ELECTRON MICROSCOPIC OBSERVATIONS ON THE SUBMICROSCOPIC MORPHOLOGY OF THE MEIOTIC NUCLEUS AND CHROMOSOMES

Thin sections of the testicular follicles of the grasshopper Laplatacris dispar were studied under the electron microscope. In the primary spermatocytes, during meiotic prophase, three main regions can be recognized within the nucleus: (1) the nucleolus and associated nucleolar material; (2) the interchromosomal regions with the dense particles; and (3) the chromosomes. The nucleolus is generally compact and is surrounded by nucleolar bodies that comprise aggregations of dense round particles 100 to 250 A in diameter. A continuous transition can be observed between these particles and those found isolated or in short chains in the interchromosomal spaces. Particles of similar size (mean diameter of 160 A) can be found associated with the nuclear membrane and in the cytoplasm. The chromosomes show different degrees of condensation in different stages of meiotic prophase. The bulk of the chromosome appears to be made of very fine and irregularly coiled filaments of macromolecular dimensions. Their length cannot be determined because of the thinness of the section but some of them can be followed without interruption for about 1000 to 2000 A. The thickness of the chromosome filaments seems to vary with different stages of prophase and in metaphase. In early prophase, filaments vary between 28 ± 7 A and 84 ± 7 A with a mean of 47 A, in late prophase the mean is about 70 A. In metaphase the filaments vary between 60 and 170 A with a mean of about 100 A. Neither the prophase nor the metaphase chromosomes have a membrane or other inhomogeneities. The finding of a macromolecular filamentous component of chromosomes is discussed in relation to the physicochemical literature on nucleoproteins and nucleic acids and as a result it is suggested that the thinnest chromosome filaments (28 ± 7 A) probably represent single deoxyribonucleoprotein molecules.     

A LIGHT AND ELECTRON MICROSCOPE STUDY OF SMALL, SPHERICAL NUCLEAR BODIES IN MERISTEMATIC CELLS OF ALLIUM CEPA, VICIA FABA, AND RAPHANUS SATIVUS

Interphase, preprophase, and prophase nuclei of meristematic cells of Allium cepa, Vicia faba, and Raphanus sativus are characterized by the presence of spherical bodies approximately 1 µ in diameter. These structures are Feulgen-negative but stain metachromatically with azure B, as the nucleolus, following fixation with glutaraldehyde. At the ultrastructural level, they consist predominantly of fibrils estimated to be between 70 and 100 A in diameter which greatly resemble those found within certain zones of the nucleolus in these plant species. Moreover, in Allium cepa, these spherules often exhibit dense particles which are also found within the fibrillar zones of the nucleolus in this species. The observations that the bodies in question are frequently located at the surface of the nucleolus and moreover show cytochemical and ultrastructural similarities with this organelle suggest that they may originate from the nucleolus. However, the common association of the spherules with chromosome strands may indicate instead that these bodies represent extranucleolar ribonucleoprotein materials synthesized by specific chromosomal loci.     

CYTOPLASMIC INCLUSIONS RESEMBLING NUCLEOLI IN SYMPATHETIC NEURONS OF ADULT RATS

A distinctive cytoplasmic inclusion consisting of a convoluted network of electron-opaque strands embedded in a less dense matrix was identified in the neurons, but not in the supporting cells, of rat sympathetic ganglia. This ball-like structure, designated "nematosome," measures ~ 0.9 µ and lacks a limiting membrane. Its strands (diameter = 400–600 A) appear to be made of an entanglement of tightly packed filaments and particles ~ 25–50 A thick. Cytochemical studies carried out with the light microscope suggest the presence of nonhistone proteins and some RNA. Usually only one such structure is present in a cell, and it appears to occur in most ganglion cells. Although they can be seen anywhere in the cell body, nematosomes are typically located in the perinuclear cytoplasm, where they are often associated with smooth-surfaced and coated vesicles. In fine structure and stainability, they bear a resemblance to the fibrous component of the nucleolus. Subsynaptic formations, which are a special feature of some of the synapses in sympathetic ganglia, appear similar to the threadlike elements in the nematosomes. The possibility that these three structures—nucleolus, nematosome, and subsynaptic formation—may be interrelated in origin and function is discussed.     

