Fig 2: TEAD and coactivator YAP directly bind the SPAG5 promoter.A Schematic representation of the SPAG5 promoter with the putative TEAD-binding sites (LASAGNA Search 2.0). The promoter regions 657–884 and 2909–3084 were amplified and cloned into pGL3 vector. B, C, E, F Chip analysis of the TEAD and YAP binding on SPAG5 promoter in MDA-MB-231 cell line after TEAD interference detected by qRT-PCR analysis. D–G Transcriptional active chromatin on SPAG5 promoter evidenced by anti-H4-Acetylate antibody. Data are shown as the mean of three independent replicates with the a relative P value. H–J Luciferase assay pGL3 vector carrying SPAG5 657–884 and SPAG5 2909–3084 promoter regions was transfected in MDA-MB-231 cell line after YAP and/or TEAD interference. H MDA-MB-231 cell lines were treated with 0.05–0.1 µM of Dasatinib (I) or with 2 µM of Verteporfin (J) and transfected with pGL3 vector carrying SPAG5 657–884 and SPAG5 2909–3084 promoter regions. Normalized luciferase-activity values and P value from three independent experiments are shown. In the lower panel, western blot shows SPAG5 expression after 0.05–0.1 µM of Dasatinib (I) or with 2 µM of Verteporfin (J) treatments. (ns nonsignificant; *P value <0.05; **P value <0.001).

Fig 3: YAP signature and SPAG5 are prognostic independent factors for disease-specific survival of breast cancer METABRIC patient dataset.A Kaplan–Meier curve for disease-specific survival on breast cancer patients stratified for different combined levels of YAP signature and SPAG5 expression. Cox regression multivariate analysis adjusted for T, N, stage, istotype, menopause, and age. B Heatmaps of standardized expression levels of SPAG5, YAP signature, and miR-10b-3p on breast cancer Metabric dataset. C Pie-chart breast cancer histotypes in the different groups of breast cancer patients stratified for combined levels of YAP signature SPAG5 and miR-10b-3p expression. D Boxplots of miR-10b-3p expression in Metabric breast cancer patient dataset stratified for different combined levels of YAP signature and SPAG5 expression. E Proposed model depicting the transcriptional and post-transcriptional modulation of SPAG5 expression in breast cancer (*P value <0.05; **P value <0.001).

Fig 4: High SPAG5 expression is associated with unfavorable OS and FP in patients with LUAD. (A) High SPAG5 expression was associated with unfavorable OS. The OS curve of 360 patients was plotted [cut-off value, 266; HR (95% CI)=2.04 (1.38-3.01); log-rank P=0.00024]. (B) High SPAG5 expression was associated with unfavorable FP. The FP curve of 170 patients was plotted [cut-off value, 216; HR (95% CI)=2.6 (1.37-4.93); log-rank P=0.0023]. (C) High SPAG5 expression was not significantly associated with PPS. The PPS curve of 125 patients was plotted [cut-off value, 305; HR (95% CI)=1.49 (0.86-2.61); log-rank P=0.15]. The online tool Kaplan Meier Plotter was used to draw the graphs. OS, overall survival; FP, first progression; LUAD, lung adenocarcinoma; SPAG5, sperm-associated antigen 5; HR, hazard ratio; PPS, post-progression survival.

Fig 5: SPAG5 is overexpressed in breast cancer tissues and anticorrelated with miR-10b-3p.A, B Heatmaps SPAG5 and miR-10b-3p expression in METABRIC cohort (A) and in TCGA dataset (B). C, D Boxplot SPAG5 expression in tumoral versus normal breast tissues in METABRIC cohort (C) and in TCGA dataset (D). E, F Boxplot SPAG5 expression in Lum-A, Lum-B, Her2, and basal-like breast cancer histotypes in METABRIC cohort (E) and in the TCGA dataset (F). G, H Kaplan–Meier plots show survival analysis of breast cancer patients with low and high expression of SPAG5 in METABRIC cohort (G) and in the TCGA dataset (H).