The characterization of free,cytoskeletal and membrane-bound polysomes in Krebs II ascites and 3T3 cells

Summary Polysomes from Krebs II ascites and 3T3 cells were separated into three populations by using a sequential extraction method. Free polysomes were released by using a combination of low salt (25 mM KCl) and NP-40 detergent in the lysis buffer. The cytoskeletal bound polysomes were subsequently released by raising the salt concentration to 130 mM and finally, polysomes bound to the membranes of the endoplasmic reticulum were extracted by the combined treatment with Triton X-100 and deoxycholate. The results presented here illustrate that the three polysome-containing fractions differ in many parameters such as polysome profiles, cytoskeletal components and phospholipid content. When polyA-containing mRNA was isolated from the three polysome fractions and translated in an in vitro system, some differences were observed in the patterns of proteins being synthesized.     

Sequential rearrangement and nuclear polymerization of actin in baculovirus-infected Spodoptera frugiperda cells.

Proper assembly of nucleocapsids of the baculovirus Autographa californica nuclear polyhedrosis virus is prevented by cytochalasin D, a drug that interferes with actin microfilament function. To investigate the involvement of microfilaments in A. californica nuclear polyhedrosis virus replication, a fluorescence microscopy study was conducted that correlated changes in distribution of microfilaments with events in the life cycle of the virus. Tetramethylrhodamine isothiocyanate-labeled phalloidin was used to label microfilaments, and monoclonal antibody was used to label p39, the major viral capsid protein. Three microfilament arrangements were found in infected cells. During uptake of virus, thick cables were formed. These were insensitive to cycloheximide, indicating that this configuration was a rearrangement of preexisting cellular actin mediated by a component of the viral inoculum. At the time of cell rounding and before viral DNA replication, ventral aggregates of actin were observed. These were sensitive to cycloheximide but not to aphidicolin, indicating that an early viral gene mediated this actin rearrangement. Ventral aggregates did not result from the rounding process itself. Uninfected cells prerounded with colchicine did not form ventral aggregates. Cells prerounded with colchicine and then infected did form aggregates. At the time of exponential production of progency virus, microfilaments were found in the nucleus surrounding the virogenic stroma. In this area (where nucleocapsid assembly is known to take place) microfilaments colocalized with p39. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western immunoblot analysis identified p39 among proteins retained on an f-actin affinity column. We postulate that microfilaments in the nucleus provide a scaffold to position capsids for proper assembly and filling with DNA.     

Phalloidin shift on velocity sedimentation sucrose gradient centrifugation for identification of microfilament-associated proteins

Velocity sedimentation by sucrose density gradient centrifugation has been used to characterize ascites microvillar microfilament cores and to identify microfilament-associated proteins. Fluoride, calcium, phalloidin and chemical cross-linking treatments of microvilli during Triton X-100 extractions increase the sedimentation rate of the microfilament core, compared with untreated control samples. Electrophoretic analyses of the distributions of actin, alpha-actinin and other microfilament-associated proteins across the gradients indicate that the primary mechanism for stabilization of the microfilament core is the reduction of fragmentation of the microfilaments. Significantly, alpha-actinin could be completely removed from the microfilaments by calcium treatment without causing a decrease in the size of the microfilament core. Because of the specificity of phalloidin in the stabilization of microfilaments, the shift on the gradients of microfilaments and their associated proteins in the presence of phalloidin provides a diagnostic tool for the identification of microfilament-associated proteins. This phalloidin shift technique should have widespread utility in the analysis of actin forms and microfilament-associated proteins from complex cell fractions.     

Selective extraction of cotton fiber cytoplasts to identify cytoskeletal-associated proteins

Cytoplasts from cotton (Gossypium hirsutum L.) fiber cells retain microtubule and microfilament cytoskeletons through extraction with non-ionic detergent and ethylene glycol bis-(β-aminoethyl ether) N,N,N',N'-tetraacetic acid. Tubulin and actin are the most abundant proteins in extracted cytoplasts; however, many other less abundant proteins are also present. To determine if minor proteins were associated with the cytoskeleton, microtubules and microfilaments were selectively removed from extracted cytoplasts by detergent extraction in an alkaline Ca²⁺ solution. Under these extraction conditions, microtubules and microfilaments were fragmented and depolymerized unless previously stabilized by taxol and phalloidin. Associated proteins were identified by their loss in conjunction with either microtubules or microfilaments. As judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, one protein, of roughly 115 kDa, appeared to be associated with microfilaments since it was present in Ca²⁺-extracted preparations only when microfilaments were stabilized with phalloidin. The failure of most minor proteins to associate with microtubules and microfilaments suggests that caution must be used when interpreting co-isolation as evidence for an association of low abundance proteins with cytoskeletons.     

