Ectopic NORs on human chromosomes 4qter and 8q11: rare chromosomal variants detected in two families.
Two different NOR bearing non-acrocentric chromosomes were detected during prenatal diagnosis performed on two probands because of advanced maternal age. In the first case, a chromosome 4 carried a NOR in the telomeric region of the long arm (4qs), while in the second case a NOR was inserted into chromosome 8q11. Family analysis showed the variant chromosomes to be transmitted through at least three generations in each family. There were no reports of reproductive problems or phenotypic effects in the carriers of these chromosomes, indicating the benign character of the aberrant chromosomes. In order to characterise the chromosomal variants more precisely, various differential banding techniques were applied. (+info)
Cell cycle-dependent regulation of RNA polymerase I transcription: the nucleolar transcription factor UBF is inactive in mitosis and early G1.
Transcription of ribosomal RNA genes by RNA polymerase (pol) I oscillates during the cell cycle, being maximal in S and G2 phase, repressed during mitosis, and gradually recovering during G1 progression. We have shown that transcription initiation factor (TIF)-IB/SL1 is inactivated during mitosis by cdc2/cyclin B-directed phosphorylation of TAFI110. In this study, we have monitored reactivation of transcription after exit from mitosis. We demonstrate that the pol I factor UBF is also inactivated by phosphorylation but recovers with different kinetics than TIF-IB/SL1. Whereas TIF-IB/SL1 activity is rapidly regained on entry into G1, UBF is reactivated later in G1, concomitant with the onset of pol I transcription. Repression of pol I transcription in mitosis and early G1 can be reproduced with either extracts from cells synchronized in M or G1 phase or with purified TIF-IB/SL1 and UBF isolated in the presence of phosphatase inhibitors. The results suggest that two basal transcription factors, e.g., TIF-IB/SL1 and UBF, are inactivated at mitosis and reactivated by dephosphorylation at the exit from mitosis and during G1 progression, respectively. (+info)
Colchicine therapy for hepatic murine schistosomal fibrosis: image analysis and serological study.
Colchicine in a dose of 200 micrograms kg body weight/day (5 days/week) was administered to groups of Schistosoma mansoni infected mice 12 weeks post infection, either alone or following previous praziquantel therapy at the 8th week of infection. Certain groups received colchicine for 6 weeks and others received it for 10 weeks. Colchicine alone did not significantly change the light microscopic appearance of schistosomal liver fibrosis, or hepatic collagen content estimated histomorphometrically, and did not reduce the elevated IL-2 serum level. Colchicine induced hepatic injury consisted of intense inflammatory reaction in granuloma and portal tracts, hepatocytic degeneration, and elevation of serum AST and ALT levels. Colchicine seemed to postpone granulomatous reaction healing and collagen deposition rather than inhibiting collagen formation or degrading it. Colchicine inhibited proliferation of hepatocytes of infected mice by expanding G2-M phases of cell cycle, thus reduced Ag NOR count and raised cell ploidy and cyclic AMP serum level. Subsidence of schistosomal infection by praziquantel prior to colchicine therapy greatly reduced inflammatory cellular reaction, significantly diminished hepatic collagen deposition and serum IL-2 level, minimized the elevated nuclear ploidy and cyclic AMP serum level that followed colchicine therapy when administered alone. (+info)
Research on nucleolar organizer regions of hippocampal neuron in Alzheimer's disease.
OBJECTIVE: To understand the cellular genetic expression of the cell's population by studying the nucleolar organizer regions (NORs) of hippocampal neuron of Alzheimer's disease (AD). METHODS: The postmortem human hippocampal tissues were divided into three groups, namely, the young, the elderly and the AD groups. Each group contained tissues from 10 patients. The study was conducted using image pattern analysis of the nucleoli-nucleoplasms ratio of the neurons of Nissl's stained pathological cerebral hippocampal tissues, the area of stain, and the integrating absorption of nucleoli of silver-stained NORs. RESULTS: The nucleoli-nucleoplasms ratio of the neurons of Nissl's stained cerebral hippocampal tissues, the area of stain, and the integrating absorption of the nucleoli of hippocampal neuron were decreased in the elderly and the AD groups as compared with the young group. However, the area of stain and the integrating absorption of the nucleoli of the hippocampal neurons were relatively increased in the AD group in comparison with the elderly group. CONCLUSION: Nissl's stain demonstrates the hypofunction of the hippocampal neurons in the elderly and the AD patients. The Silver stain of NORs shows the decline of rDNA transcription activity of the nucleoli of the hippocampal neurons in the elderly and the AD patients. However, the transcription activity of the nucleoli of the hippocampal neurons of AD patients was relatively improved, and the cellular genetic expression of the cell's population was relatively strengthened. These cellular morphological changes have probably reflected the cellular defensive system. (+info)
The mitotically phosphorylated form of the transcription termination factor TTF-1 is associated with the repressed rDNA transcription machinery.
