Fig 2: High expression of S100A1 in PTC might be a new biomarker. (A): Immunohistochemistry (IHC) staining of S100A1 was shown in human PTC and normal thyroid tissues. (B): IHC staining score of S100A1 was compared in human PTC and normal thyroid tissues (***P<0.001). (C-D): S100A1 expression was compared in paired samples and population samples in TCGA database, respectively. (*P<0.05). (E): Comparison of the expression of S100A1 in normal thyroid and various PTC subtypes in TCGA. (F): Methylation levels of S100A1 gene promoter in thyroid cancer were lower than normal thyroid based on TCGA data, which were analyzed using UALCAN web. (*P<0.05). (G): Kaplan-Meier analysis showed that a higher expression of S100A1 was associated with a worse recurrence-free survival. (H): Survival analysis was shown based on our follow-up data of 86 patients. (I): ROC curves were shown for discrimination of normal tissues and PTC tissues. An optimal cut-point derived from the maximum of Youden index was displayed as a triangle (Youden index: 0.773; Sensitivity: 82.46%, Specificity: 94.83%). ROC curves for diagnosis in stage I, stage II, stage III, and stage IV patients. PTC, papillary thyroid carcinoma; TCGA, The Cancer Genome Atlas; ROC, receiver operating characteristic; AUC, area under the curve.

Fig 3: S100A1 regulates the tumorigenicity of PTC cells in vivo and the underlying molecular mechanism. (A): Constructed S100A1 was stably silenced in K1 cells via lentivirus (***P<0.001). (B): Mouse xenograft models and tumor tissues were measured 24 days after subcutaneous injection with K1-shNC cells or K1-sh1 cells. Tumor volumes were examined every three days for 28 days (*P<0.05). (C): IHC staining of S100A1 protein was shown in nude mice transplanted tumors. (D): Western blot assays of the nuclear and cytoplasmic YAP/P-YAP protein were shown. PTC: papillary thyroid carcinoma; IHC: immunohistochemistry; YAP: yes associated protein; P-YAP: phosphorylated YAP.

Fig 4: Knockdown of S100A1 inhibited cell migration and enhanced apoptosis. (A-B): Numbers of migrating cells in the Si-1 groups were less than in the NC group, and the cell numbers were analyzed for either K1 or BCPAP cell lines (*P<0.05, **P<0.01). (C-D): Apoptosis percentage in the Si-1 group was higher than the NC group, and the cell apoptosis rate was analyzed for either K1 or BCPAP cell lines (**P<0.01). NC: negative control.

Fig 5: Knockdown of S100A1 inhibited cell proliferation. (A) Differential expression of S100A1 protein was compared between human PTC tissues and paired thyroid tissues. GAPDH: Glyceraldehyde 3-phosphate dehydrogenase. (B): The S100A1 mRNA expression was shown in four cell lines (Nthy-ori-3-1, BCPAP, TPC-1, and K1) (***P<0.001). (C): The protein expression of S100A1 in four cell lines was identified by Western blot assays. GAPDH: Glyceraldehyde 3-phosphate dehydrogenase. (D-E): The mRNA and protein expression of S100A1 were compared among three Si, the NC and the blank groups. Si-1 groups showed the lowest S100A1 protein expression in either K1 or BCPAP cell lines (***P<0.001). (F-G): Compared with the NC or blank groups, Si-1 groups showed the lowest growth rate both in either K1 or BCPAP cell lines (*P<0.05). OD value at 450 nm was measured by CCK-8 assay. (H-I): Compared to the NC group, the number of cell clones significantly decreased in Si-1 groups (*P<0.05, ***P<0.001). NC: negative control; OD: optical density.