Eplin-α expression is regulated by signalling through G-actin. (A) Four independent Affymetrix probe sets of the Lima1 gene encoding Eplin were differentially regulated by actin binding drugs. G-actin regulated genes were induced by treatment with cytochalasin D (CD, 2 μM, 90 min) and repressed by latrunculin B (LB, 5 μM). Results shown are from transcriptome analysis of NIH 3T3 fibroblasts as previously described . The q-value is the lowest false discovery rate at which the differentially expressed probe set is called significant. (B) Validation of differential regulation of Eplin-α, but not of Eplin-β, by actin binding drugs. NIH 3T3 cells were treated with cytochalasin D (2 μM) for 120 min, or with cytochalasin following 30 min pretreatment with latrunculin B (5 μM). Controls were left untreated (un.). The total mRNA was isolated and subjected to quantitative RT-PCR as described . Shown is the average induction of Eplin mRNA after normalisation to hprt. Error bars indicate SEM (n = 3) for Eplin-α, and half range for Eplin-β. (C, D) Effect of pretreatment with latrunculin B (C) or UO126 (10 μM, 30 min) on the average induction of Eplin-α mRNA by serum (FCS, 15%, 90 min). Error bars indicate SEM of at least three independent experiments. The used primers were (positions of mRNA): Eplin-α, → (1203GCTGTTTCCGATGCTCCTAC1223), ← (1382CTCATTGTCGCTCTTGCT TG1362); Eplin-β, → (183CAAGAACAAGTCATCCGCAAT204), ← (418AGGAGGGTAGTCCGCTGTGT398). Asterisk, significant activation; double asterisk, significant repression (p < 0.01, unpaired student's t-test).