PML, a multifunctional protein, is crucial for forming PML-nuclear bodies involved in stress responses. Under specific conditions, PML associates with nucleolar caps formed after RNA polymerase I (RNAPI) inhibition, leading to PML-nucleolar associations (PNAs). This study investigates PNAs-inducing stimuli by exposing cells to various genotoxic stresses. We found that the most potent inducers of PNAs introduced topological stress and inhibited RNAPI. Doxorubicin, the most effective compound, induced double-strand breaks (DSBs) in the rDNA locus. PNAs co-localized with damaged rDNA, segregating it from active nucleoli. Cleaving the rDNA locus with I-PpoI confirmed rDNA damage as a genuine stimulus for PNAs. Inhibition of ATM, ATR kinases, and RAD51 reduced I-PpoI-induced PNAs, highlighting the importance of ATM/ATR-dependent nucleolar cap formation and homologous recombination (HR) in their triggering. I-PpoI-induced PNAs co-localized with rDNA DSBs positive for RPA32-pS33 but deficient in RAD51, indicating resected DNA unable to complete HR repair. Our findings suggest that PNAs form in response to persistent rDNA damage within the nucleolar cap, highlighting the interplay between PML/PNAs and rDNA alterations due to topological stress, RNAPI inhibition, and rDNA DSBs destined for HR. Cells with persistent PNAs undergo senescence, suggesting PNAs help avoid rDNA instability, with implications for tumorigenesis and aging.

• MeSH
• buněčné jadérko * metabolismus   MeSH
• dvouřetězcové zlomy DNA MeSH
• lidé MeSH
• poškození DNA MeSH
• protein promyelocytické leukemie * metabolismus   genetika   MeSH
• ribozomální DNA * genetika   metabolismus   MeSH
• RNA-polymerasa I metabolismus   genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

In nearly all somatic cells, the ribosome biosynthesis is a key activity. The same is true also for mammalian oocytes and early embryos. This activity is intimately linked to the most prominent nuclear organelles - the nucleoli. Interestingly, during a short period around fertilization, the nucleoli in oocytes and embryos transform into ribosome-biosynthesis-inactive structures termed nucleolus-like or nucleolus precursor bodies (NPBs). For decades, researchers considered these structures to be passive repositories of nucleolar proteins used by the developing embryo to rebuild fully functional, ribosome-synthesis competent nucleoli when required. Recent evidence, however, indicates that while these structures are unquestionably essential for development, the material is largely dispensable for the formation of active embryonic nucleoli. In this mini-review, we will describe some unique features of oocytes and embryos with respect to ribosome biogenesis and the changes in the structure of oocyte and embryonic nucleoli that reflect this. We will also describe some of the different approaches that can be used to study nucleoli and NPBs in embryos and discuss the different results that might be expected. Finally, we ask whether the main function of nucleolar precursor bodies might lie in the genome organization and remodelling and what the involved components might be.

• MeSH
• buněčné jadérko metabolismus   MeSH
• buněčné jádro metabolismus   MeSH
• centromera metabolismus   MeSH
• embryo savčí cytologie   MeSH
• embryonální vývoj genetika   MeSH
• fertilizace * MeSH
• histony metabolismus   MeSH
• jaderné proteiny metabolismus   MeSH
• lidé MeSH
• molekulární chaperony metabolismus   MeSH
• oocyty metabolismus   MeSH
• organely metabolismus   MeSH
• ribozomy metabolismus   MeSH
• RNA-polymerasa I metabolismus   MeSH
• savci MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

SUMOylation is a post-translational modification that positively regulates monoallelic expression of the trypanosome variant surface glycoprotein (VSG). The presence of a highly SUMOylated focus associated with the nuclear body, where the VSG gene is transcribed, further suggests an important role of SUMOylation in regulating VSG expression. Here, we show that SNF2PH, a SUMOylated plant homeodomain (PH)-transcription factor, is upregulated in the bloodstream form of the parasite and enriched at the active VSG telomere. SUMOylation promotes the recruitment of SNF2PH to the VSG promoter, where it is required to maintain RNA polymerase I and thus to regulate VSG transcript levels. Further, ectopic overexpression of SNF2PH in insect forms, but not of a mutant lacking the PH domain, induces the expression of bloodstream stage-specific surface proteins. These data suggest that SNF2PH SUMOylation positively regulates VSG monoallelic transcription, while the PH domain is required for the expression of bloodstream-specific surface proteins. Thus, SNF2PH functions as a positive activator, linking expression of infective form surface proteins and VSG regulation, thereby acting as a major regulator of pathogenicity.

