitative immunoblots for (A) FTH1, (B) HMOX1, (C) Ribonucleotide reductase regulatory subunit M2 (RRM2), and (D) nuclear receptor coactivator 4 (NCOA4) in WT and Pink1−/− MEF, under untreated control conditions (Ctrl), after iron overload (FAC) and after two different iron depletion drugs (DFO, 22BP), administered over 48 h. Protein abundance signals were normalized to beta-Actin levels (Actin beta (ACTB)) as a loading control. The panels on the right show their densitometric quantifications, normalized to WT untreated conditions. WT n = 4–6, Pink1−/− n = 3, one exemplary set is shown. The statistical trends or levels of significance are illustrated by symbols, namely T: 0.1 > p>0.05, *: p < 0.05, **: p < 0.01, ###: p < 0.001, ****: p < 0.0001. Mutant cells are represented by dashed bars, WT cells by plain colors. Asterisks represent significance in WT MEF, treated versus untreated control, while hashtags refer to Pink1−/− MEF, treated versus untreated control. Genotype-dependent significant differences of Pink1−/− versus WT MEF are illustrated by horizontal lines below asterisks.

Fig 4: Irradiation of the pediatric GBM cells enhanced proliferation and expression of malignant-promoting proteins:(A) Growth curves of SJGBM2 and SJGBM2-10gy cells. (B) Western blot for RR M1 (94 kDa) and M2 (45 kDa) subunits and pro-cathepsin B (Pro-CatB) (43 kDa) in SJGBM2 and SJGBM2-10gy cells. (C) Direct immunofluorescence probing for RRM2 and cathepsin B (Pro-CatB) in both cells after a 24 h incubation in fresh medium. *P < 0.05.

Fig 5: Association of XBP1-gene signature with outcome in ER-positive breast cancer. A Box plot for expression of RRM2, CDC6 and TOP2A in tumour and normal tissues in human breast cancers is shown. Median is shown by horizontal black line, the box is the upper and lower quartiles and the two lines outside the box show the highest and lowest values. B KM Plotter (https://kmplot.com/analysis/) was used to determine the association of XBP1-gene signature (CDC6, RRM2 and TOP2A) with overall survival (OS) and recurrence free survival (RFS) in ER-positive breast cancer. C Web-based algorithm PROGgeneV2 (http://www.progtools.net/gene/index.php) was used to test the association between XBP1-gene signature and RFS in indicated datasets of breast cancer. D Web-based algorithm ROC plot (http://www.rocplot.org/) was used to evaluate the association between XBP1-gene signature and response to tamoxifen treatment in ER-positive breast cancer