Identification of the major membrane and core proteins of vaccinia virus by two-dimensional electrophoresis.

Vaccinia virus assembly has been well studied at the ultrastructural level, but little is known about the molecular events that occur during that process. Towards this goal, we have identified the major membrane and core proteins of the intracellular mature virus (IMV). Pure IMV preparations were subjected to Nonidet P-40 (NP-40) and dithiothreitol (DTT) treatment to separate the core proteins from the membrane proteins. These proteins were subsequently separated by two-dimensional (2D) gel electrophoresis, and the major polypeptide spots, as detected by silver staining and 35S labeling, were identified by either matrix-assisted laser desorption/ionization mass spectrometry, N-terminal amino acid sequencing, or immunoprecipitation with defined antibodies. Sixteen major spots that partitioned into the NP-40-DTT-soluble fraction were identified; 11 of these were previously described virally encoded proteins and 5 were cellular proteins, mostly of mitochondrial origin. The core fraction revealed four major spots of previously described core proteins, two of which were also detected in the membrane fraction. Subsequently, the NP-40-DTT-soluble and -insoluble fractions from purified virus preparations, separated by 2D gels, were compared with postnuclear supernatants of infected cells that had been metabolically labeled at late times (6 to 8 h) postinfection. This relatively short labeling period as well as the apparent shutoff of host protein synthesis allowed the selective detection in such postnuclear supernatants of virus-encoded proteins. These postnuclear supernatants were subsequently treated with Triton X-114 or with sodium carbonate to distinguish the membrane proteins from the soluble proteins. We have identified the major late membrane and nonmembrane proteins of the IMV as they occur in the virus as well as in infected cells. This 2D gel map should provide an important reference for future molecular studies of vaccinia virus morphogenesis.     

Association of maternal and newly synthesized ribosomes with membranous noncytoskeletal structures in Xenopus laevis embryonic cells

In Xenopus laevis embryos a high concentration of both KCl and 0.5% DOC (sodium deoxycholate) is needed for maximal extraction of ribosomes and polysomes. We studied the nature of the structures that keep ribosomes and polysomes immobilized within the cytoplasm of embryonic cells at cleavage through tailbud stages, using various combinations of a low-salt buffer (20 mM KCl), a high-salt buffer (500 mM KCl), 0.5% DOC, and 0.5% Triton X-100. With a low-salt buffer and 0.5% DOC, but not Triton X-100, 80S ribosomal monomers and polysomes were liberated from the cytoplasmic rapidly sedimenting structures (RSS) to the soluble fraction. With a high-salt buffer (500 mM KCl), ribosomes were solubilized as 60S and 40S subunits together with about one-half of the total polysomes. When cells were homogenized in a low-salt buffer with added inhibitors of the cytoskeleton (cytochalasin B or colchicine), the majority of polysomes but not ribosomes were solubilized. These results provide evidence for the following conclusions. 1) Polysomes are bound to cytoskeletal structures in Xenopus embryos, but ribosomes, both maternal and newly synthesized, are associated with membranous noncytoskeletal structures. 2) The membranous structures consist of two compartments, one high-salt sensitive and the other high-salt resistant. 3) Ribosomes of the high-salt resistant group increase in amount with developmental stage and appear to be the precursor to the ribosomes of the high-salt sensitive group.     

