Intranuclear Distribution of Poly(A) RNA Determined by Electron Microscope in Situ Hybridization

Visa, Neus; Puvion‐Dutilleul, Francine; Harper, Francis; Bachellerie, Jean‐Pierre; Puvion, Edmond

doi:10.1006/excr.1993.1218

Cited by 148 publications

(127 citation statements)

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“…Interestingly, following a 60-and 90-minute pulse, a few gold particles were present over some clusters of interchromatin granules (Fig. 9D) in which some polyadenylated RNA molecules are known to be present (Visa et al, 1993). Taken together, FU pulses of different durations demonstrate the specificity of this procedure and the absence of interrelations between the BCL6 bodies and the RNA molecules.…”

Section: Bcl6 Bodies Are Not Associated With Rnamentioning

confidence: 71%

The relationship between BCL6 bodies and nuclear sites of normal and halogenated DNA and RNA synthesis

Puvion‐Dutilleul¹,

Souquere-Besse²,

Albagli-Curiel

2003

Microscopy Res & Technique

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BCL6 is a POZ/BTB and zinc finger transcription factor that self-interacts and accumulates into discrete nuclear "bodies" of unknown function. We recently reported that BCL6 bodies associate with bromodeoxyuridine (BrdU)-substituted DNA, suggesting their implication in replication. To examine this possibility, we examine here by electron and confocal microscopy the relation between BCL6 bodies and replication foci (RF) using incorporation of various halogenated nucleotides (BrdU, chlorodeoxyuridine, CldU, and iododeoxyuridine, IdU) or PCNA (proliferating cell nuclear antigen) staining. We show that BCL6 bodies are found associated with RF, as revealed by PCNA staining. However, such association is markedly prolonged upon BrdU or CldU incorporation, but less, or not at all, upon IdU incorporation. Pulse-chase and double-labeling experiments indicate that IdU-substituted DNA leaves BCL6 bodies after a few tenths of minutes while BrdUor CldU-substituted DNA stalls in their vicinity for several hours, thereby giving the characteristic "crowns" of DNA entirely surrounding BCL6 bodies. In all cases, however, the halogenated DNA ends up undergoing a movement from BCL6 bodies toward nucleoplasm and nuclear periphery to reach euchromatin and heterochromatin, respectively. We propose that replicating DNA is prone to be bound by BCL6, while BrdU/CldU incorporation increases this propensity possibly because these two events have synergistic effects on the structure and chromatinisation of the newly synthesized DNA. Finally, despite the known proximity between nuclear sites of transcription and replication, we show via several approaches that BCL6 bodies do not appear to be involved either in RNA synthesis or storage.

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Section: Bcl6 Bodies Are Not Associated With Rnamentioning

confidence: 71%

The relationship between BCL6 bodies and nuclear sites of normal and halogenated DNA and RNA synthesis

Puvion‐Dutilleul¹,

Souquere-Besse²,

Albagli-Curiel

2003

Microscopy Res & Technique

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“…This probe also labeled the same late phase clusters that contain snRNP (compare g and h). Oligo dT probes have been previously shown to label the splicing factor rich interchromatin granule network in uninfected cells (Carter et al, 1991(Carter et al, , 1993Visa et al, 1993, see also Fig. 4 h).…”

Section: Spliced Viral Rnas and Polyadenylated Rnas Are Present In Lamentioning

confidence: 82%

“…Perichromatin fibrils are thought to be the in situ forms of newly synthesized hnRNA. Interchromatin granules are not strongly labeled by a pulse of [3H]uridine and are therefore not likely to be locations of transcription, but several studies have shown that polyadenylated RNA is present in these structures (Carter et al, 1991(Carter et al, , 1993Visa et al, 1993). SnRNP and several nonsnRNP splicing factors accumulate at both perichromatin fibrils and interchromatin granules, raising the possibility that splicing of precursor RNA may occur at these locations.…”

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confidence: 99%

Spliced exons of adenovirus late RNAs colocalize with snRNP in a specific nuclear domain.

