Nucleotide composition of the cold-sensitive heterochromatin regions of Paris hainanensis Merrill chromosomes

Cold-induced decondensation of heterochromatic regions (CSR-bands) in Paris hainanensis (= Daiswa hainanensis Merrill Takht.) (2n = 10; 10 + b) was studied. The comparison of CSR-banding patterns with those obtained by nucleotide-specific staining with fluorochromes DAPI and chromomycin A3 demonstrated that low temperatures induced decondensation only of large AT-rich heterochromatic regions. It is suggested that this is characteristic of all plant species.     

Early replication banding in <Emphasis Type="Italic">Leporinus</Emphasis> species (Osteichthyes,Characiformes) bearing differentiated sex chromosomes (ZW)

There are few examples of differentiated sex chromosomes in fishes. In the genus Leporinus, seven species present a highly differentiated ZW system, derived from heterochromatinization process. Cytogenetic analyses carried out in three of these fish species, Leporinus obtusidens, L. elongatus and L. reinhardti, through RBG-banding, showed late replication bands, coincident with heterochromatic regions in both Z and W chromosomes. A similar interstitial early replication segment was observed in the complex heterochromatic region along the Wq arms in the three species, which might correspond to a pseudoautosomal segment (SD, sex determining locus). Asynchrony related to the replication pattern among different Z chromosomes was not observed. When the identification of nuclear organizer regions by silver nitrate was performed over chromosomal preparations previously exposed to 5-bromo-2′-deoxyuridine (BrdU), remarkable positive signals at interstitial and telomeric position were observed on the q arms of W chromosomes in the species L. elongatus and L. reinhardti. The absence of 18S ribosomal RNA gene loci in this region, formerly demonstrated by FISH, indicates that this argentophilic behavior is putatively due to heterochromatin decondensation caused by BrdU incorporation, favoring such Ag+ reaction. Early and late replication bands were also observed in the heterochromatic portions of Z and W chromosomes, indicating that euchromatic and heterochromatic regions are interspersed. The present data suggest a significant level of heterochromatic complexity in the sex chromosomes of each species. On the other hand, the replication pattern shared by them supports a monophyletic origin.     

Early replicating DNA in heterochromatin ofPlagiochila ovalifolia (Liverworts)

The timing of DNA replication of heterochromatin in malePlagiochila ovalifolia was investigated by the use of³H-thymidine autoradiography. The estimated duration of the mitotic cycle was as follows: S period, 19 hr: G₂+prophase, 10 hr; G₁+meta-, ana-, telophase, 5 hr; total mitotic cycle, 34 hr. The first appearance of silver grains over the chromosomes was observed at 8 hr after the beginning of pulse labelling at which time the silver grains were only over the euchromatic regions, not over the heterochromatic regions. This labelling pattern was also observed at 10 to 15 hr. The heterochromatic regions having more grains than the euchromatic regions were observed at 20 to 25 hr. These results show that the DNA of the heterochromatin of this species is replicated earlier than the euchromatin.     

Contrasting response of euchromatin and heterochromatin to translocation in polytene chromosomes of Drosophila melanogaster

Studies on Feulgen-DNA content in the polytene chromosomes of D. melanogaster T(14)w ^m258-21 heterozygotes showed that when the euchromatic region 3D₁-E₂ is located next to the heterochromatic breakpoint it contains less DNA than in the non-translocated homologue (Hartmann-Goldstein and Cowell, 1976). In contrast to the region adjacent to the breakpoint, region 3C_1–10, which contains intercalary heterochromatin, shows more DNA in the translocated than in the non-translocated chromosome. Transposition may induce morphologically euchromatic regions containing putatively underreplicated sequences to undergo additional replication cycles. Region 2E₁-3A₄, distal to 3C₁ and at some distance from the heterochromatic breakpoint is apparently unaffected. Extended replication and reduced DNA content in regions which have undergone chromosomal rearrangement could be accounted for by varying degrees of blockage of replication in individual strands of the polytene chromosome.     

