Loss of VGLL4 suppresses tumor PD‐L1 expression and immune evasion

A, B

Correlation between protein expression of VGLL4 and IRF2BP2 in six murine tumor cell lines (A) and human cell lines (B) with the indicated antibodies.

C

Immunoblot analysis of A549 cells transfected with VGLL4‐FLAG or VGLL4‐HF4A plasmids revealed increased IRF2BP2 protein levels compared with control cells.

D

Deficiency of VGLL4 reduces IRF2BP2 protein levels. Immunoblot analysis of A549 and LLC cell lysates transfected with siRNA for control and VGLL4.

E

The indicated cell lines were transfected with control or VGLL4 siRNA and subjected to immunoblot analysis with the indicated antibodies.

F

qRT–PCR analysis of IRF2BP2 mRNA levels in control and siVGLL4 A549 cells. n = 3, mean ± SEM.

G

A549 or 16HBE cells were transfected with control or VGLL4 siRNA and treated with MG132 for the indicated times and subjected to immunoblot analysis with the indicated antibodies. Relative IRF2BP2 expression levels were quantified in the right panel. n = 3, mean ± SEM.

H

Lys48‐linked ubiquitylation of IRF2BP2 is strongly inhibited when VGLL4 was overexpressed in HEK293T cells. HEK293T cells were transfected with indicated plasmids, treated with MG132 for 3 h before harvest, and subjected to immunoprecipitation and immunoblot analysis as indicated.

I

Lys48‐linked ubiquitylation of IRF2BP2 is increased in the absence of VGLL4 in HEK293T cells. HEK293T cells were transfected with indicated siRNA and plasmids, and subjected to immunoprecipitation and immunoblot analysis as indicated.

Figure 4. VGLL4 protects proteasome‐mediated IRF2BP2 degradation.