Paraspeckles: nuclear bodies built on long noncoding RNA

Abstract

Paraspeckles are ribonucleoprotein bodies found in the interchromatin space of mammalian cell nuclei. These structures play a role in regulating the expression of certain genes in differentiated cells by nuclear retention of RNA. The core paraspeckle proteins (PSF/SFPQ, P54NRB/NONO, and PSPC1 [paraspeckle protein 1]) are members of the DBHS (Drosophila melanogaster behavior, human splicing) family. These proteins, together with the long nonprotein-coding RNA NEAT1 (MEN-epsilon/beta), associate to form paraspeckles and maintain their integrity. Given the large numbers of long noncoding transcripts currently being discovered through whole transcriptome analysis, paraspeckles may be a paradigm for a class of subnuclear bodies formed around long noncoding RNA.

Figure 1.

Visualizing paraspeckles. (A) Combined differential interference contrast and fluorescence micrograph of HeLa cells stained with anti-PSPC1 to show paraspeckles (green) as nucleoplasmic foci distinct from nucleoli (stained with B23 antibody; red). (B) HeLa cells showing reorganization of the DBHS protein PSPC1 (green) to perinucleolar caps after treatment with actinomycin D to inhibit RNA Pol II transcription. (C) HeLa cell stained with anti-PSPC1 (green), anti-SC35 (red), and DAPI (blue) to show the relationship between paraspeckles abutting nuclear speckles in the interchromatin space. (D) TEM image of a HeLa cell section immunogold labeled with anti-PSPC1. The labeled IGAZs are usually found in close proximity to the interchromatin granules (ICGs; nuclear speckles). This image was provided by S. Souquere and G. Pierron (Institut André Lwoff, Villejuif, France). Panels B and C are adapted from Fox et al. (2002) with permission from Elsevier. Bars: (A–C) 10 µm; (D) 0.5 µm.

Figure 2.

The role of nuclear retention in gene regulation in differentiated cells. In the example of the mCat2 gene, two different promoters result in two alternative transcripts: the mCat2 mRNA, which is exported for translation of the mCAT2 cation transporter protein, and Ctn RNA, which is a longer transcript including the mCAT2 protein coding region and an extended 3′ UTR containing inverted repeats. These repeats undergo A to I hyperediting, resulting in Ctn binding to DBHS proteins and retention in paraspeckles. Stress signals mediated by IFN-γ–receptor (IFN-γ–IGR) and lipopolysaccharide–toll-like receptor 4 (LPS–TLR4) interaction result in a cleavage event that liberates a shorter Ctn, which is exported for translation. This up-regulation of the mCAT2 protein results in increased nitric oxide (NO) production as a response to the cellular stress. Although detail in this figure is specific to mCAT2 protein regulation, recent research hints at a more generic retention–release mechanism that exists for other transcripts containing hyperedited inverted repeats in their 3′ UTR (Chen et al., 2008; Chen and Carmichael, 2009; Osenberg et al., 2009).

Figure 3.

Paraspeckles contain NEAT1 ncRNA and form near the NEAT1 gene. (A) NEAT1 and MALAT1 gene loci on human chromosome (Chr) 11 q13.1. Two transcripts are produced from the NEAT1 gene, 3.7-kb NEAT1_v1 and 23-kb NEAT1_v2 in humans. (B) RNA FISH against NEAT1 ncRNA (green) and immunofluorescence against P54NRB/NONO (red) shows that they colocalize in paraspeckles. The line scan is taken over a line as indicated in the merged image. (C) HeLa cells in early G1 stained with PSPC1 (red) to mark the first forming paraspeckles and DNA FISH (green) against 11q13. (D) Interphase HeLa cell with NEAT1 RNA FISH to mark paraspeckles (red) and DNA FISH to mark chromosome 11 q13.1 (green). Panels B–D are reproduced from Clemson et al. (2009) with permission from Elsevier. Bars: (B and C) 10 µm; (D) 5 µm.

Figure 4.

Model of paraspeckle formation. The schematic shows a paraspeckle forming in the interchromatin space in between two nuclear speckles. The paraspeckle forms near the NEAT1 gene locus through the interactions between newly transcribed NEAT1 RNA and DBHS protein dimers. RNA containing A to I hyperedited inverted repeats, as well as potentially other RNA, is regulated within paraspeckles via interaction with DBHS proteins.