Regulation of transcription of genes of ribosomal rna during amphibian oogenesis. A biochemical and morphological study.

Natural changes in the transcription of rRNA genes were studied in nucleoli from three oogenic stages of the newt Triturus alpestris with electron microscope, auto-radiographic, and biochemical techniques. From determinations of the uridine triphosphate pool sizes and [3H]uridine uptake, phosphorylation, and incorporation into 28S and 18S rRNAs in vivo it was estimated that the rate of rRNA synthesis was about 0.01% in previtellogenic oocytes and 13% in mature oocytes when compared to midvitellogenesis. Spread preparations of nucleoli showed significant morphological changes in the transcriptional complexes. The total number of lateral fibrils, i.e., ribonucleoproteins containing the nascent rRNA precursor, were drastically decreased in stages of reduced synthetic activity. This indicates that rRNA synthesis is regulated primarily at the level of transcription. The resulting patterns of fibril coverage of the nucleolar chromatin axes revealed a marked heterogeneity. On the same nucleolar axis occurred matrix units that were completely devoid of lateral fibrils, matrix units that were almost fully covered with lateral fibrils, and various forms of matrix units with a range of lateral fibril densities intermediate between the two extremes. Granular particles that were tentatively identified as RNA polymerase molecules were not restricted to the transciptional complexes. They were observed, although less regularly and separated by greater distances, in untranscribed spacer regions as well as in untranscribed gene intercepts. The results show that the pattern of transcriptional control of rRNA genes differs widely in different genes, even in the same genetic unit.

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In vivo evidence that TATA-binding protein/SL1 colocalizes with UBF and RNA polymerase I when rRNA synthesis is either active or inactive.

The Journal of Cell Biology ◽

10.1083/jcb.133.2.225 ◽

1996 ◽

Vol 133 (2) ◽

pp. 225-234 ◽

Cited By ~ 97

Author(s):

P Jordan ◽

M Mannervik ◽

L Tora ◽

M Carmo-Fonseca

Keyword(s):

Rna Polymerase ◽

Actinomycin D ◽

Binding Protein ◽

Tata Binding Protein ◽

Rna Polymerase I ◽

Rrna Genes ◽

Rrna Synthesis ◽

Rrna Transcription ◽

Polymerase I

Here we show that the TATA-binding protein (TBP) is localized in the nucleoplasm and in the nucleolus of mammalian cells, consistent with its known involvement in transcription by RNA polymerase I, II, and III. In the nucleolus of actively growing cells, TBP colocalizes with upstream binding factor (UBF) and RNA polymerase I at the sites of rRNA transcription. During mitosis, when rRNA synthesis is down-regulated, TBP colocalizes with TBP-associated factors for RNA polymerase I (TAF(I)s), UBF, and RNA polymerase I on the chromosomal regions containing the rRNA genes. Treatment of cells with a low concentration of actinomycin D inhibits rRNA synthesis and causes a redistribution of the rRNA genes that become concentrated in clusters at the periphery of the nucleolus. A similar redistribution was observed for the major components of the rRNA transcription machinery (i.e., TBP, TAF(I)s, UBF, and RNA polymerase I), which still colocalized with each other. Furthermore, anti-TBP antibodies are shown to coimmunoprecipitate TBP and TAF(I)63 in extracts prepared from untreated and actinomycin D-treated cells. Collectively, the data indicate that in vivo TBP/promoter selectivity factor, UBF, and RNA polymerase I remain associated with both active and inactive rRNA genes.

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In vivo regulation of rRNA transcription occurs rapidly in nondividing and dividing Drosophila cells in response to a phorbol ester and serum.

Molecular and Cellular Biology ◽

10.1128/mcb.13.2.928 ◽

1993 ◽

Vol 13 (2) ◽

pp. 928-933 ◽

Cited By ~ 9

Author(s):

S M Vallett ◽

M Brudnak ◽

M Pellegrini ◽

H W Weber

Keyword(s):

Phorbol Ester ◽

Growth Regulation ◽

Rrna Genes ◽

Rrna Synthesis ◽

Rrna Gene ◽

Rdna Transcription ◽

Transcription System ◽

Pol I ◽

Drosophila Cells

The synthesis of ribosomes is an essential cellular process which requires the transcription of the rRNA genes by RNA polymerase I (Pol I). The regulation of rRNA synthesis is known to be coupled to growth regulation. In nongrowing, slowly growing, and rapidly growing Drosophila cells, exposure to the tumor-promoting phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) increases the synthesis of precursor and mature rRNAs. Using nuclear run-on assays, we show that TPA enhances transcription of the rRNA genes. These results suggest that TPA regulates expression of RNA genes transcribed by Pol I, irrespective of the growth state of the cells. In slowly dividing Drosophila cells, increasing the serum concentration rapidly alters the accumulation of rRNA by enhancing rDNA transcription within 1 h. Thus, TPA and serum are each able to rapidly regulate rRNA gene expression in Drosophila cells. These results indicate that the RNA Pol I transcription system can be regulated by agents which have previously been shown to effect specific genes transcribed by the RNA Pol II system.

