[The perichromatin compartment of the cell nucleus].

In this review, the data on the structure and composition of the perichromatin compartment, a special border area between the condensed chromatin and the interchromatin space of the cell nucleus, are discussed in the light of the concept of nuclear functions in complex nuclear architectonics. Morphological features, molecular composition and functions of main extrachromosomal structures of the perichromatin compartment, perichromatin fibrils (PFs) and perichromatin granules (PGs) including nuclear stress-bodies (nSBs) that are derivates of the PGs under heat shock, are presented. A special attention was paid to the features of the molecular compositions of PFs and PGs in different cell types and at different physiological conditions.

[Karyosphere capsule in Tribolium castaneum oocytes].

Structure and composition of the karyosphere (karyosome) capsule were studied in the oocytes of a laboratory insect, Tribolium castaneum, with the use of electron microscopy and immunoelectron cytochemistry. Basing on the study of nuclear structure dynamics, we distinguished 8 stages that characterize the period of oocyte growth. At the diplotene stage, T. castaneum oocyte chromosomes conjoin early into a compact karyosphere, but a significant chromatin condensation does not occur. The process of karyosphere formation is accompanied by the development of an extensive extrachromosome capsule surrounding chromatin. The capsule consists of a material of different morphological types. Significant molecular components of the T. castaneum karyosphere capsule are represented by the proteins of nuclear matrix including F-actin and lamin B. Besides the structural proteins, the Sm proteins of small nuclear (sn) RNPs and mature 2,2,7-trimethyl guanosine (TMG) 5'-capped snRNAs are revealed immunocytochemically in the karyosphere capsule. The obtained data can form a basis for further expansion of ideas on the functions of the karyosphere capsule as a specialized extrachromosomal nuclear domain of the oocytes. We believe that the T. castaneum karyosphere capsule plays not only a structural role, but may be involved directly in the processes related to gene expression.

[Polyadenylated RNA and mRNA export factors in extrachromosomal nuclear domains of vitellogenic oocytes of the insect Tenebrio molitor].

The nucleus ofvitellogenic oocytes of the yellow mealworm, Tenebrio molitor, contains a karyosphere that consists of the condensed chromatin embedded in an extrachromosomal fibrogranular material. Numerous nuclear bodies located freely in the nucleoplasm are also observed. Amongst these bodies, counterparts of nuclear speckles (= interchromatin granule clusters, IGCs) can be identified by the presence of the marker protein SC35. Microinjections of fluorescently tagged methyloligoribonucleotide probes 2'-O-Me(U)22, complementary to poly(A) tails of RNAs, revealed poly(A)+ RNA in the vast majority of IGCs. We found that all T. molitor oocyte IGCs contain heterogeneous ribonucleoprotein (hnRNP) core protein Al that localizes to IGCs in an RNA-dependent manner. The extrachromosomal material of the karyosphere and a part of nucleoplasmic IGCs also contain the adapter protein Aly that is known to provide a link between pre-mRNA splicing and mRNA export. The essential mRNA export factor/receptor NXF1 was observed to colocalize with Aly. In nucleoplasmic IGCs, NXF1 was found to localize in an RNA-dependent manner whereas it is RNA-independently located in the extrachromosomal material of the karyosphere. We believe our data suggest on a role of the nucleoplasmic IGCs in mRNA biogenesis and retention in a road to nuclear export.

[Interchromatin granule clusters of mouse preovulatory oocytes, organization, molecular composition and possible functions].

At the diplotene stage of meiotic prophase, the nucleus of mouse preovulatory oocytes contains multiple interchromatin granule clusters (IGC). These nuclear compartments are universal and evolutionary conserved and enriched in pre-mRNA splicing factors. Nowadays, IGCs are believed to play an important role in gene expression events and contain different molecular components that allow coupling of many processes from transcription to mRNA export. We obtained the data on the distributions of poly(A)+RNA, hnRNPS A/B, and NXF1/TAP factor of mRNA export. These factors were found to associate with IGCs of mouse preovulatory oocytes. In the present study, we have demonstrated for the first time the dynamics of large IGCs after specific phosphorilation of SR-proteins with okadaic acid, an inhibitor of protein phosphatases. Using electron microscopy, conventional fluorescent and confocal microscopies, as well as microinjections of olygonucleotide probes in mouse oocytes, some features of structural organization and molecular compositions of IGCs in the nuclei of mouse oocyte from antral follicles were established. Possible roles of IGCs in pre-mRNA metabolism and the participation of these structures in mRNA export are discussed.

[SC35 splicing factor and coilin are colocalized within the "endobodies" in oocytes of the spider Araneus diadematus].

Oocytes of the spider Araneus diadematus are solitary developed. The oocyte nuclei are transcriptionally active, as revealed using microinjections of 5-bromouridine 5'-triphosphate into the ooplasm. The nucleus contains several extrachromosomal structures of perfectly spherical shape, the so-called endobodies. Laser scanning confocal microscopy revealed intense fluorescence of endobodies when an antibody against the splicing factor SC35, a major component of interchromatin granule clusters (IGCs), was applied. At the same time, colocalization of SC35 protein with coilin, a marker protein of Cajal bodies (CBs), was also observed within these structure. We suggest that endobodies of A. diadematus oocytes may have some features of both IGCs and CBs and represent a single nuclear domain.

