Figure 4 from Hypoxia-Induced Long Noncoding RNA HIF1A-AS2 Regulates Stability of MHC Class I Protein in Head and Neck Cancer

HIF1A-AS2 downregulates MHC-I through autophagic degradation. A, Western blots showing the expression of HLA-ABC, STAT1 in SAS cells overexpressing HIF1A-AS2 (SAS-HIF1A-AS2) versus SAS-control (SAS-Vec) treated with/without IFNγ. α-Tubulin was used as a loading control. B, A pulse-chase assay for showing the protein degradation of HLA-ABC, HLA-A, and HLA-C in SAS-HIF1A-AS2 versus SAS-Vec treated with cycloheximide to inhibit de novo protein synthesis at different time points. α-Tubulin was used as a loading control. The bottom panel is the quantification of HLA-ABC. C, Western of HLA-ABC, HLA-A, HLA-B, HLA-C, Snail, and LC3B in SAS-HIF1A-AS2 treated with the autophagic inhibitor bafilomycin A1 (Baf.), proteasome inhibitor MG132, and rapamycin (Rap.). Snail is a positive control of proteasome-degraded protein, and LC3B is the control of autophagy-degraded protein. α-Tubulin was used as a loading control. D, Western blot of HLA-ABC and LC3B in SAS-HIF1A-AS2 (left) and HSC3-HIF1A-AS2 (right) treated with/without bafilomycin A1(Baf.) for 12 hours. α-Tubulin was used as a loading control. E, Western blot of HLA-ABC, HLA-C, Snail, and LC3B in SAS overexpression HIF1A-AS2 (SAS-HIF1A-AS2; left) and HSC3 overexpression HIF1A-AS2 (HSC3-HIF1A-AS2; right) treated with/without MG132 for 12 hours. α-Tubulin was used as a loading control.