The memory function of noradrenergic activity in non-REM sleep.

There is a long-standing assumption that low noradrenergic activity during sleep reflects mainly the low arousal during this brain state. Nevertheless, recent research has demonstrated that the locus coeruleus, which is the main source of cortical noradrenaline, displays discrete periods of intense firing during non-REM sleep, without any signs of awakening. This transient locus coeruleus activation during sleep seems to occur in response to preceding learning-related episodes. In the present study, we manipulate noradrenergic activity during sleep in humans with either the α2-autoreceptor agonist clonidine or the noradrenaline reuptake inhibitor reboxetine. We show that reducing noradrenergic activity during sleep, but not during wakefulness, impairs subsequent memory performance in an odor recognition task. Increasing noradrenergic availability during sleep, in contrast, enhances memory retention. We conclude that noradrenergic activity during non-REM sleep interacts with other sleep-related mechanisms to functionally contribute to off-line memory consolidation.

Changes in expression of imprinted genes following treatment of human cancer cell lines with non-mutagenic or mutagenic carcinogens.

It remains possible that chemicals that act by mutagenic mechanisms as well as chemicals that do not induce gene mutations may affect epigenetic gene expression. To test the possibility, we investigated the ability of both types of chemicals to alter the expression of five imprinted genes, PEG3, SNRPN, NDN, ZAC and H19, using two human colon cancer cell lines and a human breast cancer cell line. The expression of imprinted genes was changed by some non-mutagenic and mutagenic carcinogens independent of their mutagenic activity. The genes most commonly exhibiting the changes in expression were SNRPN and PEG3. Alterations of the expression of NDN and ZAC were also observed in some conditions. Methylation-specific PCR and chromatin immunoprecipitation assays suggest the possibility that changes in the expression of SNRPN may be associated with DNA hypomethylation and histone acetylation of the promoters and euchromatinization of the heterochromatic domains of the promoters. Changes in expression of the imprinted genes, PEG3 and NDN, were also observed in cells immortalized by treatment of normal human fibroblasts with 4-nitroquinoline 1-oxide or aflatoxin B1. We previously demonstrated that expression of the cancer-related gene, INK4a, in these immortal cells was lost via epigenetic mechanisms. The results prove that, in cancer cells, some mutagenic or non-mutagenic carcinogens can epigenetically influence the transcription levels of imprinted genes and also suggest the possibility that some chemical carcinogens may have epigenetic carcinogenic effects in human cells.

Immunocytochemical study of estrogen receptor activation factor (E-RAF) and the proteins that interact with nuclear estrogen receptor II (nER II) in epithelial endometrial cells, in the presence and in the absence of estradiol.

The localization and abundance of the estrogen receptor activation factor (E-RAF) and a small nuclear ribonucleoprotein (snRNP) complex containing three proteins, p32, p55 and p60, which interact with the nuclear estrogen receptor II (nER II), have been studied in rat endometrial epithelial cells by means of immunofluorescence and high resolution quantitative immunocytochemistry. In the cytoplasm E-RAF is associated with the rough endoplasmic reticulum. In the nucleus it is mainly localized at the interchromatin space, and surrounding the clumps of compact or semi-condensed chromatin. Quantitative analyses show that the abundance of E-RAF in the nucleus increases after ovariectomy and decreases 3 minutes after estradiol administration. These results are in agreement with the currently available biochemical data. Double immunolocalizations demonstrate that p32, p55, p60 co-localize with other splicing-related protein. High resolution immunolocalization shows that p32, p55, p60 are associated with perichromatin fibrils (co-transcriptional splicing) and with clusters of interchromatin granules (storage of splicing-related molecules). The nuclear abundance of the snRNP complex decreases with ovariectomy, increases within 3 minutes after estradiol administration and remains higher than that in ovariectomized animals for 27 minutes. These results strongly support the previous data on the role of nER-II in the regulation of mRNA transcription and its export from the nucleus to the cytoplasm.

cAMP-dependent reorganization of the Cajal bodies and splicing machinery in cultured Schwann cells.