In vitro decondensation of the sperm chromatin in Holothuria tubulosa (sea cucumber) not affecting proteolysis of basic nuclear proteins

Sea urchin and sea star oocyte extracts contain proteolytic activities that are active against sperm basic nuclear proteins (SNBP). This SNBP degradation has been related to the decondensation of sperm chromatin as a possible model to male pronuclei formation. We have studied the presence of this proteolytic activity in Holothuria tubulosa (sea cucumber) and its possible relationship with sperm nuclei decondensation. The mature oocyte extracts from H. tubulosa contain a proteolytic activity to SNBP located in the macromolecular fraction of the egg‐jelly layer. SNBP degradation occurred both on sperm nuclei and on purified SNBP, histones being more easily degraded than protein Ø_o (sperm‐specific protein). SNBP degradation was found to be dependent on concentration, incubation time, presence of Ca²⁺, pH, and this activity could be a serine‐proteinase. Thermal denaturalization of the oocyte extracts (80°C, 10–15 min) inactivates its proteolytic activity on SNBP but does not affect sperm nuclei decondensation. These results would suggest that sperm nuclei decondensation occurs by a mechanism different from SNBP degradation. Thus, the sperm nuclei decondensation occurs by a thermostable factor(s) and the removal of linker SNBP (H1 and protein Ø_o) will be a first condition in the process of sperm chromatin remodeling.     

Isolation and characterization of microsatellite markers for Cycas hainanensis C. J. Chen (Cycadaceae)

Cycas hainanensis is an endangered cycad species endemic to Hainan Island, China. To enrich our scientific conservation for this species, we developed eight microsatellite markers using repetitive DNA enriched libraries. The number of alleles per microsatellite locus varied from 8 to 17. The expected (H _E ) and observed (H _O ) heterozygosities varied from 0.4754 to 0.8846 and from 0.3636 to 0.9600, respectively. These markers will be employed to determine whether the ex situ C. hainanensis individuals in Sourh China Botanical Garden capture a representative portion of genetic diversity of the wild populations.     

The effect of JIL-1 on position-effect variegation is proportional to the total amount of heterochromatin in the genome

In this study we have taken advantage of recent whole genome sequencing studies that have determined the DNA content in the heterochromatic regions of each Drosophila chromosome to directly correlate the effect on position-effect variegation of a pericentric insertion reporter line, 118E-10 with the total amount of heterochromatic DNA. Heterochromatic DNA levels were manipulated by adding or subtracting a Y chromosome as well as by the difference in the amount of pericentric heterochromatin between the X and Y chromosome. The results showed a direct, linear relationship between the amount of heterochromatic DNA in the genome and the expression of the w marker gene in the 118E-10 pericentric reporter line and that increasing amounts of heterochromatic DNA resulted in increasing amounts of pigment/gene activity. In Drosophila heterochromatic spreading and gene silencing is counteracted by H3S10 phosphorylation by the JIL-1 kinase, and we further demonstrate that the haplo-enhancer effect of JIL-1 is proportional to the amount of total heterochomatin, suggesting that JIL-1's activity is dynamically modulated to achieve a more or less constant balance depending on the levels of heterochromatic factors present.     

极小种群野生植物海南风吹楠的遗传多样性研究

为探讨海南风吹楠(Horsfieldia hainanensis)的濒危原因,利用限制性酶切位点相关的DNA测序技术(RAD-seq)开发单核苷酸多态性(SNPs),评估居群的遗传多样性和遗传结构.结果表明,海南风吹楠的遗传多样性较低(Ho=0.167),其中BWL居群表现出最高的遗传多样性;居群间存在中等程度的遗传分...     

The heterochromatin regions of the mitotic chromosomes in cattle]

Cytochemical and molecular peculiarities of heterochromatic regions of bovine chromosomes have been studied, using specific fluorochrome staining induced decondensation, in situ hybridization, pretreatment of restriction enzymes. The heterochromatin of autosomes demonstrated a strong homogeneity. In chromosome Y two small specific heterochromatic regions were found lacking a long repeated tandem block of nucleotides enriched in GC base pairs and having no tandem block of Bkm repeats (10(4) b.p.). This class repeats are probably interspersed in the bovine genome. A rather seldom character of mammalian karyotypes is the absence of cytochemical heterochromatin in the X chromosome.     