• MeSH
• epigeneze genetická MeSH
• glykoproteiny genetika   metabolismus   MeSH
• protozoální proteiny genetika   metabolismus   MeSH
• restrukturace chromatinu MeSH
• RNA-polymerasa I metabolismus   MeSH
• sumoylace * MeSH
• transkripční faktory genetika   metabolismus   MeSH
• Trypanosoma brucei brucei genetika   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Tick-borne encephalitis virus (TBEV), a member of the genus Flavivirus (Flaviviridae), is a causative agent of a severe neuroinfection. Recently, several flaviviruses have been shown to interact with host protein synthesis. In order to determine whether TBEV interacts with this host process in its natural target cells, we analysed de novo protein synthesis in a human cell line derived from cerebellar medulloblastoma (DAOY HTB-186). We observed a significant decrease in the rate of host protein synthesis, including the housekeeping genes HPRT1 and GAPDH and the known interferon-stimulated gene viperin. In addition, TBEV infection resulted in a specific decrease of RNA polymerase I (POLR1) transcripts, 18S and 28S rRNAs and their precursor, 45-47S pre-rRNA, but had no effect on the POLR3 transcribed 5S rRNA levels. To our knowledge, this is the first report of flavivirus-induced decrease of specifically POLR1 rRNA transcripts accompanied by host translational shut-off.

• MeSH
• genetická transkripce MeSH
• klíšťová encefalitida genetika   metabolismus   virologie   MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• prekurzory RNA MeSH
• proteosyntéza genetika   MeSH
• RNA ribozomální genetika   metabolismus   MeSH
• RNA-polymerasa I genetika   metabolismus   MeSH
• viry klíšťové encefalitidy fyziologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Ribosome biogenesis is an energy consuming process which takes place mainly in the nucleolus. By producing ribosomes to fuel protein synthesis, it is tightly connected with cell growth and cell cycle control. Perturbation of ribosome biogenesis leads to the activation of p53 tumor suppressor protein promoting processes like cell cycle arrest, apoptosis or senescence. This ribosome biogenesis stress pathway activates p53 through sequestration of MDM2 by a subset of ribosomal proteins (RPs), thereby stabilizing p53. Here, we identify human HEATR1, as a nucleolar protein which positively regulates ribosomal RNA (rRNA) synthesis. Downregulation of HEATR1 resulted in cell cycle arrest in a manner dependent on p53. Moreover, depletion of HEATR1 also caused disruption of nucleolar structure and activated the ribosomal biogenesis stress pathway - RPL5 / RPL11 dependent stabilization and activation of p53. These findings reveal an important role for HEATR1 in ribosome biogenesis and further support the concept that perturbation of ribosome biosynthesis results in p53-dependent cell cycle checkpoint activation, with implications for human pathologies including cancer.

• MeSH
• biogeneze organel * MeSH
• fyziologický stres MeSH
• genetická transkripce * MeSH
• jaderné proteiny metabolismus   MeSH
• kontrolní body buněčného cyklu MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• nádorový supresorový protein p53 metabolismus   MeSH
• proliferace buněk MeSH
• proteiny vázající RNA metabolismus   MeSH
• protoonkogenní proteiny c-mdm2 metabolismus   MeSH
• ribozomální proteiny metabolismus   MeSH
• ribozomy metabolismus   MeSH
• RNA ribozomální biosyntéza   MeSH
• RNA-polymerasa I genetika   MeSH
• signální transdukce MeSH
• vedlejší histokompatibilní antigeny metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

We studied the effect of ellagic acid (EA) on the morphology of nucleoli and on the pattern of major proteins of the nucleolus. After EA treatment of HeLa cells, we observed condensation of nucleoli as documented by the pattern of argyrophilic nucleolar organizer regions (AgNORs). EA also induced condensation of RPA194-positive nucleolar regions, but no morphological changes were observed in nucleolar compartments positive for UBF1/2 proteins or fibrillarin. Studied morphological changes induced by EA were compared with the morphology of control, non-treated cells and with pronounced condensation of all nucleolar domains caused by actinomycin D (ACT-D) treatment. Similarly as ACT-D, but in a lesser extent, EA induced an increased number of 53BP1-positive DNA lesions. However, the main marker of DNA lesions, γH2AX, was not accumulated in body-like nuclear structures. An increased level of γH2AX was found by immunofluorescence and Western blots only after EA treatment. Intriguingly, the levels of fibrillarin, UBF1/2 and γH2AX were increased at the promoters of ribosomal genes, while 53BP1 and CARM1 levels were decreased by EA treatment at these genomic regions. In the entire genome, EA reduced H3R17 dimethylation. Taken together, ellagic acid is capable of significantly changing the nucleolar morphology and protein levels inside the nucleolus.