It is well established that forskolin-induced elevation of cAMP results in activation of DNA synthesis in Schwann cell cultures. This promitotic response is partially mediated by the Cdk2, which is required for the transition from the G1 to the S phase of the cell cycle. In the present study, we analyze the effects of cAMP elevation in cultured Schwann cells on the transcriptional activity and on the organization of two nuclear compartments involved in pre-mRNA processing: Cajal bodies (CBs) and splicing factor compartments. Our immunofluorescence and quantitative studies show that forskolin treatment induces a 5.6-fold increase in the proportion of S phase Schwann cells, detected by a short pulse (20 min) of BrdU incorporation. This increase in DNA synthesis correlates with an activation of global transcription, as is indicated by the higher nuclear incorporation of BrU in nascent RNA. Forskolin treatment significantly increases the percentage of Schwann cells containing typical CBs, which concentrate spliceosomal snRNPs and the survival motor neuron (SMN) protein. This increase in the number of CBs closely correlates with the activation of transcription. Moreover, the occurrence of CBs is significantly higher in BrdU (+) cells than in BrdU (-) cells, indicating that entry in the S phase promotes the formation of CBs. During the S phase, Schwann cell nuclei display higher Cdk2 nuclear staining and concentrate this kinase in CBs. Forskolin also induces a redistribution of the pre-mRNA splicing factors in Schwann cells. Primary cultures of Schwann cells provide an excellent physiological model to demonstrate that the assembly of CBs is a transcription- and replication-dependent cellular event. Moreover, the S phase accumulation of Cdk2 observed in Schwann cells supports a functional link between CBs and DNA replication, which is mediated by the possible participation of CBs in the regulatory control of histone gene expression.

[A new class of small RNP (alpha-RNP) containing antisense RNA in K-562 cells. III. The DNA-binding activity of nuclear alpha-RNP. The Signal Transmission from Growth Factors Group of the Medical Faculty, F. Schiller University, Jena, Germany]. / Novyi klass malykh RNP (al'fa-RNP), soderzhashchikh antismyslovuiu RNK, v kletkakh linii K-562. III. DNK-sviazyvaiushchaia aktivnost' iadernykh al'fa-RNP. Gruppa "Peredacha signala s rostovykh faktorov" pri Meditsinskom fakul'tete Universiteta im. F. Shillera, Iena, Germaniia.

DNA-binding activity of small nuclear alpha-RNP identified in acid-soluble fraction of chromatin of human proerythroleukemic cell line K-562 was studied using the technique of gel retardation. We found that nuclear alpha-RNP isolated from K-562 cells through treatment with dimethylsulfoxide, an agent inducing differentiation, acquire a capacity to specific interaction with Alu repeats of DNA leading to the formation of alpha-RNP-Alu-DNA complexes; nuclear alpha-RNP from cells that were not treated with dimethylsulfoxide do not show such capacity, although they are tightly bound with chromatin in the cell. Thus, the capacity of nuclear alpha-RNP to direct interaction with DNA Alu repeats appearing after the induction of K-562 cells to differentiation along erythroid pathway is an inducible property. We discuss hypothesis about the involvement of nuclear alpha-RNP in the control of expression of inducible genes at the level of chromatin and interaction with DNA.

[A new class of small RNP (alpha-RNP) containing antisense RNA in K-562 cells. IV. The coordinated regulation of the expression of Alu-containing mRNA and alpha-RNA during differentiation]. / Novyi klass malykh RNP (al'fa-RNP), soderzhashchikh antismyslovuiu RNK, v kletkakh linii K-562. IV. Koordinirovannaia reguliatsiia ékspressii Alu-soderzhashchikh mPHK i al'fa-RNK v khode differentsirovki.

Small alpha-RNP of K-562 cells contain a small RNA as an RNA component, this RNA is homologous to Alu-repeating sequences of human DNA. When cells are exposed to dimethylsulfoxide, an agent inducing cell differentiation along the erythroid pathway, the content of both high-molecular-weight (heterogeneous nuclear and messenger) RNA enriched with Alu repeats and low-molecular-weight specific RNA, small Alu-homologous alpha-RNA undergoes a coordinated decrease. Using the technique of northern blot hybridization, we have demonstrated nonuniform distribution of Alu repeats both in the fraction of total low-molecular-weight RNA of the cytoplasm as well as in the fraction of messenger RNA. It is proposed that alpha-RNA (alpha-RNP) participates in the control of expression of non-linked Alu-containing genes.

[A new class of small RNP (alpha-RNP) containing antisense RNA in K-562 cells. I. Their characteristics and changes during erythroid differentiation]. / Novyi klass malykh RNP (al'fa-RNO), soderzhashchokh antismyslovuiu RNK, v kletkakh linii K-562. I. Kharakteristika i izmeneniia pri differentsirovke po éritroidnomu puti.

A new class of small RNP (alpha-RNP) has been detected and identified in nuclei and cytoplasm of A-562 erythroid leukemia cell line; these RNPs have a characteristic spectrum of proteins containing conservative and specific components and a special RNA component, which contains a small antisense component (alpha-RNA), a homolog of short dispersed Alu repeats. alpha-RNP is highly stable, tightly associated with chromatin in the nucleus, and is found in the free state in cytoplasm. The composition of nuclear and cytoplasmic alpha-RNP differ and have a specific pattern of changes in response to dimethylsulfoxide, an agent causing differentiation.