[Oogenesis in turbellarians of the genus Geocentrophora (Lecithoepitheliata, Prorhynchidae) studied by light and electron microscopy. VI. Immunogold labeling localization of some components essential for RNA polymerase II transcription in perichromatin regions of G. baltica oocytes].

The female gonad (germovitellarium) of the lecithoepitheliate turbellarians has a special type of organization. It consists of the peculiar follicles in which the oocytes develop surrounded by the yolk cells (vitellocytes). While an oocyte grows, chromatin remains to be decondensed, and the karyosphere is not formed. Using immunogold labeling microscopy we studied a distribution of RNA polymerase II, transcriptional coactivators CBP/p300, and TATA-box binding protein in the perichromatin regions of Geocentrophora baltica oocytes. We found these components of RNA polymerase II transcription to be associated with the perichromatin fibrils (PFs). We suggest that the presence of numerous PFs in the perichromatin regions of G. baltica oocytes reflects the active state of the nucleus during the unusual alimentary oogenesis of Lecithoepitheliata.

[Cajal bodies in insect oocytes. II. New data on the molecular composition of cajal bodies in oocytes of the house cricket Acheta domesticus with special reference to interactions between cajal bodies and interchromatin granule clusters].

Cajal bodies (CBs) in oocytes of the house cricket Acheta domesticus are large, perfectly spherical nuclear organelles with a complex internal structure. These consist of a fibrillar coilin-containing matrix and a central cavity with a prominent fibrogranular body inside; the latter has been referred to as an "internal" interchromatin granule cluster (IGC). Within the matrix of CBs we detected transcriptional co-activators CBP/p300 and TATA-box binding protein (TBP). No RNA polymerase II was revealed in CBs of both normal and actynomycin D treated oocytes. In the nucleoplasm of A. domesticus oocytes, besides CBs, free IGCs were observed. In oocytes treated with actynomycin D, the amount of "free" IGCs in the nucleoplasm increase significantly, granular and fibrillar components of IGCs were seen segregated, and RNA polymerase II and CBP/p300 were found to be accumulated in fibrillar zones of IGCs.

[Karyosphere and extrachromosomal nuclear bodies in oocytes of the scorpionfly, Panorpa communis].

Oocyte nuclear structures were studied for the scorpionfly Panorpa communis at different stages of oocyte growth, from pachytene to the first meiotic division. Using immunofluorescent and immunogold microscopy, we analyzed the nuclear distribution of RNA polymerase II, splicing factors and coilin. These factors were revealed in close association with perichromatin fibrils and, later, with some elements of the karyosphere and extrachromosomal nuclear bodies (NBs). Besides, it was shown that large amounts of P. communis oocyte NBs represent Cajal bodies (CBs) and contain CB marker protein, coilin, as well as RNA polymerase II, and in some cases an essential splicing factor, SC35. The presence of SC35 is commonly not characteristic of CBs in somatic cells. CB dynamics was traced in inactivated oocyte nuclei, during a gradual condensation of chromosomes and their final assembling into the karyosphere. It has been shown that coilin, RNA polymerase II and SC35 protein are common compounds shared by CBs and some granular material associated with these condensed chromosomes. CB remnants were demonstrated in the ooplasm after the breakdown of nuclear envelope before the first meiotic division. In inactivated oocyte nuclei, CBs serve presumably as storage compartments for some inactive components essential for gene expression.

[RNA polymerase II and pre-mRNA splicing factors in diplotene oocyte nuclei of the giant African gastropod Achatina fulica]. / RNK-polimeraza II i faktory splaisinga pre-mRNK v iadrakh diplotennykh ootsitov gigantskoi afrikanskoi ulitki Acnatina fulica.

The nuclear distribution of pre-mRNA splicing factors (snRNPs and SR-protein SC35) and unphosphorylated from of RNA polymerase II (Pol II) was studied using fluorescent and immunoelectron cytochemistry in diplotene oocytes of the gastropod Achatina fulica. Association of Pol II and splicing factors with oocyte nuclear structures was analysed. The antibodies against splicing factors and Pol II were shown to label perichromatin fibrils at the periphery of condensed chromatin blocks as well as those in interchromatin regions of nucleoplasm. The revealed character of distribution of snRNPs, SC35 protein, and Pol II, together with the decondensed chromatin and absence of karyosphere, enable us to suggest that oocyte chromosomes maintain their transcriptional activity at the diplotene stage of oogenesis. In A. fulica oocytes, sparse nuclear bodies (NBs) of a complex morphological structure were revealed. These NBs contain snRNPs rather than SC35 protein. NBs are associated with a fibrogranular material (FGM), which contains SC35 protein. No snRNPs were revealed in this material. Homology of A. fulica oocyte nuclear structures to Cajal bodies and interchromatin granule clusters is discussed.