Unraveling the Nanoscale Surface Properties of Chitin Synthase Mutants of?Aspergillus fumigatus and Their Biological Implications

Understanding the surface properties of the human opportunistic pathogen Aspergillus fumigatus conidia is essential given the important role they play during the fungal interactions with the human host. Although chitin synthases with myosin motor-like domain (CSM) play a major role in cell wall biosynthesis, the extent to which deletion of the CSM genes alter the surface structural and biophysical-biological properties of conidia is not fully characterized. We used three complementary atomic force microscopy techniques—i.e., structural imaging, chemical force microscopy with hydrophobic tips, and single-molecule force spectroscopy with lectin tips—to gain detailed insights into the nanoscale surface properties (ultrastructure, hydrophobicity) and polysaccharide composition of the wild-type and the chitin synthase mutant (ΔcsmA, ΔcsmB, and ΔcsmA/csmB) conidia of A. fumigatus. Wild-type conidia were covered with a highly hydrophobic layer of rodlet nanostructures. By contrast, the surface of the ΔcsmA mutant was almost completely devoid of rodlets, leading to loss of hydrophobicity and exposure of mannan and chitin polysaccharides. The ΔcsmB and ΔcsmA/csmB mutants showed a different behavior, i.e., the surfaces featured poorly organized rodlet layers, yet with a low hydrophobicity and substantial amounts of exposed mannan and chitin at the surface. As the rodlet layer is important for masking recognition of immunogenic fungal cell wall components by innate immune cells, disappearance of rodlet layers in all three chitin synthase mutant conidia was associated with an activation of human dendritic cells. These nanoscale analyses emphasize the important and distinct roles that the CSMA and CSMB genes play in modulating the surface properties and immune interactions of A. fumigatus and demonstrate the power of atomic force microscopy in fungal genetic studies for assessing the phenotypic characteristics of mutants altered in cell surface organization.     

A HEAT-SENSITIVE CELLULAR FUNCTION LOCATED IN THE NUCLEOLUS

Striking nucleolar lesions occur in cultured cells after exposure to supranormal temperatures. These lesions appear at 42°C and consist of a loss of the granular ribonucleoprotein (RNP) component and intranucleolar chromatin, and a disappearance of the nucleolar reticulum. The material remaining in the morphologically homogeneous nucleolus is a large amount of closely packed fibrillar RNP. The lesions remain identical as temperature increases to 45°C. These alterations are reversible when the cells are returned to 37°C and are associated with the reappearance of an exaggerated amount of intranucleolar chromatin and granular RNP. High-resolution radioautography indicates that after thermic shock nucleolar RNA synthesis is inhibited whereas extranucleolar sites are preserved: it also suggests that the granular RNP is reconverted to fibrillar RNP probably by simple unraveling. The results prove the existence of heat-sensitive cellular functions in the nucleolus which deal with the DNA-dependent RNA synthesis. The precise site of action is assumed to involve hydrogen bonds, resulting in configurational changes in nucleolar RNP and affecting the stability of the DNA molecule. The subsequent events in nucleolar RNA synthesis are discussed in light of the morphologic and biochemical effects of actinomycin D on the nucleolus.     

SARCOPLASMIC RETICULUM OF AN UNUSUALLY FAST-ACTING CRUSTACEAN MUSCLE

The fast-acting, synchronous "remotor" muscle of the lobster second antenna was examined by light and electron microscopy and was found to have a more profuse sarcoplasmic reticulum (SR) than any other muscle known. Myofibrils are widely separated from one another and occupy only about one-fourth of the volume of the muscle; most of the remaining volume is taken up by the SR, which resembles the smooth-surfaced reticulum of steroid-secreting cells. Dense granules (0.03–0.1 µ in diameter) are scattered through the reticulum. T-tubules penetrate into the fibers and form dyads along the A bands of myofibrils; however, ferritin-labeling experiments show that the volume of the T-system is very small compared with that of the SR. Myofibrils are ~0.5 µ x 1.0 µ in cross section and consist of thick filaments, which appear tubular except at the M region, and thin filaments, which are situated midway between neighboring thick filaments. The ratio of thin to thick filaments is 3:1. The extreme development of the SR in this muscle is discussed in relation to the exceedingly short duration of the contraction-relaxation cycle.     