[A new class of small RNP (alpha-RNP) containing antisense RNA in K-562 cells. II. The interaction of the RNA--the alpha-RNP component--with heterogeneous nuclear and messenger RNA in the normal state and under DMSO exposure]. / Novyi klass malykh RNP (al'fa-RNP), soderzhashchikh antismyslovuiu RNK, v kletkakh linii K-562. II. Vzaimodeistvie RNK--komponenta al'fa-RNP--s geterogennymi iadernymi i informatsionnymi RNK v norme i pod vozdeistviem DMSO.

Small antisense RNA (alpha-RNA), components of a new class of small nuclear and cytoplasmic RNP (alpha-RNP) identified in the cells of K-562 human proerythroleukemia cell line, are capable of hybridizing under stringent conditions with precursors of mRNA (heterogeneous nuclear RNA or mRNA) and with mRNA of these cells. We found that DMSO, an agent inducing differentiation in K-562 cells, is capable of regulating the composition of alpha-RNA population and concomitantly changes the content of mRNA that has regions homologous (complementary) to alpha-RNA. Specifically, it has been demonstrated that DMSO decreases the level of alpha-RNA, which hybridizes with the actin gene. Results of restriction mapping of regions of complementary interaction of alpha-RNA with the actin gene point out that alpha-RNA hybridizes with regions containing the promotor area and 3'-nontranslated area of the gene. It is proposed that small antisense alpha-RNA (alpha-RNP) participates in the control of gene expression at posttranscriptional level in cell cytoplasm.

Relationship of nucleolus-associated bodies with the nucleolar organizer tracks in plant interphase nuclei (Pisum sativum).

Nucleolus-associated bodies (NABs) have long been noted in interphase nuclei of a wide variety of plant species. We have recently shown that these bodies consist largely of snRNPs and that they are located on the nucleolar surface in the immediate vicinity of the nucleolar organizer tracks. The present study revealed that, following exposure of roots to KCN, an agent that induces nucleolar segregation, NABs were intimately associated with intranucleolar chromatin. Although immunocytochemical tests with anti-DNA indicated that NABs contained no demonstrable amounts of DNA, our observations nevertheless add further support to the notion that these bodies are somehow related to the nucleolar chromosomes.

[The epidermal growth factor induces specific changes in the expression of small RNA and of the set of small RNP in A-431 cells]. / Epidermal'nyi faktor rosta vyzyvaet spetsificheskie izmeneniia ékspressii malykh RNK i nabora belkov malykh RNP v kletkakh A-431.

Specific small ribonucleoprotein (alpha-RNP) complexes have been identified and characterized in the human epidermal carcinoma A-431 cells. The alpha-RNP complexes contain Alu-homologous small RNA, along with other small antisense RNA species. The epidermal growth factor (EGF) has been shown to induce selective specific changes in the expression of the small alpha-RNAs, the expression of the Alu-like RNA being repressed. Specific changes in the protein composition of the alpha-RNP complexes have been detected under the influence of EGF.

A herpes simplex virus type 1 immediate-early gene product, IE63, regulates small nuclear ribonucleoprotein distribution.

Herpes simplex virus 1 (HSV-1), a nuclear replicating DNA virus, has 73 identified genes of which only 4 contain introns. For this reason the virus probably makes only minimal use of the cellular RNA-splicing machinery. Antigens associated with the small nuclear ribonucleoprotein particles (snRNPs) that are subunits of splicing complexes have been reported to redistribute in the nucleus and become concentrated into the intranuclear structures, the interchromatin granules, after HSV-1 infection [Martin, T. E., Barghusen, S. C., Leser, G. P. & Spear, P. G. (1987) J. Cell Biol. 105, 2069-2082]. We observe this snRNP redistribution upon HSV-1 infection, in which the widespread snRNP staining pattern changes to a restricted punctate distribution with a concomitant loss of coiled bodies in HSV-1-infected cells. We show here that expression of the immediate-early (IE) subset of HSV-1 genes is necessary and sufficient for snRNP redistribution. Using a series of HSV-1 mutants in different IE genes, we have established that specifically the product of the viral IE63 (ICP27) gene is essential for this effect, and transfection experiments revealed that IE63 expression alone can cause the snRNP redistribution. Further, we show that the IE63 gene product colocalizes with the redistributed snRNP in the nucleus. The snRNP redistribution caused by HSV-1 infection resembles the effect seen after inhibition of transcription in uninfected cells. In HSV-1-infected cells, however, the snRNP redistribution is under the control of viral IE gene products and occurs during active virus gene transcription.