The transcription termination factor TTF-1 exerts two functions in ribosomal gene (rDNA) transcription: facilitating initiation and mediating termination of transcription. Using HeLa cells, we show that TTF-1 protein is colocalized with the active transcription machinery in the nucleolus and also with the inactive machinery present in certain mitotic nucleolar organizer regions (NORs) when rDNA transcription is repressed. We also show that TTF-1 is specifically phosphorylated during mitosis in a manner dependent on the cdc2-cyclin B kinase pathway and on an okadaic acid-sensitive phosphatase. Interestingly, the mitotically phosphorylated form of TTF-1 appearing at the G(2)/M transition phase was more easily solubilized than was the interphase form. This indicates that the chromatin-binding affinity of TTF-1 appears to be different in mitotic chromosomes compared to the interphase nucleolus. Correlated with this, the other DNA-binding factor, UBF, which interferes with chromatin conformation in the rDNA promoter, was more strongly bound to rDNA during mitosis than at interphase. The reorganization of the mitotic rDNA promoter might be induced by phosphorylation of certain components of the rDNA transcription machinery and participate in silencing of rDNA during mitosis. (+info)
Chromosomes as well as chromosomal subdomains constitute distinct units in interphase nuclei.
Fluorescence in situ hybridization has demonstrated that chromosomes form individual territories in interphase nuclei. However, this technique is not suitable to determine whether territories are mutually exclusive or interwoven. This notion, however, is essential for understanding functional organizations in the cell nucleus. Here, we analyze boundary areas of individual chromosomes during interphase using a sensitive method based on replication labeling and immunocytochemistry. Thymidine analogues IdUrd and CldUrd were incorporated during S-phase into DNA of Chinese Hamster fibroblasts. Cells labeled with IdUrd were fused with cells labeled with CldUrd. Fused nuclei contained both IdUrd or CldUrd labeled chromosomes. Alternatively, the two labels were incorporated sequentially during successive S-phases and segregated to separate chromosomes by culturing the cells one more cell cycle. Metaphase spreads showed IdUrd-, CldUrd- and unlabeled chromosomes. Some chromatids were divided sharply in differently labeled subdomains by sister chromatid exchanges. With both methods, confocal imaging of interphase nuclei revealed labeled chromosomal domains containing fiber-like structures and unlabeled areas. At various sites, fiber-like structures were embedded in other territories. Even so, essentially no overlap between chromosome territories or between subdomains within a chromosome was observed. These observations indicate that chromosome territories and chromosomal subdomains in G(1)-phase are mutually exclusive at the resolution of the light microscope. (+info)
Molecular dissection of nucleolin's role in growth and cell proliferation: new insights.
Cells require optimum protein synthetic activity in order to support cell proliferation, maintain homeostatic and metabolic integrity, and repair damage. Since growth depends on protein synthesis through ribosome biogenesis, the control of biosynthesis of ribosomes is necessarily a key element for control of growth. Nucleolin is a major nucleolar protein of exponentially growing eukaryotic cells, which is directly involved in the regulation of ribosome biogenesis and maturation. The highly conserved nucleolin contains three major domains through which it controls the organization of nucleolar chromatin, packaging of pre-RNA, rDNA transcription, and ribosome assembly. Numerous reports have implicated the involvement of nucleolin either directly or indirectly in the regulation of cell proliferation and growth, cytokinesis, replication, embryogenesis, and nucleogenesis. Nucleolin, an RNA binding protein, is also an autoantigen, a transcriptional repressor, and a switch region targeting factor. In addition, nucleolin exhibits autodegradation, DNA and RNA helicase activities, and DNA-dependent ATPase activity. An interesting aspect of nucleolin action is that it is a target for regulation by proteolysis, methylation, ADP-ribosylation, and phosphorylation by CKII, cdc2, PKC-xi, cyclic AMP-dependent protein kinase, and ecto-protein kinase. For these and other reasons, nucleolin is fundamental to the survival and proliferation of cells. Considerable progress has been made in recent years with the identification of new nucleolin binding proteins that may mediate these many nucleolin-dependent functions. Nucleolin also functions as a cell surface receptor, where it acts as a shuttling protein between cytoplasm and nucleus, and thus can even provide a mechanism for extracellular regulation of nuclear events. Exploration of the regulation of this multifaceted protein in a remarkable number of diverse functions is challenging. (+info)
Human Nopp140, which interacts with RNA polymerase I: implications for rRNA gene transcription and nucleolar structural organization.
Nopp140 is thought to shuttle between nucleolus and cytoplasm. However, the predominant nucleolar localization of Nopp140 homologues from different species suggests that Nopp140 is also involved in events occurring within the nucleolus. In this study, we demonstrated that the largest subunit of RNA polymerase I, RPA194, was coimmunoprecipitated with the human Nopp140 (hNopp140). Such an interaction is mediated through amino acids 204 to 382 of hNopp140. By double immunofluorescence, hNopp140 was colocalized with RNA polymerase I at the rDNA (rRNA genes) transcription active foci in the nucleolus. These results suggest that Nopp140 can interact with RNA polymerase I in vivo. Transfected cells expressing the amino-terminal half of hNopp140, hNopp140N382 (amino acids 1 to 382), displayed altered nucleoli with crescent-shaped structures. This phenotype is reminiscent of the segregated nucleoli induced by actinomycin D treatment, which is known to inhibit rRNA synthesis. Consistently, the hNopp140N382 protein mislocalized the endogenous RNA polymerase I and shut off cellular rRNA gene transcription as revealed by an in situ run-on assay. These dominant negative effects of the mutant hNopp140N382 suggest that Nopp140 plays an essential role in rDNA transcription. Interestingly, ectopic expression of hNopp140 to a very high level caused the formation of a transcriptionally inactive spherical structure occupying the entire nucleolar area which trapped the RNA polymerase I, fibrillarin, and hNopp140 but excluded the nucleolin. The mislocalizations of these nucleolar proteins after hNopp140 overexpression imply that Nopp140 may also play roles in maintenance of nucleolar integrity. (+info)