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. 2022 Sep 11;56:102469. doi: 10.1016/j.redox.2022.102469

Fig. 9.

Fig. 9

Interaction with AKT is required for MPST function in IBD in vitro. (A) HEK293T cells infected with a Flag-tagged MPST overexpression plasmid and an HA-tagged AKT overexpression plasmid (or the empty vector) for 48 h were collected, and total Flag-tagged MPST was immunoprecipitated using anti-Flag beads. (B) Interaction domains of AKT and MPST were explored using full-length and truncated AKT based on co-immunoprecipitation assays in HEK293T cells. (C) HEK293T cells infected with a Flag-tagged MPST (or Flag-tagged mut MPST) overexpression plasmid and an HA-tagged AKT overexpression plasmid (or the empty vector) for 48 h were collected, and total Flag was immunoprecipitated using anti-Flag beads. (D) Co-IP analyses showing the interaction between recombinant His-MPST and recombinant GST-AKT. (E) The protein levels of phosphorylation-related proteins (PI3K and PTEN) and the dephosphorylation enzyme (PP2A) were measured in MPST-deficient HT-29 cells by WB. (F) MPST-deficient HT29 cells were stimulated with 100 ng/mL TNF-α with or without LB100 (1 μM) for 24 h, and the protein level of p-AKT was measured by WB. (G) Representative immunofluorescent images of DAPI, MPST, AKT and the merge of all staining in HT29 cells.