Chromosome banding in amphibia

The distribution of constitutive heterochromatin on the chromosomes of Triturus a. alpestris, T. v. vulgaris and T. h. helveticus (Amphibia, Urodela) was investigated. Sex-specific chromosomes were determined in the karyotypes of T. a. alpestris (chromosomes 4) and T. v. vulgaris (chromosomes 5). The male animals have one heteromorphic chromosome pair, of which only one homologue displays heterochromatic telomeres in the long arms; the telomeres of the other homologue are euchromatic. This chromosome pair is always homomorphic and without telomeric heterochromatin in the female animals. There is a highly reduced crossing-over frequency between the heteromorphic chromosome arms in the male meiosis of T. a. alpestris; in T. v. vulgaris no crossing-over at all occurs between the heteromorphic chromosome arms. No heteromorphisms between the homologues exist on the corresponding lampbrush chromosomes of the female meiosis. In T. h. helveticus no sex-specific heteromorphism of the constitutive heterochromatin could be determined. The male animals of this species, however, already possess a chromosome pair with a greatly reduced frequency of chiasma-formation in the long arms. The C-band patterns and the pairing configurations of the sex-specific chromosomes in the male meiosis indicate an XX/XY-type of sex-determination for the three species. A revision of the literature about experimental interspecies hybridizations, gonadic structure of haploid and polyploid animals, and sex-linked genes yielded further evidence in favor of male heterogamety. The results moreover suggest that the heterochromatinization of the Y-chromosome was the primary step in the evolution of the sex chromosomes.     

Polymorphic patterns of heterochromatin distribution in guinea pig chromosomes

The chromosome complement and patterns of heterochromatin distribution (as demonstrated by the DNA d-r method) were studied from three different guinea pigs. Karyotype analyses showed that one of the females had a heteromorphic sex pair formed by a submetacentric X chromosome and a subterminal X chromosome originated by a shortening of the short arm (x-chromosome). The heterochromatin was mainly found in the pericentromeric areas of the autosomes and X chromosomes and in the short arm of pair 7. The Y chromosome exhibited a degree of heterochromatinization different from that of pericentromeric areas.—The analysis of the heterochromatin distribution in the X chromosomes showed that the smaller size of the heteromorphic x-chromosoine was probably due to a lack of heterochromatin in its short arm. Moreover, two out of the three animals studied had a heteromorphic pattern of heterochromatinization in the pair 21 characterized by heterochromatinization of the pericentromeric area in one chromosome and almost complete heterochromatinization of the other homologue.—It is suggested that most of the heterochromatin disclosed by the DNA d-r method is formed by repetitious DNA; and that the Y chromosome and perhaps some autosome regions in guinea pigs are formed by a type of heterochromatin with properties different from those of the constitutive and facultative heterochromatin (intermediate heterochromatin).Supported in part by NIH Grant 5-501-RR05672-02 and by NIH contract 70-2299.     

Asynchronous DNA Synthesis in a Duplicated Chromosomal Region of <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

THERE is a highly ordered temporal sequence in the replication of DNA in the polytene chromosomes of Drosophila^1–10. The mechanism underlying this replicative organization remains unknown, but it has been shown that homologous chromosome regions replicate their DNA synchronously whether or not they are paired¹¹ and, in the one case in which it has been studied, this synchrony remains evident even when one of the two homologous regions is translocated to an abnormal position¹². These observations suggest that an essential part of the system controlling replication pattern is located in each of the small chromosome regions, replication of which can be resolved autoradiographically. The simplest model consistent with these assumptions involves a chromosome constituted of numerous “replicons” with replication times geared to a common control mechanism but are independent of the anatomical ordering of the “replicons” within the genome.     