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Transcriptional control of ribosome production in regenerating rat liver

Biochemical Journal ◽

10.1042/bj2080101 ◽

1982 ◽

Vol 208 (1) ◽

pp. 101-108 ◽

Cited By ~ 19

Author(s):

M D Dabeva ◽

K P Dudov

Keyword(s):

Rat Liver ◽

Transcriptional Control ◽

18S Rrna ◽

Specific Radioactivity ◽

Rrna Synthesis ◽

Transcriptional Level ◽

Enhanced Production ◽

Regenerating Rat Liver ◽

Fold Stimulation

Kinetic experiments of labelling in vivo with [14C]orotate of cellular free UMP and/or UTP, nucleolar, nucleoplasmic and cytoplasmic rRNA in normal and 12 h-regenerating rat liver were performed. The specific-radioactivity curves obtained were analysed by computer and the rates of synthesis of precursor rRNA (45S pre-rRNA) and cytoplasmic 28S and 18S rRNA calculated. (a) The rates of synthesis of 45S pre-rRNA in normal and regenerating rat liver are 1400 and 3700 molecules/min per nucleus respectively; (b) the average rates of formation of mature 28S and 18S rRNA are identical with the rates of synthesis of 45S pre-rRNA in both normal and regenerating rat liver. Thus the synthesis of rRNA in 12h-regenerating rat liver is activated 2.7-fold. The analysis of rRNA synthesis in isolated nucleoli also shows a 2.7-fold stimulation of transcription in regenerating liver. It is concluded that all the 45S pre-rRNA molecules synthesized are processed and transferred as 28S and 18S rRNA in the cytoplasm, i.e. degradation (wastage) of newly synthesized ribosomes in the nucleus does not occur in both normal and regenerating rat liver. Thus the enhanced production of ribosomes in regenerating rat liver is regulated only at the transcriptional level.

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In vivo regulation of rRNA transcription occurs rapidly in nondividing and dividing Drosophila cells in response to a phorbol ester and serum

Molecular and Cellular Biology ◽

10.1128/mcb.13.2.928-933.1993 ◽

1993 ◽

Vol 13 (2) ◽

pp. 928-933

Author(s):

S M Vallett ◽

M Brudnak ◽

M Pellegrini ◽

H W Weber

Keyword(s):

Phorbol Ester ◽

Growth Regulation ◽

Rrna Genes ◽

Rrna Synthesis ◽

Rrna Gene ◽

Rdna Transcription ◽

Transcription System ◽

Pol I ◽

Drosophila Cells

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Ultrastructural Study of a differentiating human colon carcinoma cell line

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100158583 ◽

1990 ◽

Vol 48 (3) ◽

pp. 208-209

Author(s):

Sylvie Polak-Charcon ◽

Mehrdad Hekmati ◽

Yehuda Ben Shaul

Keyword(s):

Cell Line ◽

Morphological Changes ◽

Secretory Granules ◽

Human Colon ◽

Cell Types ◽

Culture Conditions ◽

Morphological Evidence ◽

Human Colon Carcinoma Cell ◽

Colon Cell

The epithelium of normal human colon mucosa “in vivo” exhibits a gradual pattern of differentiation as undifferentiated stem cells from the base of the crypt of “lieberkuhn” rapidly divide, differentiate and migrate toward the free surface. The major differentiated cell type of the intestine observed are: absorptive cells displaying brush border, goblet cells containing mucous granules, Paneth and endocrine cells containing dense secretory granules. These different cell types are also found in the intestine of the 13-14 week old embryo.We present here morphological evidence showing that HT29, an adenocarcinoma of the human colon cell line, can differentiate into various cell types by changing the growth and culture conditions and mimic morphological changes found during development of the intestine in the human embryo.HT29 cells grown in tissue-culture dishes in DMEM and 10% FCS form at late confluence a multilayer of morphologically undifferentiated cell culture covered with irregular microvilli, and devoid of tight junctions (Figs 1-3).

