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. 2024 Feb 15;11(16):2306915. doi: 10.1002/advs.202306915

Figure 4.

Figure 4

Circ0006646 inhibited the combination of TRIM21 and NCL to stabilize the expression of NCL. A) Cells were transfected with siNC and si‐circ0006646. Blots with antibodies recognizing the NCL and β‐Actin were shown. B) Representative IF images of NCL (green) were shown. DAPI staining represented the location of the nucleus. C) Cells were transfected with siNC and si‐circ0006646, and later treated with CHX for the indicated times. Blots with antibodies recognizing the NCL and β‐Actin were shown. D) Broken line graph represents the change of NCL expression at 0, 3, and 6 h after treatment of CHX. E) Cells were transfected with siNC and si‐circ0006646, and later treated with 10 µm MG132 for 6 h. Blots with antibodies recognizing the NCL and β‐Actin were shown. F,G) The HCCLM3 lysate was incubated with anti‐NCL and anti‐IgG, and later incubated with protein A/G magnetic beads. Blots with antibodies recognizing the UB‐K48 (F) or UB‐K63 (G) and NCL were shown. H) 293T cells transfected with HA‐tagged UB‐K48, HA‐tagged UB‐K63, siNC, and si‐circ0006646 were treated with 10 µm MG132 for 6 h. The 293T lysate was incubated with anti‐NCL, and later incubated with protein A/G magnetic beads. Blots with antibodies recognizing the HA and NCL were shown. I) The HCCLM3 lysate was incubated with anti‐NCL and anti‐IgG, and later incubated with protein A/G magnetic beads. Blots with antibodies recognizing the TRIM21 and NCL were shown. j) Representative IF images of NCL (green) and TRIM21 (red) were shown. DAPI staining represented the location of the nucleus. K) 293T cells transfected with HA‐tagged UB and MYC‐tagged TRIM21 were treated with 10 µm MG132 for 6 h. The 293T lysate was incubated with anti‐NCL, and later incubated with protein A/G magnetic beads. Blots with antibodies recognizing the HA and NCL were shown. L) 293T cells transfected with MYC‐tagged TRIM21, siNC, and si‐circ0006646 were treated with 10 µm MG132 for 6 h. The 293T lysate was incubated with anti‐NCL, and later incubated with protein A/G magnetic beads. Blots with antibodies recognizing the MYC and NCL were shown. All experiments were performed in at least triplicate samples. Data were presented as the means ± SD.