Figure 4. PDZ-RhoGEF is required for the optimal IGF-1 signaling output in MEFs and adipogenic progenitor cells.
(A) RhoA activation was determined by the RBD-binding assay in response to the indicated factors (FCS [10%], IGF-1 [100 ng/ml], LPA [20 μM]). (B) Differential PKB/Akt phosphorylation in WT and PDZ-RhoGEF KO MEFs in response to IGF-1. (C) Differential PKB/Akt phosphorylation and PDZ-RhoGEF expression in WT and PDZ-RhoGEF KO adipogenic progenitor cells in response to IGF-1. (D) IGF-1 response of PDZ-RhoGEF KO MEFs is rescued by ectopic expression of myc-tagged PDZ-RhoGEF. (E) IGF-1 stimulated tyrosine phosphorylation of IR. (F) Serine phosphorylation of IRS1 and interaction between IRS1 and PI3K in WT and PDZ-RhoGEF KO MEFs in response to IGF-1. (G) Phosphorylation of PKB/Akt S473 in response to IGF-1 is dependent on ROCK activity.
Figure 4—figure supplement 1. Stress fiber formation and mitogen-induced cell migration.
(A) Stress fiber formation 15 min after mitogen stimulation (FCS [10%], IGF-1 [100 ng/ml], LPA [20 μM]) was determined by phaloidin-rhodamine staining (B) Twenty-four hours after mitogen stimulation (as in [A]), cell migration of MEFs with the indicated genotypes was determined by a wound-healing assay. (C) Six hours after plating, IGF-1-induced migration of MEFs with the indicated genotypes was determined by the transwell migration assay.