Bridge

Riedel

Johansson

et al. 1996

The Journal of Cell Biology

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Abstract. Posttranscriptional steps in the production of mRNA include well characterized polyadenylation and splicing reactions, but it is also necessary to understand how RNA is transported within the nucleus from the site of its transcription to the nuclear pore, where it is translocated to the cytoplasmic compartment. Determining the localization of RNA within the nucleus is an important aspect of understanding RNA production and may provide clues for investigating the trafficking of RNA within the nucleus and the mechanism for its export to the cytoplasm. We have previously shown that late phase adenovirus-infected cells contain large clusters of snRNP and non-snRNP splicing factors; the presence of these structures is correlated with high levels of viral late gene transcription. The snRNP clusters correspond to enlarged interchromatin granules present in late phase infected cells. Here we show that polyadenylated RNA and spliced tripartite leader exons from the viral major late transcription unit are present in these same late phase snRNP-containing structures. We find that the majority of the steady state viral RNA present in the nucleus is spliced at the tripartite leader exons. Tripartite leader exons are efficiently exported from the nucleus at a time when we detect their accumulation in interchromatin granule clusters. Since the enlarged interchromatin granules contain spliced and polyadenylated RNA, we suggest that viral RNA may accumulate in this late phase structure during an intranuclear step in RNA transport.T HE production of eukaryotic mRNAs can be considered in several steps; these include transcription, polyadenylation, splicing, and export of mRNA from the nucleus to the cytoplasm. Export involves movement of the RNA within the nucleus from the site of transcription to the nuclear pore, followed by transport of the message through the nuclear pore and into the cytoplasm. Although it is convenient to think of these posttranscriptional processes as separate steps, they are likely to be inter-related during the production of RNA in vivo. All of these reactions occur within the structural framework of the nucleus; thus the nuclear organization of gene expression activities has recently received considerable attention (for reviews see Carter, 1994;Wansink et al., 1994;.The nucleus has a high degree of organization with the nucleolus as the most obvious and well characterized nuclear compartment (Scheer et al., 1993 Fakan, 1994). These include perichromatin fibrils, interchromatin granules, and coiled bodies. Perichromatin fibrils are strongly labeled after short pulses of [3H]uridine indicating that transcription occurs at these structures. Perichromatin fibrils are thought to be the in situ forms of newly synthesized hnRNA. Interchromatin granules are not strongly labeled by a pulse of [3H]uridine and are therefore not likely to be locations of transcription, but several studies have shown that polyadenylated RNA is present in these structures (Carter et al., 1991(Carter et al., , 1993Visa et al....

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“…Perichromatin fibrils contain hnRNP cOre proteins and snRNPs [33], Ui and U2 snRNAs [34], as weil as the splicing factor SC-35 [35]. Sinee the perichromatin fibrils have previous ly been shown to represent nascent pre-mRNA [36,37], it is highly probable that splieing takes place on these structures.…”

Section: Resul Ts and Discussionmentioning

confidence: 99%

“…The interchromatin granule clusters contain only low levels of rapidly labeled RNA [for review, see 39] and hnRNP proteins [33]. However, they do contain poly(A) (see above) and large amounts of different splicing factars [23,[33][34][35]40]. Although it is generally assumed that the interchromatin granule clusters do not represent splicing organelles, their true function in nuclear RNA metabolism remains unknown [see 30 for a recent discussion] .…”

Section: Resul Ts and Discussionmentioning

confidence: 99%

Immunodetection of Poly(A) Binding Protein II in the Cell Nucleus

Krause

Fakan

Weis

et al. 1994

Experimental Cell Research

116

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During the polyadenylation of pre-mRN A in vitro~ poly(A) binding protein 11 (PAB 11) bind. to tbe growing poly(A) taH, stimulating its extension. Tbe subcellular localization of PAß 11 was investigated witb an antibody affinity-purified from rabbit serum raised against the purified protein. Immunoßuorescence microscopy detected PAß 11 exclusively in tbe cell nueleus, botb in a widespread staining and in more intensely stained "speckles." PAß 11 was excluded from the nucleoli. ßy electron microscopy, PAß II was also found almost exclusively in the nucleus, predominantly in clusters of interchromatin granules, likely corresponding to the speckles observed by immunofluorescence microscopy, and in perichromatin fibrils, wbich represent nascent transeripts and probably tbe sites of pre-mRN A processing. In addition, electron microscopy also detected P AB 11 in nucleoli. Tbe distribution corresponds largely to that of other factors involved in the processing of premRNA and is thus in agreement witb the proposed role of the protein in polyadenylation.

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Intranuclear Distribution of Poly(A) RNA Determined by Electron Microscope in Situ Hybridization

Cited by 148 publications

References 0 publications

The relationship between BCL6 bodies and nuclear sites of normal and halogenated DNA and RNA synthesis

The relationship between BCL6 bodies and nuclear sites of normal and halogenated DNA and RNA synthesis

Spliced exons of adenovirus late RNAs colocalize with snRNP in a specific nuclear domain.

Immunodetection of Poly(A) Binding Protein II in the Cell Nucleus

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