Preparation and purification of polymerized actin from sea urchin egg extracts

Isotonic extracts of the soluble cytoplasmic proteins of sea urchin eggs, containing sufficient EGTA to reduce the calcium concentration to low levels, form a dense gel on warming to 35-40 degrees C. Although this procedure is similar to that used to polymerize tubulin from mammalian brain, sodium dodecyl sulfate-polyacrylamide gel electrophoresis shows this gel to have actin as a major component and to contain no tubulin. If such extracts are dialyzed against dilute salt solution, they no longer respond to warming, but gelation will occur if they are supplemented with 1 mM ATP and 0.020 M KCl before heating. Gelation is not temperature reversible, but the gelled material can be dissolved in 0.6-1 M KCl and these solutions contain F- actin filaments. These filaments slowly aggregate to microscopic, birefringent fibrils when 1 mM ATP is added to the solution, and this procedure provides a simple method for preparing purified actin. the supernate remaining after actin removal contains the other two components of the gel, proteins of approximately 58,000 and 220,000 mol wt. These two proteins plus actin recombine to form the original gel material when the ionic strength is reduced. This reaction is reversible at 0 degrees C, and no heating is required.     

A cytoskeleton-associated plasma membrane heparan sulfate proteoglycan in Schwann cells

Schwann cells cocultured with sensory neurons in a serum-free medium accumulate a single species of radiolabeled heparan sulfate proteoglycan (HS-PG) during incubation in medium containing 35SO4. This HS-PG was poorly extracted from cultures by solutions containing 1% Triton X-100 in low salt buffer or by solutions containing 1 M KCl, 4 M urea plus dithiothreitol, 1 mM Tris-HCl, 5 mM EDTA, or 100 micrograms/ml of heparin. The HS-PG was efficiently extracted, however, by 1% Triton X-100 in the presence of 1 M KCl or by 1% deoxycholate. These treatments solubilize both cell membranes and the Schwann cell cytoskeleton. In intact cells the HS-PG was digested by trypsin, indicating it was at least partially exposed on the cell surface. When solubilized HS-PG was applied to a column of octyl-sepharose CL-4B, more than 90% was retained by the column, but was quantitatively eluted by a solution containing 1% Triton X-100. In addition, the solubilized HS-PG could be incorporated into artificial phospholipid vesicles. These results indicate the HS-PG is an integral plasma membrane protein. The inability of low ionic strength solutions containing Triton X-100 to solubilize the HS-PG suggested it was bound to an additional structure. To determine whether the HS-PG was associated with the cytoskeleton we isolated cytoskeletons by detergent lysis of cells and centrifugation. The major protein components of isolated cytoskeletons were spectrin (Mr 225,000), vimentin (Mr 58,000), and actin (Mr 45,000). When 35SO4-labeled cells were used to prepare cytoskeletons approximately 80% of the total HS-PG was recovered in the cytoskeleton fraction. These results suggest the HS-PG is an externally exposed integral plasma membrane protein that is anchored to the Schwann cell cytoskeleton.     

Supramolecular structures in mycoplasmas

Mycoplasma pneumoniae cells treated with Triton X-100 showed a detergent-resistant cytoskeleton. This cytoskeleton consists of microfilaments which seem related to eukaryotic actin filaments, both morphologically and in some chemical properties, including specific staining by anti-actin antibodies and rhodamine-labeled phalloidin. The degree of homology, however, is still unclear. In motile cells the filaments form an irregular network in the cytoplasm of the cell body and a bundle in the frontal projection corresponding to the leading edge of the gliding cells. This particular arrangement may reflect different functions. The microfilaments could be isolated by differential centrifugation. Analysis of the microfilament fraction by SDS gel electrophoresis revealed five major polypeptide bands. One of these proteins, with a molecular weight of 42.5 kd, co-migrated with rabbit muscle actin. No filaments could be found in a nonmotile mutant, M-22.     

Monomer-polymer equilibria in the axon: direct measurement of tubulin and actin as polymer and monomer in axoplasm

The monomer-polymer equilibria for tubulin and actin were analyzed for the cytoskeleton of the squid giant axon. Two methods were evaluated for measuring the concentrations of monomer, soluble (equilibrium) polymer, and stable polymer in extruded axoplasm. One method, the Kinetic Equilibration Paradigm ( KEP ), employs the basic principles of diffusion to distinguish freely diffusible monomer from proteins that are present in the form of polymer. The other method is pharmacological and employs either taxol or phalloidin to stabilize the microtubules and microfilaments, respectively. The results of the two methods agree and demonstrate that 22-36% of the tubulin and 41-47% of the actin are monomeric. The in vivo concentration of monomeric actin and tubulin were two to three times higher than the concentration required to polymerize these proteins in vitro, suggesting that assembly of these proteins is regulated by additional mechanisms in the axon. A significant fraction of the polymerized actin and tubulin in the axoplasm was stable microtubules and microfilaments, which suggests that the dissociation reaction is blocked at both ends of these polymers. These results are discussed in relationship to the axonal transport of the cytoskeleton and with regard to the ability of axons to change their shape in response to environmental stimuli.     

