Peculiarities of chromosome organization in meiosis

Meiotic and mitotic chromosomes have a complex of differences. (1) At the early prophase I of meiosis, chromosomes acquire protein axial elements (AEs) that were absent in mitosis; in addition to somatic cohesins, AEs contain the meiosis-specific cohesins REC8, SMC1β, and STAG3. (2) At the middle prophase I, protein lateral elements (LEs) of synaptonemal complexes (SCs) are formed on the basis of AEs. The LE proteins are not conserved, but in Saccharomyces cerevisiae and Arabidopsis thaliana they contain functional domains with conserved secondary structures. Among the almost 679 thousand proteins of primitive eukaryotes that we studied by bioinformatics methods, in green and brown algae, some lower fungi, and Coelenterata, we revealed proteins or functional domains similar to SC proteins. (3) During the pachytene and diplotene stages of meiosis, chromosomes of spermatocytes and mother pollen cells acquire a general structure resembling the structure of amphibian and avian lampbrush chromosomes in miniature. Lateral chromatin loops with sizes of 90, 160, and even over 480 Kb were observed in human spermatocytes during the diplotene stage. In combination, all these observations confirm the considerable conservation of the scheme of molecular and ultrastructural organization of meiotic chromosomes in a large variety of eukaryotic organisms.     

Higher levels of organization in chromosomes.

On the basis of recent results a unified view of different aspects of the higher levels in the organization of chromatin in chromosomes is presented. Basic to these forms of organization is the arrangement of DNA in the complex with nucleosomes and recent studies suggest that at least some species of satellite DNA may maintain a fixed DNA sequence relationship to the phasing of nucleosomes. Special proteins such as the high-mobility group (HMG) proteins or other non-histone proteins could serve specific functions in the recognition of satellite DNA sequences.In the presence of histone H1 the 110 Å nucleosome fiber formed from the basic string of nucleosomes can be further condensed into a thicker 250–300 Å fiber formed by a solenoidal coiling of the 110 Å fiber with about 6–8 nucleosomes per turn and the available evidence suggests that these structures are found in mitotic chromosomes as well as other forms of inactive chromatin. A further level of coiling of the 250–300 Å solenoid has been suggested by our recent studies of disintegrated mitotic chromosomes consisting of a thin-walled tube with an outer diameter of 4000 Å referred to as the unit fiber. This structure would account for a factor of 1400 × contraction of DNA in mitotic chromosomes which in their intact state are only 5-fold more contracted. The recently described “scaffold” proteins could be responsible for this final coiling of the unit fibers in intact chromosomes.Meiotic chromosomes are generally less contracted than mitotic chromosomes. An extreme example of this are lampbrush chromosomes that apart from the axial segments which might contain some structural proteins appear to consist of naked DNA arranged in lateral loops. In the later stages of meiosis more condensed structures arise as exemplified by the synaptonemal complex during the pachytene stage in many organisms. The characteristic features of this structure are interpreted to suggest that the structure consists of lateral components containing two parallel 110 Å nucleosome fibers each representing the axial segments of two sister chromatids. From these paired regions loops protrude laterally in a manner which closely resembles the less condensed lampbrush chromosomes. The implication of this structure in the process of crossingover is discussed.Less is known about the organization of chromatin in interphase nuclei, but structures analogous to the loop-like structures in meiotic chromosomes are suggested on the basis of the isolation of supercoiled DNA loops constrained by RNA-DNA and protein-DNA interactions. The position of these loops is suggested to be fixed by specific repeated DNA sequences that could be associated with specific tenacious non-histone or HMG proteins.     

Synaptonemal Complex und Chromosomenstruktur in der achiasmatischen Spermatogenese vonPanorpa communis (Mecoptera)

Meiotic prophase in the spermatocytes ofPanorpa communis was studied. There is a proper sequence of meiotic stages in the testes. Therefore the temporal development of chromosome structure and the synaptonemal complex (SC) could be studied exactly. The structure and function of the SC are interpreted in a new model.—The chromosomes have a lambrush form from leptotene to diakinesis. At leptotene each chromatid produces an additional axis of basic protein and RNA. The axis becomes one of the lateral elements of the SC. At pachytene the DNA of the bivalents is separated into three regions: 1. Most of the DNA forms long loops outside the SC. 2. Smaller portions of the DNA filaments are twisted around the lateral elements of the SC. 3. Short DNA loops (called pairing loops) extend into the pairing space. InPanorpa the SC is composed of two lateral elements (chromosome axes), which are connected by equally spaced transverse filaments, a ladder-like central element in the middle of the pairing space and, on each side of the pairing space parallel to the lateral elements, two RNA containing strands. These are regarded as connected RNA copies of the pairing loops and are responsible for the exact pairing of homologous chromosome segments. At diplotene the axes of the sister chromatids separate to form “double complexes” with four lateral elements. The double complexes of the oocytes contain only transverse filaments between the axes of the homologous chromatids. After a short time they disappear again and the homologues separate to form the chiasmatic bivalents. In the spermatocytes all four chromatid axes are connected by transverse filaments. The pairing complex persists until diakinesis, thereby causing the suppression of the diplotene stage in the light microscope. This may be the only reason for the achiasmatic meiosis in the spermatocytes ofPanorpa.     

Synaptonemal complexes with associated chromatin in a moth,Ephestia kuehniella Z.

Chromosome structure and pairing behaviour of the pachytene bivalents in the wildtype and in W chromosome mutants were studied using a microcentrifugation technique. The spread bivalents display a characteristic lampbrush structure with lateral loops having the typical appearance of nucleosomal fibers, in autosomes as well as in the W and Z chromosomes. While the autosomal loops are always completely dispersed by the spreading forces, the loops of the heterochromatic W chromosome frequently are found to be condensed in tangles. These tangles contain supranucleosomal globular particles of a diameter of 37.7±1.2 nm. — Pairing of the WZ can be complete or partial, probably depending on the stage of the pachytene. Incomplete pairing normally is interpreted as demonstrating non-homology. Pairing was weak, however, even between homologous segments of the W chromosome, which were introduced into the genome in homozygous form by a translocation chromosome.     

Lampbrush-type chromosomes in the primary nucleus of the green alga Acetabularia mediterranea

Structures with a lampbrush-chromosome-like morphology are described in the nucleoplasm of primary nuclei of the green alga, Acetabularia mediterranea, by light and electron microscopy in sections of cells fixed in situ and in spread preparations of isolated nuclear components. These chromosomes reveal typical loops (up to 20 μm long), chromomere-like nodules (1–2 μm in diameter), and 2–4 μm large axial globules. Associations of some of these chromosomes with nucleolar structures and with the nuclear envelope are also recognized. The light microscopically identified loops are correlated with distinct fibrillogranular structures observed in the thin sections and with the very long matrix units seen in the spreadpreparations. The similarity of these structures to the lampbrush chromosomes of various animal cell types, all exclusively stages of meiotic prophase, is discussed as well as the possible relation of the appearance of lampbrush chromosomes to a defined phase of the vegetative growth of this alga.     

Die Entstehung multipler Oocytennukleolen aus akzessorischen DNS-Körpern bei Gryllus domesticus

The early stages of female and male germ cells have been investigated in Feulgen squash preparations, in unfixed state with phase contrast optics and in the electron microscope. The DNA axes of the ring-shaped multiple nucleoli in the growing oocytes of Gryllus arise from compact DNA bodies which are found in oogonia of young larvae and in oocytes prior to the growth period. The nuclei of the early oogonia contain several little DNA bodies whereas young oocytes at leptotene, zygotene and pachytene have only one body which is bigger than at earlier stages (Pig. 3). At metaphase and anaphase during oogonial mitosis the DNA body has a filamentous shape distinguishable from the compact chromosomes (Fig. 5). In oogonia as well as at leptotene and zygotene stages, nucleoli are produced in the peripheral, uncoiled parts of each DNA body whereas the compact interior is completely free of nucleolar material (Figs. 4, 12). At pachytene, the whole DNA body begins to despiralize, and single DNA strands are released into the nucleoplasm. These strands form hundreds of multiple nucleoli which finally are dispersed in the germinal vesicle (Fig. 11). — Incorporation studies with radio-active thymidine have shown that DNA synthesis in the DNA body is not synchronous with the S-phase of the chromosomes (Fig. 7). — The DNA body is an own formation distinct from the sex chromosomes (in contrast to the opinion of Sotelo and Wettstein, 1964). Although the positive heteropycnotic X-chromosome in the germ cells of the male cricket is very similar to the DNA body of the female (Fig. 8), there is no regular contact between sex chromosome and nucleolus neither in spermatogonia nor in spermatocytes (Figs. 9, 14). In all probability, the site of the nucleolar organizer is autosomal. — It is suggested that the amplification of the nucleolar genes in Gryllus oocytes results in an accumulation of ribosomal RNA for use during the early cleavage stages of the embryo     

Etude preliminaire de stades de la méiose humaine dans les spermatocytes et les ovocytes

Summary Human meiotic chromosomes, from spermatocytes and ovocytes, are described after observations of whole mount preparations under E.M. Small testicular and ovarian fragments are put in distillated water, then macerated; the cell suspension is spread on the surface of sheet copper grids covered with formvar plus collodion films. After dehydratation interesting stages are selected under L.M. before observations under E.M.Zygotene and pachytene are the most common stages. During pachytene the chromomeres are well individualized; the synaptonemal complex may be observed; chromatin fibers connect the chromosomes to nuclear pores, interchromosomal fibers joint the bivalents. Zygotene and pachytene bivalents are very similar in the male and the feminine germ cells.     

Genetic function correlated with unfolding of lampbrush loops by the Y chromosome in spermatocytes of Drosophila hydei

Summary Deficiencies of the Y chromosome of Drosophila hydei including sites which develop lampbrush loops invariably cause sterility of males. Suppression of loop unfolding in one or more sites equally results in similar morphogenetic defects of spermiogenesis. A variegated type repression of lampbrush loop unfolding observed during the spermatocyte stage results in varying morphogenetic effects on spermiogenesis. This demonstrates the existence of causal relationships between the active phase of Y chromosomal factors in spermatocytes and the differentiation processes in spermatids.In some translocated Y fragments the mode of unfolding of a particular pair of lampbrush loops may be permanently changed. As a result, lampbrush loops of a mutant phenotype are developed. Some alterations of this type are correlated with functional alterations resulting in defective spermiogenesis.Three different fragments of the Y chromosome in which lampbrush loop formation was repressed have been tested for possible reversions of loop suppression by means of X irradiations. In none of the three cases reversion has been detected among two thousand tested chromosomes.To the memory of Karl-Heinz Bier.     

Funktionsstrukturen im Oocytenkern von Locusta migratoria

Examination of living oocyte nuclei of Locusta migratoria has revealed the presence of thread-like struktures. They are paired and are thought to be the uncoiled chromosomes since they are broken into fragments by treatment with DNase. The greater part of the threads carries lateral loops like the lampbrush chromosomes of Amphibia (Fig. 14). A smaller part has no loops hut bears a series of conspicious granules with bright appearance under positive phase contrast optics (pearl-string segments) (Fig. 2). — The visibility of the chromosomes has been investigated in solutions with several ions. In hypertonic media the chromosomes contract, the granules fuse, and the pearl-string segments become lumpy (Fig. 21). In nitrogenous atmosphere and if kept at low temperature the pearl-string structures are likewise transformed into a few lumps (Fig. 19). After return to normal conditions they reconstitute their characteristic beaded appearance. — In autoradiographs obtained by injection of H³-uridine into the body cavity and by incubation of isolated nuclei in vitro, a rather uniformly distributed labelling occurs over the oocyte nuclei up to 30 min incubation time (Fig. 23). With prolonged incubation the activity of the pearl-string segments becomes more intense than the labelling of the lampbrush chromosomes (Fig. 24). After treatment with actinomycin RNA synthesis is stopped, the pearl-string axes and the lampbrush chromosomes contract, and the granules disappear more and more (Figs. 25-28). — The pearlstring segments look very much like the nucleoli in the oocytes of Amphibia, where the nucleolar substance is likewise distributed as a series of beads of rather uniform size on an axis. Therefore, the pearl-string structures may have nucleolar function in Locusta too. If so, the only difference to the Amphibian nucleoli would be the continued attachment to the lampbrush chromosomes.     

Synaptonemal complex analysis of mouse chromosomal rearrangements

Synaptonemal complex (SC) analysis by electron microscopy of spermatocytes in surface microspreads was carried out in mice heterozygous for two paracentric inversions: either In(1)1RK or In(2)5Rk. Characteristic SC inversion loops are formed at synapsis in bivalents carrying the rearrangements. Although all loops were observed to be eliminated by late pachytene through synaptic adjustment, every spermatocyte at early pachytene contained a fully synapsed loop. Cells in the earliest stage of pachytene contained the longest loops and thus had undergone minimal adjustment. The SC estimates of inversion lengths and breakpoint positions in such cells corresponded well with those from mitotic chromosome banding and could be correlated with genetic maps of chromosomes # 1 and # 2, thus demonstrating the basis for the mapping of pachytene chromosomes. The regularity of loop formation and reproducibility of the SC analysis are reflected in the constant relative positions of the estimated breakpoints. The method is sensitive enough to reflect small, real, interstitial length differences between meiotic and mitotic chromosomes. The results demonstrate the feasibility and precision of detection and quantitative characterization of inversions at early meiotic prophase by SC analysis.This paper is warmly dedicated to Prof. Dr. Wolfgang Beermann, on the occasion of his 60th birthday