Dynamic sorting of nuclear components into distinct nucleolar caps during transcriptional inhibition

Department of Molecular Cell Biology, The Weizmann Institute of Science, Rehovot, 76100 Israel.
Molecular Biology of the Cell (Impact Factor: 4.47). 06/2005; 16(5):2395-413. DOI: 10.1091/mbc.E04-11-0992
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Nucleolar segregation is observed under some physiological conditions of transcriptional arrest. This process can be mimicked by transcriptional arrest after actinomycin D treatment leading to the segregation of nucleolar components and the formation of unique structures termed nucleolar caps surrounding a central body. These nucleolar caps have been proposed to arise from the segregation of nucleolar components. We show that contrary to prevailing notion, a group of nucleoplasmic proteins, mostly RNA binding proteins, relocalized from the nucleoplasm to a specific nucleolar cap during transcriptional inhibition. For instance, an exclusively nucleoplasmic protein, the splicing factor PSF, localized to nucleolar caps under these conditions. This structure also contained pre-rRNA transcripts, but other caps contained either nucleolar proteins, PML, or Cajal body proteins and in addition nucleolar or Cajal body RNAs. In contrast to the capping of the nucleoplasmic components, nucleolar granular component proteins dispersed into the nucleoplasm, although at least two (p14/ARF and MRP RNA) were retained in the central body. The nucleolar caps are dynamic structures as determined using photobleaching and require energy for their formation. These findings demonstrate that the process of nucleolar segregation and capping involves energy-dependent repositioning of nuclear proteins and RNAs and emphasize the dynamic characteristics of nuclear domain formation in response to cellular stress.

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    • "AM-D treatment in higher eukaryotes leads to the condensation and subsequent separation of the FC and GC nucleolar regions [30]. Accordingly, in Dictyostelium different proteins leave the nucleolus in different ways after AM-D treatment which is consistent with the existence of nucleolar subcompartments [15] [16] [17]. "
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    ABSTRACT: The nucleolus is a multifunctional nuclear compartment usually consisting of two to three subcompartments which represent stages of ribosomal biogenesis. It is linked to several human diseases including viral infections, cancer, and neurodegeneration. Dictyostelium is a model eukaryote for the study of fundamental biological processes as well as several human diseases however comparatively little is known about its nucleolus. Unlike most nucleoli it does not possess visible subcompartments at the ultrastructural level. Several recently identified nucleolar proteins in Dictyostelium leave the nucleolus after treatment with the rDNA transcription inhibitor actinomycin-D (AM-D). Different proteins exit in different ways, suggesting that previously unidentified nucleolar subcompartments may exist. The identification of nucleolar subcompartments would help to better understand the nucleolus in this model eukaryote. Here, we show that Dictyostelium nucleolar proteins nucleomorphin isoform NumA1 and Bud31 localize throughout the entire nucleolus while calcium-binding protein 4a localizes to only a portion, representing nucleolar subcompartment 1 (NoSC1). SWI/SNF complex member Snf12 localizes to a smaller area within NoSC1 representing a second nucleolar subcompartment, NoSC2. The nuclear/nucleolar localization signal KRKR from Snf12 localized GFP to NoSC2, and thus also appears to function as a nucleolar subcompartment localization signal. FhkA localizes to the nucleolar periphery displaying a similar pattern to that of Hsp32. Similarities between the redistribution patterns of Dictyostelium nucleolar proteins during nucleolar disruption as a result of either AM-D treatment or mitosis support these subcompartments. A model for the AM-D-induced redistribution patterns is proposed.
    Biochemical and Biophysical Research Communications 12/2014; 433(4). DOI:10.1016/j.bbrc.2014.12.050 · 2.30 Impact Factor
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    • "2. The nucleolus as a stress sensor Because the process of ribosome biogenesis is extremely demanding in terms of energy and resources, its fidelity is closely inspected and virtually any type of severe cellular stress will result in an immediate shutdown of rRNA transcription (Fig. 1B) [19]. In response to such stress conditions, including exposure to different genotoxic agents like doxorubicin or inhibition of rRNA transcription using low levels of Actinomycin D (ActD), the nucleolus undergoes distinct structural changes, including condensation and segregation into structures called nucleolar caps, composed of nucleolar proteins and RNA [17] [20] [21]. Consequently, detection Table 1 A list of reported participants in ribosome biogenesis stress signaling (upper) and key regulators of ribosome biogenesis (lower). "
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    ABSTRACT: Cell proliferation and cell growth are two tightly linked processes, as the proliferation program cannot be executed without proper accumulation of cell mass, otherwise endangering the fate of the two daughter cells. It is therefore not surprising that ribosome biogenesis, a key element in cell growth, is regulated by many cell cycle regulators. This regulation is exerted transcriptionally and post-transcriptionally, in conjunction with numerous intrinsic and extrinsic signals. Those signals eventually converge at the nucleolus, the cellular compartment that is not only responsible for executing the ribosome biogenesis program, but also serves as a regulatory hub, responsible for integrating and transmitting multiple stress signals to the omnipotent cell fate gatekeeper, p53. In this review we discuss when, how and why p53 is activated upon ribosomal biogenesis stress, and how perturbation of this critical regulatory interplay may impact human disease.
    FEBS letters 04/2014; 588(16). DOI:10.1016/j.febslet.2014.04.014 · 3.17 Impact Factor
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    • "Nucleophosmin is an important nucleolar protein supporting translation of ribosomal proteins and associated with the nucleolar fibrillar component. Its localization depends on the functional state of nucleoli28. Although the connection between nuclear and nucleolar distribution of ΔNp63 and nucleophosmin is unclear, localization of nucleophosmin can serve as an additional indicator of TP53-related effects in MCF10A cells. "
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    ABSTRACT: Multiple observations suggest a cell type-specific role for TP53 in mammary epithelia. We developed an in vitro assay, in which primary mouse mammary epithelial cells (mMECs) progressed from lumenal to basal-like phenotypes based on expression of Krt18 or ΔNp63, respectively. Such transition was markedly delayed in Trp53(-/-) mMECs suggesting that Trp53 is required for specification of the basal, but not lumenal cells. Evidence from human basal-like cell lines suggests that TP53 may support the activity of ΔNp63 by preventing its translocation from nucleoplasm into nucleoli. In human lumenal cells, activation of TP53 by inhibiting MDM2 or BRCA1 restored the nucleoplasmic expression of ΔNp63. Trp53(-/-) mMECs eventually lost epithelial features resulting in upregulation of MDM2 and translocation of ΔNp63 into nucleoli. We propose that TP63 may contribute to TP53-mediated oncogenic transformation of epithelial cells and shed light on tissue- and cell type-specific biases observed for TP53-related cancers.
    Scientific Reports 04/2014; 4:4663. DOI:10.1038/srep04663 · 5.58 Impact Factor
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