Fractionation of micrococcal nuclease-digested chromatin solubilized at physiologic ionic strength

When mouse brain nuclei are optimally digested with micrococcal nuclease, most of the chromatin is soluble in a 180 mM salt/1 mM EDTA buffer [1]. At this ionic concentration, chromatin maintains its native structure [2]. In an attempt to selectively extract different fractions of chromatin from digested nuclei, we have examined the differential solubility of chromatin in the 180 mM salt buffer containing concentrations of MgCl2 ranging from 2 to 0 mM. The results suggest that digested chromatin may be fractionated into specific soluble chromatin fractions which correspond to nuclease-sensitive chromatin, bulk chromatin, and heterochromatin. These soluble fractions have a high molecular weight (up to 20 kbp), and contain a full complement of histones as well as a complex assortment of non-histone proteins. The residual insoluble fraction may be equivalent to a native, nuclear matrix-bound chromatin fraction.     

Translation and the cytoskeleton: a mechanism for targeted protein synthesis

This review describes the critical evidence that in eukaryotic cells polyribosomes, mRNAs and components of the protein synthetic machinery are associated with the cytoskeleton. The role of microtubules, intermediate filaments and microfilaments are discussed; at present most evidence suggests that polyribosomes interact with the actin filaments. The use of non-ionic detergent/deoxycholate treatment in the isolation of cytoskeletal-bound polysomes is described and the conclusion reached that at low salt concentrations this leads to mixed preparations of polysomes derived from both the cytoskeleton and the endoplasmic reticulum. At present the best approach for isolation of cytoskeletal-bound polysomes appears to involve extraction with salt concentrations greater than 130 mM after an initial non-ionic detergent treatment. Such polysomes appear to be enriched in certain mRNAs and thus it is suggested that they are involved in translation of a unique set of proteins. The evidence for mRNA localisation is presented and the role of the cytoskeleton in transport and localisation of RNA discussed. Recent data on the role of the 3 untranslated region in the targeting of mRNAs both to particular regions of the cell and for translation on cytoskeletal-bound polysomes is described. The hypothesis is developed that the association of polysomes with the cytoskeleton is the basis of a mechanism for the targeting of mRNAs and the compartmentalization of protein synthesis.Abbreviations CBP cytoskeletal-bound polysomes - FP free polysomes - MBP membrane-bound polysomes - ER endoplasmic reticulum     

Alterations in the cell surface proteins of Drosophila during morphogenesis

Summary Unevaginated and evaginated Drosophila imaginal discs were surface-labeled with ¹²⁵I. Relative labeling was greater in eleven peptides and lower in three peptides of evaginated discs compared to unevaginated discs. These results are compared to the effects of 20-hydroxyecdysone (20-HOE) on metabolic labeling of membrane proteins fractionated from imaginal discs, and on cell surface labeling of a hormone-responsive Drosophila tissue culture line. A group of ³⁵S-methionine labeled membrane fraction peptides whose metabolic labeling is 20-HOE dependent have isoelectric points and apparent molecular weights very similar to those of a group of proteins only labeled in iodinated evaginated discs, supporting the conclusion that these are hormone-dependent, cell surface proteins (Rickoll and Fristrom 1983). Based upon two-dimensional gel electrophoretic and immunological criteria three of the proteins showing increased labeling in evaginated discs are related to three proteins induced by 20-HOE in tissue culture cells. Two different subsets of radiolabeled peptides were observed in the imaginal discs based upon detergent solubility. Some of the proteins which are soluble in NP-40 plus urea but insoluble in NP-40 alone may be localized in the basal lamina of the imaginal discs, a structure which labels heavily with ¹²⁵I and is lacking in tissue culture cells. In discs, the majority of hormone-dependent changes in radiolabeled peptides were seen in the fraction solubilized by NP-40 and urea with a sulfhydryl reducing agent, while in tissue culture cells, the majority of differences is seen in the fraction solubilized by NP-40 only. We speculate that these proteins may be involved in similar processes, e.g., cell rearrangement, that occur during both disc morphogenesis and 20-HOE induced aggregation in tissue culture cells.This work was supported by grants CD-205 from the American Cancer Society, RR08132 from NIH to C.A.P. and GM 19937 from NIH to J.W.F.     