A test of homology between the B chromosome of maize and abnormal chromosome 10, involving the control of nondisjunction in B's

Summary Nondisjunction of B and B-translocation chromosomes occurs regularly in maize at the second pollen mitosis (Roman, 1947; Blackwood, 1956). The mechanism of nondisjunction was studied using the A-B interchange, TB-9b. The B⁹ chromosome of the interchange undergoes nondisjunction at the second pollen mitosis, while the 9^B chromosome does not (Roman, 1947). It was shown that the 9^B chromosome must be present in a plant for nondisjunction of the B⁹ to occur. This is consistent with the reports of Roman on TB-4a (1949) and Longley on TB-10a (1956). It was also demonstrated that the influence of the 9^B chromosome is limited to pollen grains containing it, and does not extend to all the pollen of a plant.A test of homology between the B chromosome and abnormal chromosome 10 was also made. The ability of abnormal 10 to substitute for the 9^B chromosome and induce nondisjunction of the B⁹ was tested. Nondisjunction did not occur at a detectable rate in the presence of abnormal 10, and the results failed to support Ting's proposal (1958) concerning the origin of abnormal 10.     

Kinetics of DNA replication in the Indian muntjac chromosomes as studied by quantitative autoradiography

DNA replication patterns of individual chromosomes and their various euchromatic and heterochromatic regions were analyzed by means of quantitative autoradiography. The cultured cells of the skin fibroblast of a male Indian muntjac were pulse labeled with ³H-thymidine and chromosome samples were prepared for the next 32 h at 1–2 h intervals. A typical late replication pattern widely observed in heterochromatin was not found in the muntjac chromosomes. The following points make the DNA replication of the muntjac chromosomes characteristics: (1) Heterochromatin replicated its DNA in a shorter period with a higher rate than euchromatin. (2) Two small euchromatic regions adjacent to centromeric heterochromatin behaved differently from other portions of euchromatin, possessing shorter Ts, higher DNA synthetic rates and starting much later and ending earlier their DNA replication. (3) Segmental replication patterns were observed in the chromosomes 2 and 3 during the entire S phase. (4) Both homologues of the chromosome 3 showed a synchronous DNA replication pattern throughout the S phase except in the distal portion of the long arms during the mid-S phase.     

DNA replication patterns in somatic chromosomes of Leptodactylus ocellatus (Amphibia,Anura)

The morphology and pattern of replication in the somatic chromosomes of Leptodactylus ocellatus (Amphibia, Anura) was studied by means of H³-thymidine autoradiography. A total of 300 metaphases from leukocyte cultures and 200 metaphases from spleen cell cultures were analysed.The diploid chromosome number in Leptodactylus ocellatus is 22. The pairs 1, 2, 3, 4, 7 and 8 could be easily identified on the basis of their size, centromere position, and location of secondary constrictions. In 30% of metaphases the pair 10 could be recognized on account of an end-to-end homologous association, which originated from a satellite fusion.The continuous H³-thymidine labelings carried out in the last 10, 5 and 3 hours of a culture indicated that the G₂ period was 3.5 hours. The labeled metaphases were divided in two groups. In the first one all those cells showing radioactivity along the entire length of every chromosome were included. The second group was formed by metaphases with extensive unlabeled chromosome regions. The former and the latter group were identified as representatives of the intermediate and final stages of the S period, respectively.The pattern of chromosome labeling indicates that secondary constrictions are associated with late replicating regions. However, the presence of chromosome areas, which in spite of being late in finishing duplication did not bear any kind of constriction, suggests that regions other than those associated with constrictions also may replicate late. No interchromosomal asynchrony of replication at the end of the S period was noticed. However, very often in pair 10 one chromosome had about two times as much labeling as its homologue. No sex-linked differences in chromosome morphology or in patterns of chromosome replication could be noticed.     

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).      

X-inactivation patterns in lymphocytes and skin fibroblasts of three cases of X-autosome translocations with abnormal phenotypes

Summary X-inactivation patterns were studied by replication analyses both in lymphocytes and skin fibroblasts of two patients carrying balanced X-autosome translocations, t(X;10)-(pter;q11) and t(X;17)(q11;q11), and one patient with an unbalanced translocation t(X;22)(p21;q11). Preferential late replication of the normal X chromosome was found in lymphocytes of both patients carrying balanced translocations and in skin fibroblasts of the patient carrying the translocation t(X;17). However, skin fibroblasts of the patient with a translocation t(X;10) showed preferential late replication of the abnormal der(X) chromosome with no spreading of late replication to the autosomal segment. In the case of unbalanced translocation t(X;22) there was preferential late replication of the der(X) chromosome both in lymphocytes and skin fibroblasts. The abnormal phenotype of the patients is discussed in relation to the observed X-inactivation patterns and the variability of the patterns in different tissues.     

