Fig. 4: Arginine mediated nuclear retention of TEAD4.

a Cells were first incubated in arginine-free media and harvested at different time points as indicated. Fresh arginine was then added into media and cells were harvested at different time points as indicated. Western blotting shows that arginine (Arg) did not affect TEAD4 protein level, but affected the ratio of nucleus and cytosol fractions (b). c Cells were first incubated in arginine-free media for 24 h. Arginine was then added into media for 24 h. Immunofluorescent staining of TEAD4 shows cytosolic translocation after arginine deprivation (scale bar = 20 μm). The quantification of immunofluorescent staining is shown in d. e Cells were incubated in arginine-free media and harvested at different time points as indicated. Western blotting shows that arginine depletion induced p38 phosphorylation in a time-dependent manner, which is consistent with TEAD4 cytosol translocation (Supplementary Fig. 3a). f Cells were incubated in arginine-free media with or without p38 inhibitor (p38i) SB203580 treatment for 24 h. Western blotting shows that inhibition of p38 activation prevented TEAD4 cytosolic translocation upon arginine deprivation. The total TEAD4 expression is shown in Supplementary Fig. 3b. g Immunofluorescent staining of TEAD4 shows that inhibition of p38 activation impeded TEAD4 cytosolic translocation upon arginine deprivation (scale bar = 20 μm). The quantification of immunofluorescent staining is shown in (h). i After starvation overnight, cells were treated with arginine for 24 h and then harvested for immunoblotting. Western blotting shows that arginine did not affect the phosphorylations of YAP1. j YAP1 was knocked down via shRNA lentivirus infection for 24 h, following an antibiotic selection for 1 week. ChIP-qPCR data show that silencing of YAP1 did not significantly affect the binding activity of TEAD4 on OXPHOS promoter region. k Cells were transduced with two individual YAP1 shRNAs for 24 h, following an antibiotic selection for 1 week. Seahorse assay for mitochondrial respiration activities after silencing of YAP1 via two individual shRNA clones. O oligomycin, F FCCP, R/A rotenone/antimycin. l ChIP-qPCR data show the binding activity of TEAD4, YAP1, PGC1a, and NRF1 on OXPHOS promoters. m ChIP-qPCR data show that the presence (+) or absence (−) of arginine regulates the recruitment of TEAD4 and PGC1a on OXPHOS promoters. Data are presented as mean value ± SEM of independent experiments (n = 3 in d, h, j, l, m; n = 4 in k). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, ns not significant, using unpaired two-tailed Student’s t test. Source data are provided as a Source data file.