Chemical composition and electron microscopy of the rodlet layer ofAspergillus nidulans conidia

The rodlet layers of wild-type and white mutantAspergillus nidulans conidia were purified by a simple centrifugation procedure after conidial suspensions were subjected to sonication. Chemical analysis showed that the major components of wild-type rodlets were protein and melanin in almost equal amounts, followed by carbohydrate. White mutant rodlets differed from those of the wild-type strain in that the melanin content was very low. Histidine, aspartic acid, glutamic acid, glycine, and alanine were the most prominent amino acids in the rodlet layer of the white mutant, whereas cystine and methionine were not found. Electron microscopy studies showed that the rodlets of both white mutant and wild-type strains were grouped into fascicles, which varied from 80 to 160 nm in width. Individual rodlets measured 125–360 nm in length and 7 nm in diameter.     

FILAMENT LENGTHS IN STRIATED MUSCLE

Filament lengths in resting and excited frog muscles have been measured in the electron microscope, and investigations made of the changes in length that are found under different conditions, to distinguish between those changes which arise during preparation and the actual differences in the living muscles. It is concluded that all the measured differences in filament length are caused by the preparative procedures in ways that can be simply accounted for, and that the filament lengths are the same in both resting and excited muscles at all sarcomere lengths greater than 2.1 µ, viz., A filaments, 1.6 µ; I filaments, 2.05 µ. The fine periodicity visible along the I filaments also has been measured in frog, toad, and rabbit muscles and found to be 406 A.     

Relaxed tarantula skeletal muscle has two ATP energy-saving mechanisms

Myosin molecules in the relaxed thick filaments of striated muscle have a helical arrangement in which the heads of each molecule interact with each other, forming the interacting-heads motif (IHM). In relaxed mammalian skeletal muscle, this helical ordering occurs only at temperatures >20°C and is disrupted when temperature is decreased. Recent x-ray diffraction studies of live tarantula skeletal muscle have suggested that the two myosin heads of the IHM (blocked heads [BHs] and free heads [FHs]) have very different roles and dynamics during contraction. Here, we explore temperature-induced changes in the BHs and FHs in relaxed tarantula skeletal muscle. We find a change with decreasing temperature that is similar to that in mammals, while increasing temperature induces a different behavior in the heads. At 22.5°C, the BHs and FHs containing ADP.P_i are fully helically organized, but they become progressively disordered as temperature is lowered or raised. Our interpretation suggests that at low temperature, while the BHs remain ordered the FHs become disordered due to transition of the heads to a straight conformation containing Mg.ATP. Above 27.5°C, the nucleotide remains as ADP.P_i, but while BHs remain ordered, half of the FHs become progressively disordered, released semipermanently at a midway distance to the thin filaments while the remaining FHs are docked as swaying heads. We propose a thermosensing mechanism for tarantula skeletal muscle to explain these changes. Our results suggest that tarantula skeletal muscle thick filaments, in addition to having a superrelaxation–based ATP energy-saving mechanism in the range of 8.5–40°C, also exhibit energy saving at lower temperatures (<22.5°C), similar to the proposed refractory state in mammals.     

Self-assembly of proteins into a three-dimensional multilayer system: Investigation of the surface of the human fungal pathogen Aspergillus fumigatus

Hydrophobins are small surface active proteins that fulfil a wide spectrum of functions in fungal growth and development. The human fungal pathogen Aspergillus fumigatus expresses RodA hydrophobins that self-assemble on the outer conidial surface into tightly organized nanorods known as rodlets. AFM investigation of the conidial surface allows us to evidence that RodA hydrophobins self-assemble into rodlets through bilayers. Within bilayers, hydrophilic domains of hydrophobins point inward, thus making a hydrophilic core, while hydrophobic domains point outward. AFM measurements reveal that several rodlet bilayers are present on the conidial surface thus showing that proteins self-assemble into a complex three-dimensional multilayer system. The self-assembly of RodA hydrophobins into rodlets results from attractive interactions between stacked β-sheets, which conduct to a final linear cross-β spine structure. A Monte Carlo simulation shows that anisotropic interactions are the main driving forces leading the hydrophobins to self-assemble into parallel rodlets, which are further structured in nanodomains. Taken together, these findings allow us to propose a mechanism, which conducts RodA hydrophobins to a highly ordered rodlet structure. The mechanism of hydrophobin assembly into rodlets offers new prospects for the development of more efficient strategies leading to disruption of rodlet formation allowing a rapid detection of the fungus by the immune system.     

