Fig 2: Knockdown validation of miR-142-3p targets.qPCR expression analysis of Cfl2, Wasl, and Grlf1 genes in NIH-3T3 fibroblasts that were transduced with either lentivrial vectors encoding shRNAs directed against miR-142-3p targets (shCfl2, shWasl, shGrlf1; grey bars), or with lentivirus harboring control shRNA directed against RFP (shRFP; black bars). Data are presented as mean + SEM, and normalized to Hprt expression and to the mRNA expression in shRFP-transduced cells. **p<0.005; ***p<0.0005.DOI: http://dx.doi.org/10.7554/eLife.01964.020

Fig 3: Cell migration assay. (A) SH-SY5Y cells either transfected with Nox4 or scrambled siRNA were grown to 80% confluency in a 24-well plate containing a hydrogel spot (time point 0 h). After removing of the spot the cells transfected with scrambled siRNA but not the Nox4 siRNA started to migrate into the cell free area (time point 24 h). (B) The remaining free area in the open spots was determined as described in the Section “Materials and Methods.” Results are mean ± SD and were marked for significance as described in the Section “Materials and Methods.” (C) The western blot for N-WASP normalized to GAPDH indicates downregulation of N-WASP when Nox4 expression was suppressed in three independent homogenates from hepatoma cells. Equal amounts of protein were loaded onto the SDS-PAGE.

Fig 4: Model of C. jejuni internalization.C. jejuni invasion of host cells. Step 1: C. jejuni binds to fibronectin (Fn) via the two C. jejuni Fn binding proteins CadF (blue dots) and FlpA (yellow dots) [43,44] causing activation of the α5β1 integrin receptors and the epidermal growth factor receptor (EGFR) [27,45]. Step 2: Activation of the α5β1 integrin leads to the recruitment and partial activation of FAK and paxillin [27,46]. Step 3: The delivery of the Campylobacter invasion antigens (e.g., CiaD shown in red) to the host cell [16,19,27,47] leads to the maximal activation of key components of the focal complex (i.e., FAK, paxillin, vinculin, p130Cas, Src, and the CrkII/DOCK-180/ELMO complex) [27,28,45, Konkel et. al, Invasion of epithelial cells by Campylobacter jejuni is independent of caveolin-1, In Submission]. Step 4: Focal complex activation, in conjunction with CiaD, leads to the phosphorylation of Erk 1/2. Caveolin-1, Vav2, Rac1, and Cdc42 are also activated following focal complex activation [27,28,45]. Step 5: Activation of Erk 1/2 and Src leads to the phosphorylation of cortactin, which allows for the Rho GTPases Rac1 and Cdc42 to activate N-WASP associated with phosphorylated cortactin, promoting actin cytoskeletal reorganization. Highlighted in this model is the role of CiaD in C. jejuni internalization. Specifically, CiaD is necessary for the maximal activation of the Erk 1/2 and cortactin signaling pathways. Components of the focal complex and focal complex associated proteins are shown in blue. The newly identified components of the C. jejuni invasion complex are shown in green.

Fig 5: Numb and Numblike regulate Z-disc assembly and integrity through ACTN.(A) Double immunofluorescence staining of Numb and ACTN in the CMs of E14.5 CTL and MLC-2v-cre DKO mice after 72-hour culture in vitro (ACTN, green; Numb, red; original magnification, ×630, zoom in ×2; amplified images: original magnification, ×630, zoom in ×5). (B) Double immunofluorescence staining of Numb and ACTN in the SKMs of CKMM-cre DKO mice on P60 (ACTN, green; Numb, red; original magnification, ×630, zoom in ×2). (C) Coimmunoprecipitation of N-WASP with other ABPs in heart and SKM lysates of CTL and CKMM-cre DKO mice on P60. (D and E) Relative protein level of ABPs immunoprecipitated by N-WASP in heart and SKM lysates of CTL and CKMM-cre DKO mice on P60 (n = 3). (F) Coimmunoprecipitation of α-Actin (ACTC1) with ACTN in heart lysates of CTL and α-MHC-cre DKO mice on P60. (G) Relative ACTN protein level immunoprecipitated by ACTC1 in heart lysates of CTL and α-MHC-Cre DKO mice on P60 (n = 4). Scale bars: 5 μm. Two-tailed Student’s t test with error bars showing mean ± SD.