Extended Data Fig. 6. Senescent cells have a reduced capacity to form aggresomes.
(a) Bright-field image of cycling and senescent HFF-1 cells that were subjected to a senescence associated β-galactosidase (SA-β-gal) assay. The percentage of cells displaying SA-β-gal activity is indicated. Scale bar 10 μm. (b) Extracts from cycling and senescent HFF-1 cells treated with or without MG132 were probed for CP110, CEP290, Ki67, p53, phospho-p38, p38, p21, pHSP27 and total HSP27, as indicated. α-tubulin was used as a loading control. (c) Cycling and senescent HFF-1 cells were treated with DMSO or MG132, then stained for Ub+ and CP110. DNA was stained with DAPI. Scale bar 10 μm. (d) crFAM83 and crPCM1 IMR-90 cells were stained for PCM1. Scale bar 10 μm. (e) Bright-field image of crFAM83G and crPCM1 IMR-90 cells that were subjected to a senescence associated β-galactosidase (SA-β-gal) assay. Scale bar 20 μm (f) Quantification of the percentage of senescent cells as determined by SA-β-gal activity in crFAM83 and crPCM1 IMR-90 cells, n = 2 biological replicates. (g) Senescent HFF-1 cells were transfected with GFP-CP110 for 48 hours before treatment with MG132. Cells were stained as indicated. Scale bar 10 μm, inset 2 μm. (h) Histogram of the percentage of senescent HFF-1 cells transfected with GFP or GFP-CP110 that formed an aggresome. Data displayed as mean and range. n = 2 independent experiments. (i) Senescent HFF-1 cells were transfected with GFP-CP110, treated with MG132 and stained for CEP97, CEP290 and PCM1. Scale bar 10 μm, inset 2 μm. Unprocessed immunoblots and numerical data are provided as source data.