ON THE ROLE OF MICROTUBULES IN MOVEMENT AND ALIGNMENT OF NUCLEI IN VIRUS-INDUCED SYNCYTIA

Infection of baby hamster kidney (BHK21-F) cells with the parainfluenza virus SV5 causes rapid and extensive cell fusion. Time-lapse cinematography shows that when cells fuse, their nuclei migrate straight to the center of the syncytium at rates of 1–2 µ/min. Nuclei are often arranged in long, tightly packed, parallel rows in syncytia derived from the fibroblastic BHK21-F cells. Polarization microscopy shows birefringent material between and parallel to these rows of nuclei, and electron microscopy shows bundles of cytoplasmic microtubules, ~250 A in diameter, and filaments, ~80 A in diameter, parallel to and between the rows of nuclei. Colchicine treatment causes disappearance of microtubules from BHK21-F cells and an apparent increase in the number of 80-A filaments. Although colchicine-treated, SV5-infected cells fuse, their nuclei do not migrate or form rows but remain randomly scattered through the syncytial cytoplasm. Incubation at 4°C does not disrupt microtubules in BHK21-F cells. Rows of nuclei have been isolated from SV5-induced syncytia, and the nuclei in them have been found to be intimately associated with microtubules but not with other cytoplasmic structures. These results suggest that microtubules demarcate cytoplasmic channels through which nuclei migrate and that they may also be involved in the mechanism of nuclear movement.     

High-resolution cell surface dynamics of germinating Aspergillus fumigatus conidia

We used real-time atomic force microscopy with a temperature-controlled stage (37°C) to probe the structural and physicochemical dynamics of single Aspergillus fumigatus conidia during germination. Nanoscale topographic images of dormant spores revealed the presence of a layer of rodlets made of hydrophobins, in agreement with earlier electron microscopy observations. Within the 3-h germination period, progressive disruption of the rodlet layer was observed, revealing hydrophilic inner cell wall structures. Using adhesion force mapping with hydrophobic tips, these ultrastructural changes were shown to correlate with major differences in cell surface hydrophobicity. That is, the rodlet surface was uniformly hydrophobic due to the presence of hydrophobins, whereas the cell wall material appearing upon germination was purely hydrophilic. This study illustrates the potential of real-time atomic force microscopy imaging and force spectroscopy for tracking cell-surface dynamics.     

Characteristics of Streptomyces coelicolor A3(2) aerial spore rodlet mosaic

Cytochemical analysis of Streptomyces coelicolor (A3(2) indicated that the aerial growth rodlet mosaic is a polysaccharide. Statistical analysis of frequency distributions of individual rodlet lengths from control and ether-reoriented spore mosaics indicated that the rodlet fibrillar image is the result of individual particulates, rather than evaginations in a continuous sheet of material. A model of the mature sport envelope was developed from freeze-etch-replicated, thin-sectioned, and critical point dried S. coelicolor A3(2) mature spores. The rodlet mosaic was situated between the outer spore wall and an external granuloma matrix. Mixture spore envelope layers from the inner surface to the external surface are plasma membrane, inner spore wall, outer spore wall, rodlet mosaic, an undefined granular matrix, and the sheath. The granular matrix had an uneven thickness and much of the matrix was frequently absent from the interspore spaces of mature spore chains. Streptomyces coelicolor A3(2) mosaic rodlets were isolated by acetic acid refluxing, then ethanol precipitation. Complete acid hydrolysis of rodlets released on sugar which cochromatographed with D-glucosamine-HCl and released acetic acid at 139% of the expected level. Cell associated rodlet mosaics and isolated mosaic rodlets were hydrolyzed with chitinase. Infrared spectra of isolated rodlets were similar to crab chitin spectra.     

MYOSIN-LIKE AGGREGATES IN TRYPSIN-TREATED SMOOTH MUSCLE CELLS

Segments of the lower small intestine of the toad Bufo marinus were excised and soaked for approximately 2 hr in Ringer's solution (pH 7.4 or 7.8) containing crystalline trypsin and then fixed for electron microscopy at approximately the same pH. Thin sections of the tunica muscularis of these specimens show smooth muscle cells ranging in appearance from severely damaged at one extreme to apparently unaffected at the other. Among these are cells at intermediate stages, including some which exhibit large and conspicuous populations of thick filaments closely resembling artificially prepared aggregates of smooth muscle myosin. The thick filaments have the form of tactoids ~ 250–300 A in diameter in their middle regions and are ~ 0.5–1.0 µ in length. In some preparations they also display an axial periodicity approximating 143 A. They are usually randomly oriented and segregated from the thin filaments, which tend to form closely packed, virtually crystalline bundles at the periphery of these cells. "Dense bodies" are absent from cells showing these changes. The simplest interpretation of these data is that smooth muscle myosin normally exists among the actin filaments in a relatively disaggregated state and that trypsin induces aggregation by altering the conformation of the myosin molecule. Alternatively, trypsin may act indirectly through an effect on some other smooth muscle protein which normally forms a stable complex with relatively disaggregated myosin.     