Karyotypic comparison among Cebuella pygmaea,Callithrix jacchus and C. emiliae (Callitrichidae,Primates) and its taxonomic implications

The karyotype of Cebuella pygmaea (2n=44) obtained by G-, C-banding, and NOR-staining is described. This species presents a heteromorphic C band in the intersticial region of the short arm of chromosome 2. The data obtained were compared with those previously described for the karyotypes of Callithrix jacchus and Callithrix emiliae. The three species differ in the amount and distribution of non-centromeric constitutive heterochromatin. The importance of the variation in constitutive heterochromatin for the phylogeny of the group is discussed. Comparison of the karyotypes in terms of G-banding patterns showed that C. pygmaea and C. emiliae differ from C. jacchus by a Robertsonian translocation and a paracentric inversion, whereas C. pygmaea and C. emiliae differ from each other by a reciprocal translocation between an acrocentric autosome and the short arm of the submetacentric chromosomes that distinguishes their karyotypes from that of C. jacchus. The possible evolutionary paths followed by the karyotypes of the three species are discussed.     

Rye chromosome translocations in hexaploid wheat: a re-evaluation of the loss of heterochromatin from rye chromosomes

Summary Using in situ hybridization techniques, we have been able to identify the translocated chromosomes resulting from whole arm interchanges between homoeologous chromosomes of wheat and rye. This was possible because radioactive probes are available which recognize specific sites of highly repeated sequence DNA in either rye or wheat chromosomes. The translocated chromosomes analysed in detail were found in plants from a breeding programme designed to substitute chromosome 2R of rye into commercial wheat cultivars. The distribution of rye highly repeated DNA sequences showed modified chromosomes in which (a) most of the telomeric heterochromatin of the short arm and (b) all of the telomeric heterochromatin of the long arm, had disappeared. Subsequent analyses of these chromosomes assaying for wheat highly repeated DNA sequences showed that in type (a), the entire short arm of 2R had been replaced by the short arm of wheat chromosome 2B and in (b), the long arm of 2R had been replaced by the long arm of 2B. The use of these probes has also allowed us to show that rye heterochromatin has little effect on the pairing of the translocated wheat arm to its wheat homologue during meiosis. We have also characterized the chromosomes resulting from a 1B-1R translocation event.From these results, we suggest that the observed loss of telomeric heterochromatin from rye chromosomes in wheat is commonly due to wheat-rye chromosome translocations.     

Sex chromosome evolution in fish: the formation of the neo-Y chromosome in Eigenmannia (Gymnotiformes)

Chromosomes of a species of Eigenmannia presenting a X₁X₁X₂X₂:X₁X₂Y sex chromosome system, resulting from a Y-autosome Robertsonian translocation, were analyzed using the C-banding technique, chromomycin A₃ (CMA₃) and mithramycin (MM) staining and in situ digestion by the restriction endonuclease AluI. A comparison of the metacentric Y chromosome of males with the corresponding acrocentrics in females indicated that a C-band-positive, CMA₃/MM-fluorescent and AluI digestion-resistant region had been lost during the process of translocation, resulting in a diminution of heterochromatin in the males. It is hypothesized that the presence of a smaller amount of G+C-rich heterochromatin in the sex chromosomes of the heteromorphic sex when compared with the homomorphic sex may be associated with the sex determination mechanism in this species and may be a more widely occurring phenomenon in fish with differentiated sex chromosomes than was initially thought. Received: 1 April 1999; in revised form: 16 October 1999 / Accepted: 4 December 1999     

The pattern of DNA replication in the chromosomes of Puschkinia libanotica

The uptake of H³-thymidine into the chromosomes of Puschkinia libanotica has been studied in plants possessing or lacking a heterochromatic B chromosome. The pattern of H³-thymidine uptake by the A chromosomes at the end of the S phase is similar in plants of both genotypes. Regions around the centromere take up more H³-thymidine at the end of S than do more distal regions. The rate of uptake into the heterochromatin of the B chromosome increases towards the end of S, but there is no evidence that synthesis in the B chromosome carries on after the completion of DNA synthesis in the euchromatic A complement. It is proposed that at the end of the S phase more replicons in the heterochromatin of the B chromosome are engaged in DNA synthesis than in euchromatin.     