• MeSH
• buněčné dělení účinky léků   MeSH
• buněčné jadérko chemie   účinky léků   ultrastruktura   MeSH
• chromozomální proteiny, nehistonové analýza   MeSH
• daktinomycin farmakologie   MeSH
• epigeneze genetická účinky léků   MeSH
• G2 fáze účinky léků   MeSH
• guanylátcyklasa analýza   antagonisté a inhibitory   MeSH
• HeLa buňky chemie   účinky léků   MeSH
• histony analýza   metabolismus   MeSH
• intracelulární signální peptidy a proteiny analýza   MeSH
• kyselina ellagová farmakologie   MeSH
• lidé MeSH
• metylace MeSH
• nádorové proteiny analýza   MeSH
• organizátor jadérka chemie   účinky léků   ultrastruktura   MeSH
• poškození DNA MeSH
• posttranslační úpravy proteinů účinky léků   MeSH
• promotorové oblasti (genetika) MeSH
• ribozomální DNA účinky léků   genetika   MeSH
• RNA-polymerasa I analýza   MeSH
• signální adaptorové proteiny CARD analýza   antagonisté a inhibitory   MeSH
• transkripční iniciační komplex Pol1 - proteiny analýza   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Although actin monomers polymerize into filaments in the cytoplasm, the form of actin in the nucleus remains elusive. We searched for the form and function of β-actin fused to nuclear localization signal and to enhanced yellow fluorescent protein (EN-actin). Our results reveal that EN-actin is either dispersed in the nucleoplasm (homogenous EN-actin) or forms bundled filaments in the nucleus (EN-actin filaments). Formation of such filaments was not connected with increased EN-actin levels. Among numerous actin-binding proteins tested, only cofilin is recruited to the EN-actin filaments. Overexpression of EN-actin causes increase in the nuclear levels of actin-related protein 3 (Arp3). Although Arp3, a member of actin nucleation complex Arp2/3, is responsible for EN-actin filament nucleation and bundling, the way cofilin affects nuclear EN-actin filaments dynamics is not clear. While cells with homogenous EN-actin maintained unaffected mitosis during which EN-actin re-localizes to the plasma membrane, generation of nuclear EN-actin filaments severely decreases cell proliferation and interferes with mitotic progress. The introduction of EN-actin manifests in two mitotic-inborn defects-formation of binucleic cells and generation of micronuclei-suggesting that cells suffer aberrant cytokinesis and/or impaired chromosomal segregation. In interphase, nuclear EN-actin filaments passed through chromatin region, but do not co-localize with either chromatin remodeling complexes or RNA polymerases I and II. Surprisingly presence of EN-actin filaments was connected with increase in the overall transcription levels in the S-phase by yet unknown mechanism. Taken together, EN-actin can form filaments in the nucleus which affect important cellular processes such as transcription and mitosis.

• MeSH
• aktiny metabolismus   MeSH
• bakteriální proteiny metabolismus   MeSH
• buněčné jádro metabolismus   MeSH
• faktory depolymerizující aktin MeSH
• genetická transkripce MeSH
• HEK293 buňky MeSH
• lidé MeSH
• luminescentní proteiny metabolismus   MeSH
• mikrofilamenta metabolismus   MeSH
• mitóza genetika   MeSH
• nádorové buněčné linie MeSH
• protein 3 související s aktinem biosyntéza   metabolismus   MeSH
• restrukturace chromatinu MeSH
• RNA-polymerasa I genetika   MeSH
• RNA-polymerasa II genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

RNA polymerase I (Pol I) transcription is essential for the cell cycle, growth and protein synthesis in eukaryotes. In the present study, we found that phosphatidylinositol 4,5-bisphosphate (PIP2) is a part of the protein complex on the active ribosomal promoter during transcription. PIP2 makes a complex with Pol I and the Pol I transcription factor UBF in the nucleolus. PIP2 depletion reduces Pol I transcription, which can be rescued by the addition of exogenous PIP2. In addition, PIP2 also binds directly to the pre-rRNA processing factor fibrillarin (Fib), and co-localizes with nascent transcripts in the nucleolus. PIP2 binding to UBF and Fib modulates their binding to DNA and RNA, respectively. In conclusion, PIP2 interacts with a subset of Pol I transcription machinery, and promotes Pol I transcription.