Genotypes affecting the condensation and transmission of heterochromtic B chromosomes in the mealybug Pseudococcus affinis

In male mealybugs (Pseudococcide: Homoptera) the set of chromosomes of paternal origin becomes heterochromatic (H) and genetically inactive in early embryogenesis. During spermatogenesis the two sets segregate and only the meiotic products with the euchromatic (E) set form sperm. Individuals of Pseudococcus affinis (Maskell) may carry a B chromosome (B) which is usually heterochromatic. During prophase I of spermatogenesis, however, the B becomes even less condensed than the E set and usually segregates with the E set. We have previously shown that natural populations of P. affinis contain genotypes that can reduce the rate of transmission (k) of Bs from more than 0.9 to less than 0.1. We now demonstrate that these genotypes suppress k either by enhancing or by preventing the decondensation of the B, which in turn affects the position of the B on the metaphase plate and its segregation. We also demonstrate that radiation-induced fragment of the B, and a piece of a B which has been translocated onto a piece of an H chromosome, retain their characteristic pattern of condensation and thus that the condensation of the B is not controlled by one or a few cis-acting centers or loci.     

Pseudoterminalisation,terminalisation, and non-chiasmate modes of terminal association

Terminal associations occur commonly between meiotic homologues of the two smallest (S10, S11) chromosomes in the northern race of Cryptobothrus chrysophorus when they are either heterozygous or homozygous for distal supernumerary heterochromatic segments. A detailed examination of the origin and behaviour of these associations provides convincing evidence that they are non-chiasmate in character and so cannot be explained by either pseudoterminalisation or terminalisation. The same is true of the terminal associations involved in the persistent pseudomultiples that develop between non-homologues of Heteropternis obscurella when one or both of these carry distal heterochromatic segments. In both situations the C-bands involved in such terminal associations are entire and are never interrupted by non-banded material. In Cryptobothrus, similar associations can also develop between centromere regions when these are heterozygous or homozygous for proximal supernumerary heterochromatic segments.     

Differentielle Darstellung der Metaphasechromosomen von Cypripedium debile mit Chinacrin- und Giemsa-Färbung

The somatic metaphase chromosomes of C. debile stain differentially with quinacrine and Giemsa respectively. After quinacrine staining the heterochromatic regions show fainter fluorescence than euchromatic regions. With Giemsa the heterochromatic regions are more deeply stained than the euchromatic regions. This is true also for slides which were pre-treated with trypsin or trichloroacetic acid. From these findings it may be supposed, that the heterochromatic regions are G-C rich. Furthermore, differences in protein composition are expected between hetero- and euchromatic regions. The results of differential staining of metaphase chromosomes of C. debile, including those which were published in previous papers, are summarized here.     

Differential sensitivity of constitutive and facultative heterochromatin in orthopteran chromosomes to digestion by DNaseI

In situ pancreatic DNaseI digestions were used as probes to study the structural organization of facultative and constitutive heterochromatin during both mitotic and meiotic divisions. Three different types of heterochromatic regions from three insect species were chosen for this study. These regions had been previously characterized by in situ treatments with restriction endonucleases (AT and GC rich DNA sequences). Progressive increase in DNaseI concentration (from 10 to 200 ng/ml) or in incubation time (from 5 to 30 min) revealed a specific pattern of sequential digestion of the constitutive heterochromatic regions, the centromeric ones (AT-rich DNA) being the most resistant to DNaseI action. The interstitial C-bands (with AT or GC-rich DNA) were more sensitive to DNaseI, and the band 4.4 from Baetica ustalata was the most resistant of the non-centromeric bands. Similar results were obtained during meiosis, but increased accessibility to DNAseI was observed compared to mitosis. DNA methylation in the non-centromeric band 4.4 of B. ustulata could be responsible for its differential digestion with respect to the remaining intercalar heterochromatin. Facultatively heterochromatic regions (X chromosomes) were found to exhibit a differential response to DNaseI attack from mitosis to meiosis. While they behaved as cuchromatin during mitosis, they were the most resistant together with centromeric heterochromatin regions, during metaphase I and II. The different responses to digestion of the X chromosome and X-derived regions between somatic and meiotic divisions are probably a consequence of the changes in the organization of this chromosome during the facultative heterochromatinization process.     