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Spectroscopic imaging of human disease

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100166889 ◽

1996 ◽

Vol 54 ◽

pp. 884-885

Author(s):

D.J. Meyerhoff

Keyword(s):

Magnetic Field ◽

Magnetic Resonance ◽

Field Gradient ◽

Human Disease ◽

Morphological Changes ◽

Magnetic Field Gradient ◽

Spectroscopic Imaging ◽

Resonance Spectroscopy ◽

Tissue Water

Magnetic Resonance Imaging (MRI) observes tissue water in the presence of a magnetic field gradient to study morphological changes such as tissue volume loss and signal hyperintensities in human disease. These changes are mostly non-specific and do not appear to be correlated with the range of severity of a certain disease. In contrast, Magnetic Resonance Spectroscopy (MRS), which measures many different chemicals and tissue metabolites in the millimolar concentration range in the absence of a magnetic field gradient, has been shown to reveal characteristic metabolite patterns which are often correlated with the severity of a disease. In-vivo MRS studies are performed on widely available MRI scanners without any “sample preparation” or invasive procedures and are therefore widely used in clinical research. Hydrogen (H) MRS and MR Spectroscopic Imaging (MRSI, conceptionally a combination of MRI and MRS) measure N-acetylaspartate (a putative marker of neurons), creatine-containing metabolites (involved in energy processes in the cell), choline-containing metabolites (involved in membrane metabolism and, possibly, inflammatory processes),

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The Anti-tumor Activity and Mechanisms of rLj-RGD3 on Human Laryngeal Squamous Carcinoma Hep2 Cells

Anti-Cancer Agents in Medicinal Chemistry ◽

10.2174/1871520619666191022160024 ◽

2020 ◽

Vol 19 (17) ◽

pp. 2108-2119

Author(s):

Yang Jin ◽

Li Lv ◽

Shu-Xiang Ning ◽

Ji-Hong Wang ◽

Rong Xiao

Keyword(s):

Wound Healing ◽

Western Blot ◽

Morphological Changes ◽

In Vivo Studies ◽

Migration And Invasion ◽

Tunel Staining ◽

Dna Ladder ◽

Western Blot Assay

Background: Laryngeal Squamous Cell Carcinoma (LSCC) is a malignant epithelial tumor with poor prognosis and its incidence rate increased recently. rLj-RGD3, a recombinant protein cloned from the buccal gland of Lampetra japonica, contains three RGD motifs that could bind to integrins on the tumor cells. Methods: MTT assay was used to detect the inhibitory rate of viability. Giemsa’s staining assay was used to observe the morphological changes of cells. Hoechst 33258 and TUNEL staining assay, DNA ladder assay were used to examine the apoptotic. Western blot assay was applied to detect the change of the integrin signal pathway. Wound-healing assay, migration, and invasion assay were used to detect the mobility of Hep2 cells. H&E staining assay was used to show the arrangement of the Hep2 cells in the solid tumor tissues. Results: In the present study, rLj-RGD3 was shown to inhibit the viability of LSCC Hep2 cells in vitro by inducing apoptosis with an IC50 of 1.23µM. Western blot showed that the apoptosis of Hep2 cells induced by rLj- RGD3 was dependent on the integrin-FAK-Akt pathway. Wound healing, transwells, and western blot assays in vitro showed that rLj-RGD3 suppressed the migration and invasion of Hep2 cells by integrin-FAKpaxillin/ PLC pathway which could also affect the cytoskeleton arrangement in Hep2 cells. In in vivo studies, rLj-RGD3 inhibited the growth, tumor volume, and weight, as well as disturbed the tissue structure of the solid tumors in xenograft models of BALB/c nude mice without reducing their body weights. Conclusion: hese results suggested that rLj-RGD3 is an effective and safe suppressor on the growth and metastasis of LSCC Hep2 cells from both in vitro and in vivo experiments. rLj-RGD3 might be expected to become a novel anti-tumor drug to treat LSCC patients in the near future.