Ligand-induced association of surface immunoglobulin with the detergent insoluble cytoskeleton may involve alpha-actinin

B cell surface immunoglobulin (SIg) plays an important role in antigen recognition and cellular activation. Cross-linking of SIg by bivalent antibody converts it into a detergent insoluble state. The resultant SIg may be partially solubilized by incubating the detergent insoluble cytoskeleton in buffers that convert F actin to G actin. Immunoprecipitation of SIg from the detergent soluble fraction of [35S]methionine-labeled B cells results in the co-isolation of 112 kDa, 42 kDa, (actin), and three additional proteins in the 70- to 73-kDa molecular mass range, isoelectric point 4.8 to 5.6. Analysis of anti-Ig immunoprecipitates made after preclearing with anti-alpha-actinin showed complete depletion of the 112-kDa protein, suggesting that the 112-kDa protein is immunologically related to alpha-actinin. These immunoprecipitates also showed partial depletion of 70- to 73-kDa proteins and mu and delta heavy chains. After treatment of a rat B cells with anti-Ig, much of the Ig-associated 112-kDa protein and 70- to 73-kDa proteins became detergent insoluble, concomitant with most of the SIg. The migration of the SIg-associated 112-kDa and 70- to 73-kDa proteins from the detergent soluble fraction to the detergent insoluble fraction after ligand treatment, suggests that these proteins might be involved in linking SIg to the underlying cytoskeleton and could be involved in the transmission of mitogenic signals.     

Efficient removal of detergents from proteins and peptides in a spin column format

Detergents are commonly used in protein–chemistry protocols and may be necessary for protein extraction, solubilization, and denaturation; however, their presence interferes with many downstream analysis techniques, including mass spectrometry (MS). To enable downstream analysis, it is critical to remove unbound detergents from protein and peptide samples. In this study, we describe a high-performance resin that offers exceptional detergent removal for proteins and peptides. When used in a spin column format, this resin dramatically improves protein and peptide MS results by more than 95% removal of 1–5% detergents, including sodium dodecyl sulfate (SDS), sodium deoxycholate, Chaps, Triton X-100, Triton X-114, NP-40, Brij-35, octyl glucoside, octyl thioglucoside, and lauryl maltoside, with high recovery of proteins and peptides. Postcolumn liquid chromatography–tandem MS (LC–MS/MS) analysis of trypsin digests of bovine serum albumin (BSA) and HeLa cell lysate revealed excellent sequence coverage, indicating successful removal of detergent from the peptides. Matrix-assisted laser desorption/ionization (MALDI)–MS analysis of unprocessed and processed samples further confirmed efficient removal of detergents. The advantages of this method include speed (<15 min), efficient detergent removal, and high recovery of proteins and peptides.     

Effects of detergents on the West Nile virus protease activity

Detergents such as Triton X-100 are often used in drug discovery research to weed out small molecule promiscuous and non-specific inhibitors which act by aggregation in solution and undesirable precipitation in aqueous assay buffers. We evaluated the effects of commonly used detergents, Triton X-100, Tween-20, Nonidet-40 (NP-40), Brij-35, and CHAPS, on the enzymatic activity of West Nile virus (WNV) protease. Unexpectedly, Triton X-100, Tween-20, and NP-40 showed an enhancement of in vitro WNV protease activity from 2 to 2.5-fold depending on the detergent and its concentration. On the other hand, Brij-35, at ?0.001% enhanced the protease activity by 1.5-fold and CHAPS had the least enhancing effect. The kinetic analysis showed that the increase in protease activity by Triton X-100 was dose-dependent. Furthermore, at Triton X-100 and Tween-20 concentrations higher than 0.001%, the inhibition of compound B, one of the lead compounds against WNV protease identified in a high throughput screen (IC₅₀ value of 5.7 ± 2.5 μM), was reversed. However, in the presence of CHAPS, compound B still showed good inhibition of WNV protease. Our results, taken together, indicate that nonionic detergents, Triton X-100, Tween, and NP-40 are unsuitable for the purpose of discrimination of true versus promiscuous inhibitors of WNV protease in high throughput assays.     