Inhibition of cation-induced DNA condensation by intercalating dyes

Several intercalating dyes are shown to inhibit the cation-induced condensation of λ-DNA when Co³⁺(NH₃)₆ is the condensing agent. The dyes that have been studied are ethidium, propidium, proflavin, quinacrine, and actinomycin D. Earlier work has shown that intercalating dyes inhibit ψ-DNA condensation. [Lerman, L. S. (1971) Prog. Mol. Subcell. Biol. 2 , 382–391; Cheng, S. & Mohr, S. C. (1975) Biopolymers 14 , 663–674.] Dye-induced decondensation of intramolecularly condensed DNA has been studied by making use of conditions in which Co³⁺(NH₃)₆ produces intramolecular condensation without significant aggregation. Some aggregation is caused, however, during dye-induced decondensation. Dye titration curves of DNA decondensation have been measured by excess light scattering to monitor decondensation and by fluorescence to monitor intercalation. All of the dyes studied act as competing cations in displacing the condensing cation Co³⁺(NH₃)₆ from the DNA. Competition occurs both in and below the transition zone for condensation. The effectiveness of a dye as a competing cation increases with its net positive charge. Before decondensation begins, no intercalated dye can be detected, suggesting that intercalation might be incompatible with the proper helix packing needed for cation-induced DNA condensation. To test this last point, methidium–spermine was synthesized: it contains an intercalating methidium head group combined with a polyamine tail. Methidium–spermine is found to cause λ-DNA condensation, but aggregation accompanies condensation, as has been found earlier for spermine and spermidine. Fluorescence and absorption spectra indicate that the methidium group is intercalated when the DNA is condensed, indicating that intercalation need not be incompatible with DNA condensation. The presence of aggregates among the condensed DNA molecules makes this last conclusion tentative.     

A New Retroposed Gene in <Emphasis Type="Italic">Drosophila</Emphasis> Heterochromatin Detected by Microarray-Based Comparative Genomic Hybridization

A genomic pattern of new gene origination is often dependent on a genomic method that can efficiently identify a statistically adequate number of recently originated genes. The heterochromatic regions have often been viewed as genomic deserts with low coding potential and thus a low flux of new genes. However, increasing reports revealed unexpected roles of heterochromatic regions in the evolution of genes and genomes. We identified recently retroposed genes that originated in heterochromatic regions in Drosophila, by developing microarray-based comparative genomic hybridization (CGH) with multiple species. This new gene family, named Ifc-2h, originated in the common ancestor of the clade of D. simulans, D. mauritiana, and D. sechellia. The sequence features and phylogenetic distribution indicated that Ifc-2h resulted from the retroposition from its parental gene, Infertile crescent (Ifc), and integrated into heterochromatic region of common ancester of the three sibling species 2 million years ago. Expression analysis revealed that Ifc-2h had developed a new expression pattern by recruiting a putative regulatory element from its target sequence. The distribution of indel variation in Ifc-2h of D. simulans and D. mauritiana revealed a significant sequence constraint, suggesting that the Ifc-2h gene may be functional. These analyses cast fresh insight into the evolution of heterochromatin and the origin of its coding regions. Electronic Supplementary Material Electronic Supplementary material is available for this article at and accessible for authorised users. [Reviewing Editor: Dr. Martin Kreitman]     

Pseudomultiple production in Ageneotettix deorum deorum

Meiosis in Ageneotettix deorum deorum is characterised by extensive pseudomultiple formation during prophase. The association of non-homologous chromosomes takes place prior to pairing and chiasma formation and occurs to a varying degree in all prophase cells. These pseudomultiples originate during interphase as a consequence of the association of heterochromatin. All autosomes carry procentric heterochromatic segments of variable size and the L₁, L₃ and M₅ chromosomes also possess terminal heterochromatic regions. The association of non-homologous chromosomes during zygotene and pachytene does not appear to impede pairing or the frequency and distribution of chiasmata. — A majority of the pseudomultiples dissociate after diakinesis, during orientation and congression on the spindle. However in 4% of the cells examined, associations, mainly quadrivalents, persist through metaphase. — Heterochromatic associations of non-homologous chromosomes are again evident during second prophase, though here they involve only the centric heterochromatic regions; 9% of these associations persist through second metaphase. — The nature and behaviour of the pseudomultiples in Ageneotettix are pertinent to the interpretation of terminal associations in other Orthoptera and provide evidence that persistent associations can arise following a non-chiasmate association.On educational leave from the Forest Research Laboratory, Fredericton, N.B. Canada.     

