Extended Data Fig. 1: Mini-PSs are present within NSs, although NEAT1 transcription occurs outside of NSs. | Nature Cell Biology

Extended Data Fig. 1: Mini-PSs are present within NSs, although NEAT1 transcription occurs outside of NSs.

From: Shell protein composition specified by the lncRNA NEAT1 domains dictates the formation of paraspeckles as distinct membraneless organelles

Extended Data Fig. 1

a, Schematics of the NEAT1_2 configuration in the WT-PS and mini-PS. b, SRM images of PSs (RNA-FISH with NEAT1_5′ probe) and NSs (SRRM2 IF) in WT and mini-NEAT1 mutant cells in the absence of MG132. Scale bar, 500 nm. c, Graph showing the proportion of segregated or incorporated PSs in b. Data were collected from two independent experiments. WT: n = 167, mini-NEAT1: n = 210. d, SRM images of the PSs (RNA-FISH with NEAT1_5′ probe) and NSs (SRRM2 IF) in WT and another mini-NEAT1 mutant clone treated with MG132 (5 μM for 6 h). Scale bar, 10 μm. e, PSs and NSs were detected by smFISH with NEAT1_5′ probes (magenta) and SON IF (green) in the presence of MG132 (5 μM for 6 h). Nuclei were stained with DAPI. Line profiles of the PSs and NSs are shown (right). f, SRM images of PSs (smFISH with NEAT1_5′ probe) and NSs (SON IF) in WT and mini-NEAT1 cells treated with MG132 (5 μM for 6 h). Scale bar, 500 nm. g, PSs and NSs in WT and mini-NEAT1 cells were detected by RNA-FISH with NEAT1_5′ probe and MALAT1 probe (green and magenta, respectively) in the presence of MG132 (5 μM for 6 h). Nuclei were stained with DAPI. Line profiles of the PSs and NSs are shown (right). Scale bar, 10 μm. h, SRM images of PSs (RNA-FISH with NEAT1_5′ probe) and NSs (RNA FISH with MALAT1 probe) in WT and mini-NEAT1 cells treated with MG132 (5 μM for 6 h). Scale bar, 500 nm. i, EM observation of multiple, but unconnected, mini-PSs within an NS. NONO localization was detected as gold particles. Dotted magenta circles indicate the mini-PS position. Dotted black circles indicate the NS position. Scale bar, 500 nm. j, PSs and other nuclear bodies in WT and mini-NEAT1 cells treated with MG132 (5 μM for 6 h). PSs were visualized by NEAT1 RNA-FISH with NEAT1_5′ probe (green). Nuclear bodies (parentheses) were visualized by IF of marker proteins (magenta). Nuclei were stained with DAPI. Scale bar, 10 μm. Source numerical data are available in source data.

Source data

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