Additional file 1 of Deubiquitinase USP10 promotes osteosarcoma autophagy and progression through regulating GSK3β-ULK1 axis

Supplementary material 1: Figure S1.The expression levels of USP10 in 143B and U2OS cells were evaluated by western blotting and q-RT-PCR following USP10 knockdown or overexpression.USP10 expression was downregulated using shRNAs in HOS and MG-63 cells. Subsequently, protein levels of LC3B-II, p62, ULK1, and p-ULK1were analyzed by western blotting.Upon USP10 overexpression in 143B and U2OS cells, the protein levels of LC3B-II, p62, ULK1, and p-ULK1were determined via western blotting.ULK1 was downregulated specifically in 143B cells. Subsequently, protein levels of USP10 were analyzed using western blotting.USP10 was downregulated or overexpressed specifically in 143B cells. Subsequently, protein levels of ATG13 were analyzed using western blotting.The process of autophagosome maturation was examined through immunofluorescence analysis in RFP-GFP-LC3 cells. These cells were stably transfected with either shUSP10 or shNC and subsequently cultured in EBSS for a duration of 4 h. Scale bars:10 μm. Quantifying RFP-LC3-only puncta and RFP-GFP overlay puncta in indicated cells.USP10 knockdown in HOS and MG-63 cells or USP10 overexpression in 143B cells and U2OS cells, cell growth was examined through colony formation.USP10 knockdown in HOS and MG-63 cells or USP10 overexpression in 143B cells and U2OS cells, cell invasion was detected using a transwell assay