Evidence that insulin increases the proportion of polysomes that are bound to the cytoskeleton in 3T3 fibroblasts

The association of polysome redistribution with changes in protein synthesis was investigated in insulin-stimulated fibroblasts. Free polysomes were released by Nonidet-P40 and 25 mM KCl, cytoskeletal-bound polysomes were retained at 25 mM KCl but released at 130 mM, while membrane-bound polysomes were released by deoxycholate. Insulin increased the proportion of polysomes which were retained at 25 mM KCl but had no effect when extraction was carried out at 130 mM KCl, suggesting that more polysomes were associated with the cytoskeleton. Insulin also reduced the amount of actin released from the detergent-insoluble cytoskeleton indicating that the hormone affects microfilament organization.     

Detection of activity and mass spectrometric identification of mouse liver carboxylesterase and aldehyde dehydrogenase separated by non-denaturing two-dimensional electrophoresis after extraction with detergents

To examine the activities and identity of enzymes associated with organelles such as microsomes and mitochondria, proteins from mouse liver were extracted using the non-ionic detergents Nonidet P-40 (NP-40), polyoxyethylene sorbitan monooleate (Tween 80), polyoxyethylene isooctylphenyl ester (Triton X), n-octyl beta-D-glucoside (octyl glycoside) or anionic detergent sodium dodecylsulfate (SDS) after the removal of cytosolic proteins. The proteins extracted by detergents were separated by non-denaturing two-dimensional electrophoresis (2-DE). The activities of esterase and aldehyde dehydrogenase were retained by non-denaturing 2-DE after treatment with each non-ionic detergent, but the activities were reduced or lost when the proteins were extracted with more than 0.5% SDS. For proteomic analysis of the organelle-associated proteins in mouse liver, proteins were separated by non-denaturing 2-DE and were identified using electrospray ionization tandem mass spectrometry (ESI-MS/MS) after the proteins were solubilized by octyl glycoside, NP-40 and 0.1% SDS. Several organelle-associated proteins such as carboxylesterase, aldehyde dehydrogenase, glucose regulated protein and HSP60 were identified. These results indicate that the activities and identity of detergent-soluble enzymes can be examined by this non-denaturing 2-DE and mass spectrometry.     

Peripheral nuclear matrix actin forms perinuclear shells

Perinuclear actin shells have been reported in a variety of organisms. The shells have been identified by staining perinuclear material with fluorescently-labelled phalloidin, but have not been localized to a specific subcellular compartment at the ultrastructural level. We show here that the shells of 3T3 cells lie in the peripheral nuclear matrix. Nuclear shells and matrix actin in other parts of the nucleus are not usually detected by immunohistochemical staining because they are inaccessible to antibodies or to phalloidin. Immunohistochemical detection of nuclear actin is only possible during its deposition at the end of mitosis, or in interphase nuclei that have been extracted with detergent, digested with nucleases and washed with high salt buffers. J. Cell. Biochem. 70:240–251, 1998. © 1998 Wiley-Liss, Inc.     

Intranuclear actin microfilaments in the oocytes of the common frog

For identification and distribution of actin microfilaments in hand-isolated nuclei of R. temporaria oocytes (stage 6, according to Dumont, 1972) different methods were used: heavy meromyosin decoration, antiactin immunofluorescence with monoclonal antibodies, staining with rhodamine phalloidin, and electrophoresis in polyacrylamide gel. The nuclei of R. temporaria oocytes contain a considerable quantities of actin microfilaments which form intranuclear meshwork. Microfilaments are connected with the nucleoli, nucleolar RNP-complexes and nuclear envelope. Immunofluorescence with antiactin monoclonal antibodies reveals a strong staining of microfilaments and nucleoli. A slight staining of nucleoli is observed after the treatment of nuclei with rhodamine phalloidin. A specific role of intranuclear microfilaments in direct transport of nucleolar material from the nucleus into the oocyte cytoplasm, in stabilization of the karyosphere (the late diplotene oocyte complex of chromosomes with numerous nucleoli) is discussed in addition to its keeping in a definite region of the nucleus. A supposition is drawn on the functional significance of the connection between microfilaments and nuclear matrix. Based on our own and literature data, a conclusion is drawn, that the intranuclear filament actin may be one of the leading components in morpho-functional organization of the nucleus as the whole.