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. 2017 Jul 31;127(9):3527–3542. doi: 10.1172/JCI93488

Figure 7. Mutant GATA4 V267M does not recruit the mediator complex and is transcriptionally less active.

Figure 7

(A) Impaired activation of HNF4A and CEBPD by GATA4 V267M versus GATA4. Cells transiently transfected with expression vector for GATA4, GATA4 V267M, or empty vector control. QRT-PCR, relative to nontransfected cells. (B) Western blot for GATA4, c-MYC, p27/CDKN1B, and actin. (C) Cell proliferation. (D) GATA4 and GATA4 V267M binding to GATA response elements. PLC HCC cells transfected with expression vectors for GATA4 or GATA4 V267M. Nuclear lysates incubated with DNA probes containing GATA4 response elements X3-biotin tag or with control scrambled probes. Bound protein detected by Western blot after streptavidin pull-down. (E) Mediator was absent from the GATA4 V267M interactome. LC-MS/MS analysis of protein interactome of GATA4 and GATA4 V267M immunoprecipitated from transfected PLC cells using anti-Flag antibody. Circle size indicates protein abundance in the coimmunoprecipitate (Supplemental Figure 10). (F) Confirmation by immunoprecipitation/Western blot of absence of mediator, but not other coactivators. Triplicate results are shown in Supplemental Figure 11.