[Cajal bodies in insect oocytes. I. Identification and immunocytochemical characteristics of Cajal bodies in vitellogenic oocytes of the yellow mealworm beetle]. / Tel'tsa Kakhala v ootsitakh nasekomykh. 1. Identifikatsiia i immunotsitochimicheskaia kharakteristika telest Kakhala v vitellogennykh ootsitakh zhuka-chernotelki.

In vitellogenic oocytes of Tenebrio molitor (inactive stage), numerous fibrogranular nuclear bodies (NBs) are present. Using immunofluorescent microscopy, these NBs were shown to contain pre-mRNA splicing factors (small nuclear [sn] RNPs and SR-protein, SC35) as well as RNA polymerase II. A limited set of NBs also contained coilin, a marker protein for Cajal bodies (CBs). We suggest that in T. molitor oocytes, coilin-containing NBs, which also contain splicing factors and RNA polymerase II, seem to represent CBs. In the species studied, no morphological features of CBs were established as compared with other NBs, which do not contain coilin. Microinjectons in oocytes of myc-tagged coilin mRNA, followed by revealing newly translated protein with antibody specific for this tag, have shown targeting of myc-coilin with CBs. The own and literary data on the morphology and molecular composition of CBs are discussed in terms of searching for criteria for CB identification in cells of different origin, and at active and inactive stages.

[Extrachromosomal structures containing small nuclear RNP and coilin in the late vitellogenic oocytes of hibernating grass frogs]. / Ekstrakhromosomnye struktury, soderzhashchie malye iadernye RNP i koilin, v iadrakh pozdnikh vitellogennykh ootsitov zimuiushchikh travianykh liagushek.

We have studied extrachromosomal structures in the germinal vesicle (GV) of the late vitellogenic oocytes of hibernating frogs Rana temporaria. During this period of oogenesis, chromosomes are completely inactivated to be surrounded by a fibrillar karyosphere capsule (Gruzova, Parfenov, 1993). Using immunostaining and in situ nucleic acid hybridization, we have identified three types of extrachromosomal structures: Cajal bodies (CB), nucleoli, and micronucleoli. Immunostaining of GV spreads has revealed that CB and nucleoli contain coilin, a marker protein for CB. The nucleoli were also positively stained with antibodies against Sm-epitope and trimetylguanosine cap of small nuclear (sn) RNP. According to the results of in situ nucleic acid hybridization, the nucleoli contain U6 snRNA. To further investigate a distribution of coilin in GV of the late vitellogenic oocytes of R. temporaria, we injected myc-tagged transcripts of full length human coilin (Wu et al., 1994) into the cytoplasm of oocytes and followed the pathway of the newly translated protein with an antibody specific for the tag. Immunofluorescent staining of spread GV contents demonstrated a specific staining of the nucleoli within 3 h after injection. We suggest that the newly synthesized myc-coilin may be phosphorilated and targeted to the nucleoli.

[Oogenesis in turbellarians of the genus Geocentrophora studied by light and electron microscopy. V. Oocyte nuclear structures containing factors of pre-mRNA splicing and pre-rRNA processing]. / Oogenez tubelliarii roda Geocentrophora po dannym svetovoi i élektronnoi mikroskopii V.Iadernye struktury ootsitov, soderzhashchie faktory splaisinga pre-mRNK i protsessinga pre-rRNK.

An immunoelectron study of nuclear distribution of pre-mRNA splicing and pre-rRNA processing factors was carried out for oocytes of two turbellarian species: the Baikal endemic Geocentrophora wagini and a cosmopolitan G. baltica. Using monoclonal antibodies against Sm-epitope of small nuclear RNPs (snRNPs) and SR-protein SC35, it has been shown that on different stages of oocyte growth splicing factors (snRNPs and SC35) are distributed within the whole nucleus. A fibrogranular material located near heterochromatin clumps is labeled with these antibodies. A fibrillar part of this material seems to represent perichromatin fibrils. The features of intranuclear distribution of splicing factors in Geocentrophora oocyte nuclei and their ultrastructural features suggest that pre-mRNA synthesis and splicing may occur up to the end of diplotene. In Geocentrophora oocyte nuclei a few nuclear bodies (NBs) were found. Splicing factors (snRNPs and SC35) and fibrillarin were revealed in these NBs. Homology of Geocentrophora oocyte NBs to coiled bodies of oocyte and somatic cell nuclei of other animals is discussed. During diplotene, Geocentrophora oocyte nucleoli were found to lose their granular component and to change to large fibrillar structures named "postnucleoli". The postnucleoli contain both fibrillarin and non-nucleolar spliceosomal components (snRNPs and SC35). Geocentrophora oocyte postnucleoi are compared with similar structures of mammalian oocyte nuclei, taken as an example of morphological convergence of nuclear structure organization in phylogenetically distant animal species.