Fig 2: The mechanism for the AT1R inhibition of anti-senescence through mitochondrial quality control. ox-LDL induced mitochondrial fission through the association of LOX-1 and AT1R and the CRAF/MEK/ERK pathway, resulting in mitochondrial dysfunction and senescence. AT1R inhibition attenuated senescence by suppression of this cascade and induced Rab9-dependent alternative autophagy through the CRAF/MEK axis and aspects of the mitochondrial quality control process in human VSMCs.

Fig 3: FAM83D promotes activation of the oncogenic pathways. (A) The protein levels of FAM83D, EGFR, p‐EGFR and c‐Raf, p‐c‐Raf, ERK1/2, p‐ERK1/2, AKT and p‐AKT in the NM‐vector and NM‐FAM83D cells were quantified using Western blotting. (B) The protein levels of MMP2, N‐cadherin and ZO1 in the NM‐vector and NM‐FAM83D cells were quantified using Western blotting. (C) Schematic diagram summarizing the molecular mechanism through which FAM83D enhanced the proliferation and migration of ovarian cancer cells. EGFR, epidermal growth factor receptor, NM, non‐metastatic; MMP2, matrix metalloproteinase‐2

Fig 4: USP15 regulates CRAF expression levels. A, samples shown in Fig. 4A were probed with the indicated antibodies. Quantitation shows the mean of three biological replicates (error bars, S.E.; two-tailed paired t test compared with control; USP15-1, p < 0.05; USP15-17, p < 0.025). IB, immunoblot. B, CRAF was immunoprecipitated (IP) from lysates shown in Fig. 4B and probed with anti-ubiquitin antibodies (Ab). C, samples from epomoxicin (Epo) and vehicle-treated cells (Fig. 4C) were probed with anti-CRAF antibodies. D, HeLa cells were treated with control reagents or USP15 siRNA (Oligo #17) for 66 h before cycloheximide (CHX; 100 μg/ml) treatment. Cells were lysed at indicated time points in Nonidet P-40 lysis buffer, and lysates were probed with anti-CRAF antibody. E, HEK293T cells were treated with control reagents or USP15 siRNA (Oligo#1) for 24 h before transfection with myc-CRAF or an empty vector for another 48 h. Cell lysates were probed with anti-USP15 and anti-CRAF antibodies. Endogenous CRAF levels are only visible in the darker exposure.

Fig 5: Perturbation of the RSK2‐mediated NFB increases the efficiency of RAS, MEK, and ERK targeting drugs Schematic representation of the optoFGFR (RSK2‐mediated feedback dependent) and optoSOS (RSK2‐mediated feedback independent) systems targeted with the B/CRAF (RAF709), the MEK (U0126) or the ERK (SCH772984) inhibitor.Single‐cell ERK amplitudes from sustained high optoFGFR input (D = 18 mJ/cm2) or optoSOS input (D = 0.6 J/cm2) under different concentrations of the MAPK inhibitors, extracted at a fixed time point (t fixed optoFGFR = 15 min, t fixed optoSOS = 10 min, N = 200 cells with high optoFGFR or optoSOS expression per condition randomly selected from 3 technical replicates).A Hill function was fit to the normalized mean ERK activity as shown in (B) (N min = 200 cells per condition). Shaded area indicates the 95% CI and dashed lines the EC50.Normalized standard deviation of ERK amplitudes shown in (B) (N min = 200 cells per condition).Schematic representation of the optoFGFR system treated with CTRL KD (RSK2‐mediated feedback dependent) or RSK2 KD (RSK2‐mediated feedback independent) targeted with the B/CRAF (RAF709), the MEK (U0126) or the ERK (SCH772984) inhibitor.Single‐cell ERK amplitudes from sustained high optoFGFR input (D = 18 mJ/cm2) under different concentrations of the MAPK inhibitors, extracted at a fixed time point (t fixed optoFGFR = 15 min, N = 70 cells with high optoFGFR expression per condition (apart from RSK2 KD + 0 μM U0126 (32 cells)), randomly selected from 2 technical replicates for RSK2 KD and 1 replicate for CTRL KD).A Hill function was fit to the normalized mean ERK activity as shown in (F) (N min = 32 cells per perturbation). Shaded area indicates the 95% CI and dashed lines the EC50.Normalized standard deviation of ERK amplitudes shown in (F) (N min = 32 cells per perturbation). Data information: in (B) and (F), boxes indicate the upper and lower quartiles, the central bands indicate the median and whiskers extend to individuals up to 1.5 interquartile away from the median.