• MeSH
• buněčné jadérko genetika   metabolismus   MeSH
• chromozomální proteiny, nehistonové genetika   metabolismus   MeSH
• DNA vazebné proteiny genetika   metabolismus   MeSH
• fosfatidylinositol-4,5-difosfát genetika   metabolismus   MeSH
• genetická transkripce genetika   MeSH
• HeLa buňky MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• prekurzory RNA genetika   metabolismus   MeSH
• promotorové oblasti (genetika) genetika   MeSH
• RNA-polymerasa I genetika   metabolismus   MeSH
• transkripční iniciační komplex Pol1 - proteiny genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

To maintain growth and division, cells require a large-scale production of rRNAs which occurs in the nucleolus. Recently, we have shown the interaction of nucleolar phosphatidylinositol 4,5-bisphosphate (PIP2) with proteins involved in rRNA transcription and processing, namely RNA polymerase I (Pol I), UBF, and fibrillarin. Here we extend the study by investigating transcription-related localization of PIP2 in regards to transcription and processing complexes of Pol I. To achieve this, we used either physiological inhibition of transcription during mitosis or inhibition by treatment the cells with actinomycin D (AMD) or 5,6-dichloro-1β-d-ribofuranosyl-benzimidazole (DRB). We show that PIP2 is associated with Pol I subunits and UBF in a transcription-independent manner. On the other hand, PIP2/fibrillarin colocalization is dependent on the production of rRNA. These results indicate that PIP2 is required not only during rRNA production and biogenesis, as we have shown before, but also plays a structural role as an anchor for the Pol I pre-initiation complex during the cell cycle. We suggest that throughout mitosis, PIP2 together with UBF is involved in forming and maintaining the core platform of the rDNA helix structure. Thus we introduce PIP2 as a novel component of the NOR complex, which is further engaged in the renewed rRNA synthesis upon exit from mitosis.

• MeSH
• buněčné jadérko metabolismus   MeSH
• buněčný cyklus MeSH
• chromozomální proteiny, nehistonové metabolismus   MeSH
• genetická transkripce MeSH
• HeLa buňky MeSH
• lidé MeSH
• mitóza MeSH
• nádorové buněčné linie MeSH
• organizátor jadérka metabolismus   MeSH
• rekombinantní proteiny metabolismus   MeSH
• ribozomální DNA MeSH
• RNA ribozomální MeSH
• RNA-polymerasa I metabolismus   MeSH
• transkripční iniciační komplex Pol1 - proteiny metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Nuclear actin and nuclear myosin I (NMI) are important players in transcription of ribosomal genes. Transcription of rDNA takes place in highly organized intranuclear compartment, the nucleolus. In this study, we characterized the localization of these two proteins within the nucleolus of HeLa cells with high structural resolution by means of electron microscopy and gold-immunolabeling. We demonstrate that both actin and NMI are localized in specific compartments within the nucleolus, and the distribution of NMI is transcription-dependent. Moreover, a pool of NMI is present in the foci containing nascent rRNA transcripts. Actin, in turn, is present both in transcriptionally active and inactive regions of the nucleolus and colocalizes with RNA polymerase I and UBF. Our data support the involvement of actin and NMI in rDNA transcription and point out to other functions of these proteins in the nucleolus, such as rRNA maturation and maintenance of nucleolar architecture.

• MeSH
• aktiny metabolismus   MeSH
• buněčné jadérko metabolismus   MeSH
• genetická transkripce fyziologie   MeSH
• HeLa buňky MeSH
• imunohistochemie MeSH
• lidé MeSH
• myosin typu I metabolismus   MeSH
• ribozomální DNA metabolismus   MeSH
• RNA ribozomální metabolismus   MeSH
• RNA-polymerasa I metabolismus   MeSH
• transkripční iniciační komplex Pol1 - proteiny metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH