Quantitative analysis of NOR expression in a B chromosome of the grasshopper <Emphasis Type="Italic">Eyprepocnemi</Emphasis>s <Emphasis Type="Italic">ploran</Emphasis>s

The B₂₄ chromosome in the Torrox population of the grasshopper Eyprepocnemis plorans is recurrently attached to a nucleolus in diplotene cells, indicating the activity of its distally located ribosomal DNA (rDNA). The frequency of males expressing the B chromosome nucleolus organizer region (B-NOR) almost doubled in 4 years. The likelihood of expressing the B-NOR increased with the B number and, in males expressing it, about 20% of their cells showed a nucleolus attached to the B. When active, the B-NOR contributed more than 25% of total cell nucleolar area (NA). Within males expressing the B-NOR, total cell NA did not differ between cells showing the active or inactive B-NOR, suggesting that total cell NA is tightly regulated in this species. However, this parameter tended to increase in this population from 1999 to 2004, in parallel to the neutralization process which is taking place in this population. Finally, an analysis of A chromosome NOR interdependence for activity revealed a positive correlation among autosomes but a negative correlation between autosomes and the X chromosome, the manifestation of which depends on B-NOR activity. These results are discussed in the context of the nucleolus as a sensor of the stress caused by parasitic B chromosomes.     

End-to-end chromosome attachments in mitotic interphase and their possible significance to meiotic chromosome pairing

Cytological studies on telophase and early prophase in roottip cells of several plant species (Allium cepa, 2n=16; four Crepis species, including Crepis capillaris, 2n=6; Callitriche hermaphroditica, 2n=6; Nigella arvensis, 2n=12; Secale cereale, 2n=14) revealed that chromosome ends are attached two by two forming chains of chromosomes (interphase associations). In these chains homologous chromosomes are presumably located adjacent to each other. In Crepis capillaris it was observed that the two nucleolar chromosomes form a separate ring one end attached to the ring of the four remaining chromosomes and the other end attached to the nucleolus. It is proposed that these end-to-end attachments have significance for chromosome pairing in meiosis. The adjacent location of homologous chromosomes in the interphase associations would facilitate rapid and regular synapsis.     

Amounts of ribosomal DNA inAllium (Liliaceae)

Eight species ofAllium subgen.Allium sect.Allium have been studied at the cytological level by means of karyological analyses and at the biochemical level with regard to the proportions of ribosomal DNA. All the species have a basic genome of x = 8.A. sativum, A. commutatum, A. ampeloprasum, andA. vineale possess approximately 0.050% rDNA and two nucleolus organizer regions per basic chromosome set.A. sphaerocephalon andA. arvense have two nucleolus organizers, andA. amethystinum three nucleolus organizers per haploid (n = x) genome: the three species possess approximately 0.075% rDNA.A. acutiflorum has five nucleolus organizer regions per haploid genome and 0.121% rDNA. An attempt to relate these differences with functional and ecological characteristics indicates that evolutionary variation of rDNA proportions is not casual. Such data also can help to define systematic affinities and circumscribe infrageneric taxa.     

Fragmentation par les rayons X de l'organisateur d'une différenciation de chromosome en écouvillon (lampbrush), chez Pleurodeles waltlii

X-rays have been used to induce heritable changes in the specific morphology of the lampbrush chromosomes in the newt Pleurodeles waltlii. The karyotype organization of female progeny of irradiated males was studied. Nine out of ten females were found to have chromosomal aberrations. In one of the nine, one of the breaks occurred at the sphere organizer, the sphere being part of the striking morphological features of chromosome IV. On irradiation the normal sphere organizer had been broken into two fragments each of which, when recombined with other chromosome breaks, still forms a sphere. The relationship of these observations to genome redundancy is discussed.     

A new method of in situ hybridization

A new method for gene mapping at the chromosome level using in situ hybridization and scanning electron microscopy is described and has been applied to mapping the rRNA genes of Drosophila melanogaster. Biotin is covalently attached to Drosophila rRNA via a cytochrome c bridge at a ratio of one cytochrome-biotin per 130 nucleotides by a chemical procedure. Polymethacrylate spheres with a diameter of ca. 60 nm are prepared by emulsion polymerization and are covalently attached to the protein avidin at a ratio of 5–20 avidins per sphere. The biotin-labeled rRNA is hybridized to denatured DNA in a chromosome squash. Upon incubation with a sphere solution, some of the biotin sites become labeled with spheres because of the strong non-covalent interaction between biotin and avidin. The chromosome squash is examined in the scanning electron microscope (SEM). Polymer spheres, which are visible in the SEM, are observed to label the nucleolus, where the rRNA genes are located.Contribution number 5121 from the Department of Chemistry.     

The genetic control of nucleolus formation in wheat

The wheat variety Chinese Spring has four pairs of nucleolus organisers of known rDNA content. The genetic control of these has been investigated in root tip cells by cytologically scoring the number of nucleoli per cell in (a) aneuploid derivatives each having a different dosage of a particular chromosome or chromosome arm and (b) in substitution lines where nucleolus organiser chromosomes have been replaced by homologues possessing different amounts of rDNA. It has been assumed that nucleolus organiser activity is correlated with nucleolus size and thus with the presence of a cytologically visible nucleolus. Those nucleolus organisers on chromosomes 1A and 5D, which together possess only 10% of the rDNA form a visible nucleolus only infrequently in the presence of the larger nucleolus organisers on chromosomes 1B and 6B. When a major pair of organisers on chromosomes 1B or 6B is deleted, the smaller nucleolus organisers form a visible nucleolus more frequently. Similarly, when the major nucleolus organisers are replaced by organisers with less rDNA, the smaller nucleolus organisers form visible nucleoli more frequently. When a small nucleolus organiser is replaced by one with much more rDNA, a larger nucleolus is formed. These and other findings lead to the general conclusions that there is a frequently, but not invariably, seen correlation between rRNA gene number and nucleolus size. However the relative size of the nucleolus formed depends principally upon the proportion of the total active rRNA genes in the cell which are localised at the nucleolus organiser in question. Varying the dosage of at least 13 non nucleolus organiser chromosomes also resulted in changes in the number of visible nucleoli per cell. This implies the genetic control of individual nucleolus organisers is complex. Inclusion in the wheat genome of the nucleolus organiser chromosome from Aegilops umbellulata, causes suppression of the wheat nucleolus organisers, the Aegilops umbellulata organiser remaining active. This suppression is similar to that observed in many interspecific plant and animal hybrids.     

A chromosome rearrangement in Neurospora that produces segmental aneuploid progeny containing only part of the nucleolus organizer

In translocation T(ILVL)OY321 of Neurospora crassa a distal portion of the nucleolus organizer chromosome, including ribosomal DNA sequences and the nucleolus satellite, is interchanged with a long terminal segment of IL. When OY321 is crossed by Normal sequence, one-fourth of the meiotic products are segmental aneuploids that contain two copies of the long IL segment and that are deficient for the distal portion of the organizer. Each such product forms a nucleolus and is viable. The complementary aneuploid products are deficient for the IL segment and are therefore inviable. — In crosses of OY321xOY321, each product is capable of making two nucleoli; nucleoli formed by the separated nucleolus organizer parts usually fuse, but most 8-spored asci contain some nuclei in which two separate nucleoli can be seen. One nucleolus is then terminal on its chromosome while the second is interstitial and somewhat smaller. — In crosses of OY321 x Normal, half of the meiotic products are capable of making two nucleoli. However, only about 15% of 8-spored asci have one or more nuclei containing separate nucleoli. At pachytene and later in prophase I, the single fusion nucleolus is associated with three bivalent chromosome segments. Each nucleus of every ascus contains at least one nucleolus, even in asci where some nuclei display two nucleoli. — Crosses of Aneuploid x Normal are usually semibarren, producing a reduced number of ascospores, some of which are inviable. Some aneuploid cultures become fully fertile by reverting to a quasinormal sequence lacking a satellite. In some crosses of Aneuploid x Normal, individual asci may show at prophase I either complete loss, partial loss, or pycnosis of the translocated IL segment. This observation of pycnosis suggests chromosome inactivation. — Growth from aneuploid ascospores is initially slow, but can accelerate to the wild-type rate.     

Differences in the number of nucleolus organizers in Chironomus tepperi shown by in situ hybridization

Two stocks of Chironomus tepperi could be isolated. One stock, N(IV)⁺, contains nucleolus organizers in chromosome I and IV, whereas the other one, N(IV)^–, shows only one nucleolus in chromosome I. It is demonstrated by in situ hybridization with radioactive rRNA that the absence of the nucleolus in chromosome IV of stock N(IV)^– is not related to an inactivation of the nucleolar DNA, as might have been suggested, but is due to the lack of ribosomal cistrons.     

The Location of the nucleolus organizer regions in Drosophila hydei

The positions of the nucleolus organizer regions in metaphase chromosomes of Drosophila hydei were detected by in situ hybridization experiments. In agreement with earlier conclusions the nucleolus of the X chromosome was found to originate in a terminal region of the heterochromatic arm. The Y chromosome contains two nucleolus organizers, one in a terminal position of the long arm, and the other in the short arm. The implications with respect to the evolution of the Y chromosome are discussed.     

Comparative Cytogenetic Variation of the Salivary Gland Polytene Chromosomes in Chironomus riparius Mg., 1804 (Diptera, Chironomidae) from Two Polluted Biotopes of Bulgaria and Russia

The structural–functional variation ofChironomus riparius salivary gland polytene chromosomes was studied in two geographically isolated Palearctic regions, Bulgaria (village Pancharevo) and Russia (St. Petersburg). The two biotopes, where larvae were collected, were polluted with various heavy metals from anthropogenic sources. Hereditary paracentric heterozygous inversions were characteristic of the Russian population, whereas somatic paracentric or pericentric heterozygous inversions were more common in the Bulgarian one. All inversions occurred at low frequencies. Other aberrations found in the two populations included somatic deletions resulting in a pompon structure of chromosome IVG, heterozygous translocation between chromosomes IVG and IIIF, enlargement of individual disks, and the appearance of a heterozygous block close to the centromere of chromosome IVG. In addition, changes in functional activity of the nucleolus organizer and Balbiani rings (BRc/BRb) were observed. Several aberration breakpoints proved to coincide with satellites of the Alu and Hinf families.     

Regulation of karyotype stability in tobacco tissue cultures of normal and tumorous genotypes

Summary Callus cultures of Nicotiana glauca, N. langsdorffii and of their tumor-forming hybrid plants contained a high frequency of cells with irregular chromosome numbers and chromosome aberrations (hypo-, hyper-, polyploid, aneuploid cells; bridges, polytene, broken, fragmented chromosomes, megachromosomes, etc.). Meristematic cells of shoot tips regenerated from the same cultures contained only regular chromosome numbers with normal chromosome structures. Variability in chromosome numbers is a consequence of abnormal mitoses. The data suggest genome segregation in the cultures. Cytological instability appears to be independent of genome segregation composition, genotype, tumorous condition, hormonal requirement and level of ploidy. The karyotype stability of the cultures is only dependent on the degree of organization of tissues and is regulated by factors involved in the control mechanisms of organizational processes.     

Nucleolus size varies with sex,ploidy and gene dosage in insects

The nucleolus constitutes a cytologically visible phenotype for ribosomal DNA (rDNA). Nucleolar size, as determined by silver staining, is a good indicator of cell proliferation rate and biosynthetic activity. Nevertheless, the relationship between rDNA content and sexual dimorphism for nucleolar size is not well documented. In the present study, the impact of sex and ploidy level on nucleolar size is investigated in three haplo/diploid and three diplo/diploid species of insect. Nucleolar sizes are found to be proportional to ploidy level in the haplo/diploid hymenopterans Trypoxylon albitarse and Nasonia vitripennis. Conversely, in the ant Messor barbarus, nucleolar sizes are larger in haploid males (winged) than diploid females (apterous). Among the diplo/diploid species, evidence for gene dosage compensation on nucleolar activity is suggested by the absence of sex differences in Drosophila simulans, a species in which rDNA is limited to the X chromosome. By contrast, in the grasshopper Stenobothrus festivus, another species with rRNA genes restricted to the X chromosome, the size of the nucleolus is significantly larger in females than in males. Additionally, in the grasshopper Chorthippus parallelus, where rDNA is distributed evenly on several autosomes of males and females, the females also show larger nucleoli than males. In both grasshopper species, the magnitude of the female/male ratio for nucleolus area is very similar to the body size ratio, suggesting that body size, as well as sex, ploidy, gene dosage and physiological activity, may be an important determinant of nucleolus area.     

Circadian clock phenotypes of chromosome aberrations with a breakpoint at the per locus

Summary The circadian rhythm phenotypes of eight chromosome aberrations with a breakpoint in the region of the per locus (3B1-2) were analyzed. Two duplications and five deficiencies with a 3B1-2 breakpoint produce either a wild-type or an arrhythmic clock phenotype while one translocation with a 3B1-2 breakpoint, T(1;4)JC43, produces locomotor-activity rhythms with either very-long period (31–39 h), rhythms that grade into arrhythmicity, or completely arrhythmic phenotypes. This is a unique phenotype that had not previously been observed for mutants at the per locus. An extensive complementation analysis of 3B1-2 chromosome aberrations and per mutant alleles provided no compelling evidence for genetic complexity at the per locus. This is in contrast to the report of Young and Judd (1978). Analysis of both the locomotor-activity and eclosion phenotypes of 3B1-2 chromosome aberrations did not uncover differences in the genetic control of these two rhythms. The clock phenotypes of 3B1-2 chromosome aberrations, the three per mutant alleles, and per ⁺ duplications suggest that mutations at the per locus shorten, lengthen, or eliminate periodicity by respectively increasing, decreasing, or eliminating per activity.     

Evidence for Heterochromatin Involvement in Chromosome Breakage in Maize Callus Culture

ng from delayed separation of chromatids and typical bridgeswere observed in Feulgen preparations. The analysis of C-bandedanaphases showed that delayed chromatids were held togetherat heterochromatic knob sites (primary event), and the presenceof typical bridges with and without bands corresponding to knobs.These events suggest the occurrence of breakage-fusion-bridge(BFB) cycles initiated by chromosome arms broken during theprimary event. Additional evidence for such a mechanism wasthe presence of gross aberrations involving chromosome 7, detectedin several C-banded metaphases of some cultures. It is hypothesizedthat such aberrations are duplication deficiencies producedby BFB cycles and chromosome healing that would have occurredafter some cell divisions. Zea mays L.; maize; tissue culture; chromosome breakage; heterochromatin; C-banding     

Scanning electron microscopy of isolated Chironomus polytene chromosomes

Individual polytene chromosomes have been isolated from Chironomus stigmaterus for scanning electron microscope observations. The three dimensional ultrastructure of these chromosomes consists of a series of chromatin strands extended in the interbands and more tightly coiled or folded in the banded regions. The nucleolus is observed to be a dense disc or doughnut shaped structure surrounding the chromosome while the Balbiani Rings appear as diffuse regions consisting of both fibrillar and granular elements.     

Comparison of the karyotypes ofPsathyrostachys juncea andP. huashanica (Poaceae) studied by banding techniques

The karyotypes ofP. juncea (Elymus junceus) andP. huashanica (both outbreeders) were investigated by Feulgen-staining and by C-, N-, and Agbanding, based on a single plant in cach case. Both species have 2n=2x=14 and large chromosomes, possibly a generic character. The karyotype ofP. juncea has 8 metacentrics and 6 SAT-chromosomes with minute, heterochromatic satellites while that ofP. huashanica has 9 metacentrics and 5 SAT-chromosomes only, 2 of which with small, heterochromatic satellites. The C-banding patterns ofP. juncea chromosomes comprise from one to five, mostly small, bands at distal, and terminal positions, while those ofP. huashanica chromosomes are characterized by large telomeric bands in most arms. Banding patterns and chromosome morphology allow identification of the homologues of the seven chromosome pairs inP. juncea, but of two pairs inP. huashanica only. The patterns of both taxa are polymorphic, supporting that both taxa are outbreeders. The karyotypic characters suggest thatP. juncea is more closely related toP. fragilis than either is toP. huashanica. N-banding stains weakly. Silver nitrate staining demonstrates that nucleolus organizers of both species have different nucleolus forming capacities. The presence of micronucleoli suggests that both species have an extra unidentified chromosome with nucleolus forming capacity.     

Actinomycin D effects on mitosis and chromosomes: sticky chromatids and localized lesions

When Indian muntjac and Chinese hamster cells in culture were treated with Actinomycin D (1 g/ml) for 1–2 hours, the sister chromatids, especially the distal segments, appeared to have difficulty separating in anaphase. The separated proximal segments progressively became stretched. The nucleolus organizer regions seemed to be most susceptible to stretching, and breaks in these regions were frequently observed. Electron microscopic observations showed that the sticky chromatids (and less frequently sticky chromosomes) contain connecting submicroscopic chromosome strands. When the treated cells were allowed to grow in a drug-free medium for several days, a high frequency of endoreduplicated mitotic figures was found. Chromosome and chromatid breaks and other aberrations were common, mainly localized at G band negative areas particularly nucleolus organizer regions.     

Nucleolar organizing chromosomes ofRicinus

Summary Pachytene chromosome morphology was compared in nine races ofRicinus communis L. (2n = 20), using pollen mother cells (PMCs) and light microscopy. Of the ten bivalents, only the two possessing nucleolar organizing regions (NORs), chromosomes 2 and 7, exhibit structural variations among the races. The NORs are located in the short arms of these two chromosomes. Most of the observed structural variations affect these short arms, which are similar morphologically and consist largely of heterochromatic segments. The PMCs contain a single nucleolus and this is associated with the NOR of each of the two chromosomes at a particular frequency in each race. In eight races, a nucleolar constriction (NC) is present in either chromosome 2 or chromosome 7. In these races, the nucleolus is associated with the chromosome possessing an NC at a frequency of 100% and with the chromosome lacking an NC at a frequency ranging between 5.6 and 100%, depending upon the race. No microscopically visible NC is present in the ninth race. In this race, the nucleolus is associated with both chromosomes 2 and 7 at a frequency of 100%. The association of the nucleolus with a chromosome possessing an NC is at the NC and with a chromosome lacking an NC is at the terminal heterochromatic segment of the short arm. Several interpretations are offered to account for the variations in frequency of association between the nucleolus and each of the nucleolar organizing chromosomes. It is suggested that the two non-linked NORs have evolved through some intragenomic changes rather than polyploidy, that this species is highly intolerant to structural variations other than those occurring in or near the NORs, and that structural variations in the nucleolar organizing chromosomes are not associated with racial variations in plant phenotype.Paper of the Journal Series, New Jersey Agricultural Experiment Station