The modification of crossing over in maize by extraneous chromosomal elements

Summary Five regions of the maize genome were tested for their response to endogenous factors influencing recombination. These included heterochromatic B chromosomes and abnormal chromosome 10 as well as the sex in which recombination occurred.The frequency of recombination in the proximal A ₂-Bt and Bt-Pr segments of chromosome 5 was increased in the presence of B chromosomes, with the male meiocytes showing a greater response than the female meiocytes. In addition, experiments involving 0, 1, 2 and 4 B's revealed a dosage effect of B chromosomes on crossing over in chromosome 5. Recombination in the proximal Wx-Gl ₁₅ interval of chromosome 9 was found to be slightly higher than normal in male flowers when two B chromosomes were present. This increase was accompanied by a decrease in the adjacent Sh-Wx segment. Crossing over in the distal C-Sh segment and in the C-Sh-Wx-Gl ₁₅ regions of female flowers was unaffected by B's.Comparisons of plants heterozygous for abnormal chromosome 10 (K10 k10) and homozygous for the standard chromosome 10 (k10 k10) showed that abnormal 10 greatly enhances crossing over in the A ₂-Bt and Bt-Pr segments of chromosome 5. In contrast to the finding with B's, the effect is greater in female than in male sporocytes. K10 showed no significant effect on recombination in the C-Sh-Wx-Gl ₁₅ region of chromosome 9 except in male sporocytes, where there was a slight increase in the Sh-Wx region of 0 B K10 k10 plants and a possible interaction with B chromosomes to raise the level of recombination between Wx and Gl ₁₅. The fact that the regions adjacent to the centromere of chromosome 9 show little or no response to the presence of K10 indicates that the proximal heterochromatin of this chromosome differs qualitatively from that of other maize chromosomes. This conclusion is supported by a comparison of the effects of B chromosomes, K10 and sex on crossing over in chromosomes 5 and 9.Dedicated to Dr. M. M. Rhoades on the occasion of his seventieth birthday.     

Orientation of interphase chromosomes as detected by Giemsa C-bands

The orientation of Giemsa C-bands has been studied in mitotic and interphase cells of Allium cepa, A. sativum and of Aloe vera. The C-bands in these three species are located at the telomeres, secondary constriction region of the nucleolar chromosomes and the centromeric regions, respectively. Observations in A. cepa and Aloe indicate clearly that the interphase chromosomes are non-random in their orientation and possibly maintain their telophase configuration through the attachment of telomeres and perhaps of kinetochores with the nuclear membrane. Electron micrographs of onion cells also reveal that certain heterochromatic segments are associated with the nuclear membrane. — The nucleolar interstitial C-bands in A. sativum remain free in the nucleoplasm and may come close to each other due to heterochromatic attraction. Such a heterochromatic attraction is also evident between telomeric regions and between centromeres. However, a two by two attachment could not be noticed. A diagrammatic representation of the orientation of interphase chromosomes has been presented.The major part of this work was presented at the First International Congress on Cell Biology, Boston, Sept. 5–10, 1976 (Platform Session 36, J. Cell Biol. 70, 418a (1976)     

Die intraspezifische Variabilität des heterochromatischen Armes eines Chromosoms bei der GattungCarabus L. (Coleoptera)

The chromosome complement ofC. auronitens Fabr. is 2n _♂=26+XY. One autosomal pair—called A-chromosomes—is relatively long.A-chromosomes consist of a euchromatic and a heterochromatic arm. Labelling of mitotic chromosomes with³H-thymidine shows that replication of the heterochromatic arm continues when it has ended in the euchromatic arm. In males and females the length of the heterochromatic arm varies intraindividually. In 47 of 99 males the heterochromatic arms were heteromorphic. Calculations of the quotient length of the euchromatic/length of the heterochromatic arm have shown that at least 6 different types of the A-chromosome exist. These types differ from each other in the number of heterochromatic sections separated by constrictions. The longest heterochromatic arm observed consisted of 8 such sections. The genetic significance of the heterochromatin in the genus ofCarabus is at present unknown (Zusammenfassung see p.305).