MORPHOLOGY OF RIGOR-SHORTENED BOVINE MUSCLE AND THE EFFECT OF TRYPSIN ON PRE- AND POSTRIGOR MYOFIBRILS

Bovine semitendinosus muscles were sampled immediately after death, after 24 hr postmortem with storage at 2°, 16°, or 37°C, and after 312 hr postmortem with storage at 2° and 16°C. A biopsy technique was used to prevent shortening during glutaraldehyde fixation. Postfixation in osmium tetroxide was followed by embedding in an Epon-Araldite mixture. Bovine muscle was supercontracted after 24 hr storage at 27deg; but was only slightly contracted after storage at 16° for 24 hr. Muscle held at 37° for 24 hr was slightly less supercontracted than the 2° muscle. Striking similarities existed between muscles stored at 16° and at 2°C for 312 hr. Both were slightly shortened with narrowed I bands and an area of increased density, probably due to overlap of thin filaments in the middle of the A band. Postmortem shortening was accompanied by banding-pattern changes similar to those predicted for contracting muscle by Huxley and Hanson's sliding filament model. Treatment of myofibrils with 0.05% trypsin resulted in a rapid loss of Z lines and, in supercontracted myofibrils, caused a return of the banding pattern of resting muscle.     

ELECTRON MICROSCOPE AND EXPERIMENTAL INVESTIGATIONS OF THE NEUROFILAMENTOUS NETWORK IN DEITERS'' NEURONS : Relationship with the Cell Surface and Nuclear Pores

The assembly of filamentous elements and their relations to the plasma membrane and to the nuclear pores have been studied in Deiters' neurons of rabbit brain. Electron microscopy of thin sections and of ectoplasm spread preparations have been integrated with physicochemical experiments and differential interference microscopy of freshly isolated cells. A neurofilamentous network extends as a continuous, three-dimensional, semilattice structure throughout the ectoplasm, the "plasma roads," and the perinuclear zone of the perikaryon. This space network consists of ~90-Å wide neurofilaments arranged in fascicles which are interconnected by an exchange of neurofilaments. The neurofilaments consist of intercoiled ~20-Å wide unit-filaments and are associated through cross-associating filaments with other neurofilaments of the fascicle and with microfilaments. The ~20–50-Å wide microfilaments display intimate associations with the plasma membrane and with the nuclear pores. Electron microscopy of thin sections from glycerinated and heavy meromyosin-treated Deiters' neurons shows that actin-like filaments are present in the pre- and postsynaptic regions of synapses terminating on these neurons. It is proposed that the neurofilamentous space network serves a transducing function by linking plasma membrane activities with the genetic machinery of the neuron.     

The structure and behavior of the nucleolus organizers in mammalian cells

The regularly occurring secondary constrictions on metaphase chromosomes of mammalian cells prove to be nucleolus organizers as expected. The expression of nucleolus organizers as secondary constrictions, however, varies from cell to cell and from tissue to tissue, including cultivation in vitro. Electron micrographs of the organizer region show that the nucleolus organizer at metaphase is not a constriction. The width of the organizer area is the same as the condensed chromosomal arms; but the filaments, which are the major components of this region, show a diameter of 50–70 Å. The condensed chromosome arms consist of filaments 150–200 Å in diameter. In some mammalian species, structures similar to the nucleolus organizer are located at the end of chromosomes. These may be terminal nucleolus organizers.Supported in part by Research Grants DRG-269 from Damon Runyon Memorial Fund for Cancer Research, E-286 from American Cancer Society, and HD-2590 from National Institutes of Health.     

