Fig. 6: RIF1-L interacts with BRCA1 potentially at broken replication forks. | Nature Communications

Fig. 6: RIF1-L interacts with BRCA1 potentially at broken replication forks.

From: The human RIF1-Long isoform interacts with BRCA1 to promote recombinational fork repair under DNA replication stress

Fig. 6

A Representative images of GFP-RIF1-L or GFP-RIF1-L-pp1bs signal in HeLa cells, with indicated treatments. unt: not treated with HU; HU: 4 mM, 24 h. Scalebar: 10 µm. B Quantification of GFP-RIF1-L or GFP-RIF1-L-pp1bs foci in cells from the experiment as (A). Means and standard errors of three independent experiments are plotted. p values (two-tailed) calculated by Student’s t test. C Left: Representative images of GFP-RIF1 fluorescence and BRCA1 immunofluorescence in HeLa RIF1 KO, +GFP-RIF1-L and +GFP-RIF1-L-pp1bs cells. All samples were treated with 4 mM 24 h HU. Scalebar: 10 µm. Right: BRCA1 nuclear signal intensity fold change. Median intensity values from three independent experiments were recorded (see Supplementary Fig. 6B for one representative experiment). Fold changes were determined by normalising to values of the RIF1 KO sample. Means and standard errors were shown. p values (two-tailed) calculated by one-sample t test. Source data are provided as a Source Data file. D Left: Representative images of BRCA1 immunofluorescence in HeLa cells treated with siCtrl or siRIF1. All samples were treated with 4 mM 24 h HU. Scalebar: 10 µm. Right: BRCA1 nuclear signal intensity fold change (normalised to siCtrl cells). Data plotted as described in (C). p values (two-tailed) calculated by one-sample t test. E An example of co-localisation between GFP-RIF1-L-pp1bs, BRCA1 and γH2AX in HeLa cells. This sample was treated with 4 mM 24 h HU. BRCA1 and γH2AX signals were generated by immunostaining. Scalebar: 5 µm. F Quantification of co-localisation between GFP-RIF1-L-pp1bs, BRCA1 and γH2AX in cells from the experiment as (E). Means and standard errors of four independent experiments are plotted. G An example of co-localisation between GFP-RIF1-L-pp1bs, BRCA1 and RAD51 in HeLa cells. This sample was treated with 4 mM 24 h HU. BRCA1 and RAD51 signals were generated by immunostaining. Scalebar: 5 µm. H Quantification of co-localisation between GFP-RIF1-L-pp1bs, BRCA1 and RAD51 in cells from the experiment as (G). Means and standard errors of three independent experiments are plotted.

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