Fig. 4: Contributions of IAP RNA to condensate localization in vivo and condensate formation in vitro.
From: Hijacking of transcriptional condensates by endogenous retroviruses
a, Schematic model of the ERV shRNA knockdown experiments. b, qRT–PCR data as FC-normalized to the DMSO treatment control. Data are presented as mean values ± s.d. from three biological replicates. P values are from two-tailed t-tests. ****P < 1 × 10−4, ***P < 1 × 10−3, **P < 1 × 10−2. c, Log2 FC values in total RNA-seq data at intergenic SEs and genes. Data are from three biological replicates. P values are from two-sided Wilcoxon–Mann–Whitney tests. ****P < 1 × 10−4. d, Representative images of individual z-slices (same z) of RNA-FISH and IF signal, and an image of the merged channels. The nuclear periphery determined by DAPI staining is highlighted as a white contour (scale bars, 2.5 μm). Also shown are averaged signals of either RNA-FISH or RNAPII IF centered on the miR290-295 FISH foci or randomly selected nuclear positions (scale bars, 0.5 μm). r denotes a Spearman’s correlation coefficient. e, Representative images of mixtures of fluorescein-labeled IAP RNA and purified recombinant RNAPII CTD-mCherry in droplet formation buffer. Scale bar, 5 μm. f, Partitioning ratio of RNAPII CTD-mCherry into droplets at the indicated IAP RNA concentrations. Every dot represents a detected droplet. P values are from two-sided t-tests. g, Quantification of the enrichment of fluorescein-labeled IAP RNA in RNAPII CTD-mCherry droplets. P values are from two-sided t-tests. h, Quantification of the partitioning of (left) MED1 IDR and (right) HPIα into droplets in the presence of IAP RNA. Values are normalized against the partition ratio at no RNA added. Corresponding images are found in Extended Data Fig. 8a. The displayed quantification is the same as displayed in Extended Data Fig. 8b. i, Representative images of droplet formation by purified NFYC-IDR-mEGFP (1 μM) and MED1 IDR-mCherry (5 μM) fusion proteins in the presence of in vitro transcribed Cy5-labeled IAP RNA fragment. Scale bar, 5 μm. j, Partitioning ratio of NFYC-IDR-mEGFP, MED1 IDR-mCherry and IAP RNA into droplets at the indicated IAP RNA concentrations. Every dot represents a detected droplet. All pairwise P values <2.2 × 10−16 (Welch’s t-test). k, Schematic model of the experiment mimicking IAPEz transcription. l, qRT–PCR data as FC-normalized to the DMSO control treatment. n.d., not detectable. Data are presented as mean values ± s.d. P values are from two-tailed t-tests. ****P < 1 × 10−4. m, qRT–PCR data as FC-normalized to the DMSO control treatment. Data are presented as mean values ± s.d. from three biological replicates. P values are from two-tailed t-tests. ****P < 1 × 10−4, ***P < 1 × 10−3, **P < 1 × 10−2, NS, not significant. In panels f, g, h and j, data for quantification were acquired from at least five images of two independent image series per condition.