EARLY STAGES IN THE DEVELOPMENT OF PLASTID FINE STRUCTURE IN RED AND FAR-RED LIGHT

Developmental changes in fine structure were studied in plastids of etiolated bean leaves during the time required for the protochlorophyllide-chlorophyllide transformation and the following lag phase prior to chlorophyll accumulation. In agreement with some other workers, two distinct stages of change in the fine structure of proplastids were found to occur upon illumination during this period. The first involves a dissociation of the previously fused units in the prolamellar bodies of the proplastids and occurs simultaneously with the protochlorophyllide-chlorophyllide conversion in light of 655 mµ, but not of 682, 700, or 730 mµ. The effect of the red light could not be reversed by a simultaneously supplied stronger far-red irradiation. The energy requirements for these structural changes parallel those for the pigment conversion. During the following step the vesicles which arose from the fused units of the prolamellar body were dispersed in rows through the stroma, and the prolamellar bodies themselves disappeared. For these changes to occur, higher light energies were required and the leaves had to be illuminated for longer periods. A red preillumination seemed to accelerate the development somewhat. The structural changes could be induced by light of 655 mµ, but also, to a lesser degree, of 730 mµ. No measurable additional chlorophyll accumulated during this period. Thus, the structural changes observed were independent of major changes in pigment content.     

Conidial Hydrophobins of Aspergillus fumigatus

The surface of Aspergillus fumigatus conidia, the first structure recognized by the host immune system, is covered by rodlets. We report that this outer cell wall layer contains two hydrophobins, RodAp and RodBp, which are found as highly insoluble complexes. The RODA gene was previously characterized, and ΔrodA conidia do not display a rodlet layer (N. Thau, M. Monod, B. Crestani, C. Rolland, G. Tronchin, J. P. Latgé, and S. Paris, Infect. Immun. 62:4380-4388, 1994). The RODB gene was cloned and disrupted. RodBp was highly homologous to RodAp and different from DewAp of A. nidulans. ΔrodB conidia had a rodlet layer similar to that of the wild-type conidia. Therefore, unlike RodAp, RodBp is not required for rodlet formation. The surface of ΔrodA conidia is granular; in contrast, an amorphous layer is present at the surface of the conidia of the ΔrodA ΔrodB double mutant. These data show that RodBp plays a role in the structure of the conidial cell wall. Moreover, rodletless mutants are more sensitive to killing by alveolar macrophages, suggesting that RodAp or the rodlet structure is involved in the resistance to host cells.     

A CORRELATED LIGHT AND ELECTRON MICROSCOPE STUDY OF THE NUCLEOLAR MATERIAL DURING MITOSIS IN VICIA FABA

Root meristematic cells of Vicia faba were examined, with both light and electron microscopes, in order to study the behaviour of the nucleolar material during the mitotic process. Under light microscopy, the preprophase nucleolus is seen to consist of a densely stained material in which are embedded several unstained vacuole-like structures of varying size. The electron microscope reveals that the dense nucleolar material is formed of two structurally distinct components, each segregated into irregularly shaped zones blending with one another. One of these components is represented by 150 A granules which, in places, are arranged into thread-like structures approximately 0.1 µ in diameter; the other component apparently consists of fibrils 60 to 100 A in diameter. The large and medium sized intranucleolar vacuoles contain loosely scattered granules and fibrils similar to those just described. The granular and fibrillar components of the denser portion of the nucleolus persist as such during prophase and disperse throughout the nuclear cavity at the time of nucleolar disintegration. After nuclear membrane breakdown, these granules and fibrils, as well as those of the nucleoplasm, mix freely with similar elements already present within the forming spindle. No evidence has been obtained that, during or after nucleolar disintegration, the structural components of the nucleolus become associated as such with the chromosomes to form an external or internal matrix. Our observations suggest the existence, of a matrix substance within late prophase, metaphase, and anaphase chromosomes, the fine structure of which bears strong resemblance to that of their constituent coiled chromonemata. Data are presented, moreover, that indicate that part of this matrix substance, presumably formed at some time during prophase, is released from the chromosomes during their anaphasic movement. A number of observations indicate that the main bulk of the next nucleolus is derived from a prenucleolar fibrillogranular material, arranged into thread-like structures some 0.1 µ in diameter, which collect in the interchromosomal spaces during early and midtelophase. Finally, our data would seem to favour the view that most of this prenucleolar material results from a resumption of the synthetic activity of the early and midtelophase chromosomes rather than from a mere shedding of a preexisting matrix substance.