Fig 2: Model of cisplatin-stimulated nuclear FAK protection against ERK-associated cell death in ovarian cancer. Left, Patient tumors surviving neoadjuvant chemotherapy exhibit a high level of active FAK staining in the cell nucleus. This is a tumor cell–intrinsic response, as subcytotoxic cisplatin treatment was associated with FAK nuclear accumulation in human and mouse ovarian tumor cells. Cisplatin stress also increased MKP1 tyrosine phosphatase expression that limits phosphorylation and activity of ERK. Inhibition of MKP1 results in elevated cisplatin-stimulated ERK activity associated with cell death. FAK inhibition also results in increased ERK activity, and FAK inhibition potentiated cisplatin-induced cell death (28). Right, Mutational inactivation of FAK nuclear targeting (FAK-NLS−) does not prevent basal or cisplatin-stimulated FAK-NLS− phosphorylation at Y397. FAK-NLS− does not accumulate in the nucleus upon cisplatin addition to cells. FAK-NLS− cells exhibit sensitivity to cisplatin cytotoxicity, reduced MKP1 expression, and elevated ERK activation. Our results support the notion that nuclear FAK supports MKP1 expression and that combinatorial small-molecule targeting of FAK kinase and MKP1 phosphatase activities may function as an effective therapy combination with cisplatin for HGSOC. Created in BioRender (RRID: SCR_018361). Schlaepfer, D. (2024) BioRender.com/n24i207.

Fig 3: Ninj1 deficiency attenuated IR-induced intrahepatic inflammation and neutrophil infiltration by DUSP1-dependent inhibition of p38 and JNK pathways in vivo. WT→WT and KO→WT mice were treated with either DUSP1-siRNA or NS-siRNA before the onset of liver ischemia, as described in the Materials and Methods section. The liver tissues and blood specimens were collected at 24 hours after reperfusion. (A) Western blot analysis of DUSP1, p-P38, T-p38, p-JNK, T-JNK, and β-actin expression. Relative intensity was analyzed by ImageJ software (National Institutes of Health). (B) Quantitative RT-PCR analysis of liver inflammatory cytokine and chemokine gene induction (IL6, IL1β, TNF-α, and CXCL-1). (C) Enzyme-linked immunosorbent assay (ELISA) of inflammatory cytokine and chemokine levels in serum. All results are representative of at least 2 independent experiments. (D and E) Representative immunofluorescence staining of LY6G (red) and 4’,6-diamidino-2-phenylindole dihydrochloride (DAPI) (blue) in livers from each group, quantification of LY6G+ cells per high-power field (HPF), Scale bar: 100 μm; n = 6 mice/group. ∗P < .05. All data are presented as the means ± SD. GAPDH, glyceraldehyde-3-phosphate dehydrogenase. T-p38, Total-p38; T-JNK, Total-JNK; LY6G, lymphocyte antigen 6 complex locus g.

Fig 4: Ninj1 deficiency diminished the inflammatory response in KCs through DUSP1-dependent deactivation of JNK and p38 in vitro. KCs isolated from WT and Ninj1 KO mice were stimulated with LPS for 6 hours. The cells and culture supernatant were collected. (A) Western blot analysis of MyD88 and β-actin expression. Relative intensity analyzed by ImageJ software (National Institutes of Health). (B) Western blot analysis of p-P38, T-p38, p-JNK, T-JNK, p-ERK, T-ERK, and β-actin expression. Relative intensity analyzed by ImageJ software. (C) Western blot analysis of Ninj1, p-ASK1, p-TAK1, DUSP1, and β-actin expression. Relative intensity analyzed by ImageJ software. WT and KO KCs were transfected transiently with DUSP1-siRNA or NS-siRNA followed by LPS stimulation for 6 hours. The cells and culture supernatant were collected. (D) Western blot analysis of DUSP1, p-P38, T-p38, p-JNK, T-JNK, and β-actin expression. Relative intensity analyzed by ImageJ software. (E) Quantitative RT-PCR analysis of KC inflammatory cytokine and chemokine gene induction (IL6, IL1β, TNF-α, and CXCL-1). (F) Enzyme-linked immunosorbent assay of inflammatory cytokine and chemokine levels in cell supernatant. All results are representative of at least 2 independent experiments. ∗P < .05. All data are presented as the means ± SD. ELISA, enzyme-linked immunosorbent assay; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; PBS, phosphate-buffered saline. T-p38, Total-p38; T-JNK, Total-JNK; T-ERK, Total ERK.

Fig 5: The interaction between TRIM11 and DUSP1 in AC16 cells (a, b). After injection with the oeTRIM11 lentivirus, the mRNA (a) and protein (b) expressions of TRIM11 and DUSP1 were detected. ***p < .001 versus vector. (c) Co‐immunoprecipitation assay of the interaction between TRIM11 and DUSP1. (d) DUSP1 ubiquitination assay. (e) After oeTRIM11 or MG132 (10 µmol/L) treatment, the protein levels of TRIM11 and DUSP1 were detected