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Harnessing the Nucleolar DNA Damage Response in Cancer Therapy

Genes ◽

10.3390/genes12081156 ◽

2021 ◽

Vol 12 (8) ◽

pp. 1156

Author(s):

Jiachen Xuan ◽

Kezia Gitareja ◽

Natalie Brajanovski ◽

Elaine Sanij

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Cancer Therapy ◽

Dna Damage Response ◽

Rrna Genes ◽

Rrna Synthesis ◽

Pol I ◽

Clinical Investigations ◽

Damage Response ◽

Synthesis And Processing

The nucleoli are subdomains of the nucleus that form around actively transcribed ribosomal RNA (rRNA) genes. They serve as the site of rRNA synthesis and processing, and ribosome assembly. There are 400–600 copies of rRNA genes (rDNA) in human cells and their highly repetitive and transcribed nature poses a challenge for DNA repair and replication machineries. It is only in the last 7 years that the DNA damage response and processes of DNA repair at the rDNA repeats have been recognized to be unique and distinct from the classic response to DNA damage in the nucleoplasm. In the last decade, the nucleolus has also emerged as a central hub for coordinating responses to stress via sequestering tumor suppressors, DNA repair and cell cycle factors until they are required for their functional role in the nucleoplasm. In this review, we focus on features of the rDNA repeats that make them highly vulnerable to DNA damage and the mechanisms by which rDNA damage is repaired. We highlight the molecular consequences of rDNA damage including activation of the nucleolar DNA damage response, which is emerging as a unique response that can be exploited in anti-cancer therapy. In this review, we focus on CX-5461, a novel inhibitor of Pol I transcription that induces the nucleolar DNA damage response and is showing increasing promise in clinical investigations.

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Dental Implant Site Drilling and Induced Morphological Changes Correlated with Temperature in Pig’s Rib Used as the Human Jaw Model

Applied Sciences ◽

10.3390/app11062493 ◽

2021 ◽

Vol 11 (6) ◽

pp. 2493

Author(s):

Karol Kirstein ◽

Michalina Horochowska ◽

Jacek Jagiełło ◽

Joanna Bubak ◽

Aleksander Chrószcz ◽

...

Keyword(s):

Bone Tissue ◽

Morphological Changes ◽

Bone Destruction ◽

Microscopic Investigation ◽

In Vivo Studies ◽

Drilling Speed ◽

Tissue Destruction ◽

Destruction Zone ◽

Jaw Model

The bone tissue destruction during drilling is still one of the crucial problems in implantology. In this study, the influence of drilling speed, coolant presence, and its temperature on bone tissue was tested using swine rib as a biological model of human jaws. The same method of drilling (with or without coolant) was used in all tested samples. The microscopic investigation estimated the size of the destruction zone and morphology of bone tissue surrounding the drilling canal. The achieved results were statistically elaborated. The study proved that the optimal drilling speed was ca. 1200 rpm, but the temperature of the used coolant had no significant influence on provoked bone destruction. Simultaneously, the drilling system without coolant compared to this with coolant has statistical importance on drilling results. Further in vivo studies will verify the obtained results.

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Osteocytes as main responders to low-intensity pulsed ultrasound treatment during fracture healing

Scientific Reports ◽

10.1038/s41598-021-89672-9 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Tatsuya Shimizu ◽

Naomasa Fujita ◽

Kiyomi Tsuji-Tamura ◽

Yoshimasa Kitagawa ◽

Toshiaki Fujisawa ◽

...

Keyword(s):

Fracture Healing ◽

Transcriptional Control ◽

Target Genes ◽

Pulsed Ultrasound ◽

Activated T Cells ◽

In Vivo Models ◽

Low Intensity Pulsed Ultrasound ◽

Low Intensity ◽

Ultrasound Stimulation

AbstractUltrasound stimulation is a type of mechanical stress, and low-intensity pulsed ultrasound (LIPUS) devices have been used clinically to promote fracture healing. However, it remains unclear which skeletal cells, in particular osteocytes or osteoblasts, primarily respond to LIPUS stimulation and how they contribute to fracture healing. To examine this, we utilized medaka, whose bone lacks osteocytes, and zebrafish, whose bone has osteocytes, as in vivo models. Fracture healing was accelerated by ultrasound stimulation in zebrafish, but not in medaka. To examine the molecular events induced by LIPUS stimulation in osteocytes, we performed RNA sequencing of a murine osteocytic cell line exposed to LIPUS. 179 genes reacted to LIPUS stimulation, and functional cluster analysis identified among them several molecular signatures related to immunity, secretion, and transcription. Notably, most of the isolated transcription-related genes were also modulated by LIPUS in vivo in zebrafish. However, expression levels of early growth response protein 1 and 2 (Egr1, 2), JunB, forkhead box Q1 (FoxQ1), and nuclear factor of activated T cells c1 (NFATc1) were not altered by LIPUS in medaka, suggesting that these genes are key transcriptional regulators of LIPUS-dependent fracture healing via osteocytes. We therefore show that bone-embedded osteocytes are necessary for LIPUS-induced promotion of fracture healing via transcriptional control of target genes, which presumably activates neighboring cells involved in fracture healing processes.

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