Chromosome 1 in crested and marbled newts (Triturus)

The heteromorphic chromosomes 1 of Triturus cristatus carnifex and T. marmoratus were studied in mitotic metaphase after staining with the Giemsa C-banding technique and with the fluorochromes, DAPI (AT-specific) and mithramycin (GC-specific). They were also examined in the lampbrush form under phase-contrast before fixation and after fixation and staining with Giemsa. Chromosomes 1 of T.c. carnifex are asynaptic and achiasmatic throughout most of their long arms. They are also heteromorphic in most of their long arms for the patterns of Giemsa and fluorochrome staining and the distribution of distinctive lampbrush loops. The heteromorphic regions correspond to the regions that are asynaptic and achiasmatic. They stain more strongly with mithramycin and more weakly with DAPI than the remainder of the chromosomes, signifying that their DNA is relatively rich in GC. The patterns of staining with Giemsa and fluorochromes and the distributions of distinctive lateral loops vary from one animal to another in the same species and even in the same population. The asynaptic and achiasmatic regions of chromosomes 1 in T. marmoratus extend throughout the whole of the long arms and well beyond the heterochromatic region. Chiasmata form only in the short arm and occasionally in the short euchromatic segment at the tip of the long arms. The staining patterns of chromosomes 1 in T. marmoratus differ from those in T.c. carnifex although, like carnifex, their DNA is relatively GC-rich. The chromosomes 1 of T. marmoratus are more submetacentric than those of T.c. carnifex. In T. marmoratus chromosome 1B is about 12% shorter than 1A. There is a short paracentric inversion heterozygosity in the long arm of chromosome 1B in T. marmoratus which probably accounts for the lack of chiasmata in the euchromatin that separates the centromere from the start of the heterochromatin. In both carnifex and marmoratus, embryos that are homomorphic for chromosome 1 arrest and die at the late tailbud stage of development. The same applies to F₁ hybrid embryos T.c. carnifex x T. marmoratus, and this has permitted identification of chromosomes 1A and 1B in both species. There is no correspondence between patterns of Giemsa or fluorochrome staining of the heteromorphic regions of chromosome 1 and any feature of the lampbrush chromosomes. However, the short euchromatic ends of the long arms of chromosomes 1 in both species are distinguished in the lampbrush form by a series of uniformly small loops of fine texture associated with very small chromomeres. The Giemsa C-staining patterns of both chromosomes 1A and 1B are different in each of the four subspecies of T. cristatus. T.c. karelinii stands out by having unusually large masses of Giemsa C-staining centromeric heterochromatin on all but 1 of its 12 chromosomes. A scheme is proposed for the evolution of chromosome 1 in T. cristatus and T. marmoratus, based on all available cytological and molecular data.     

Inheritance of chromosome heteromorphisms analyzed by high-resolution bivariate flow karyotyping.

The DNA content of the mitotic chromosomes from 10 children and their parents in four families were quantified by bivariate flow karyotyping. In all cases, each chromosome peak in the flow karyotype of the child could be traced to one of the two parents. The measured absolute difference in homologue DNA content between children and their parents averaged 0.8%, or approximately 1 Mbp over all chromosome types. This study demonstrates that flow karyotypes of a proband's parents can be an aid in detecting and quantifying the size of de novo deletions that involve heteromorphic chromosome types.     

Transmission and phenotypic effect of a duplicate chromosome segment in maize

Summary The chromosome B⁴, extracted from the translocation TB-4a (involving chromosome 4 and a B chromosome) was transferred into stocks with normal complement. This chromosome carried 75% of the short arm of chromosome 4 and was provided with a B centromere. Loss in somatic tissues, in meiotic divisions and through gametophyte competition in the pollen was investigated by cytological and genetical means. Nondisjunction in the second microspore division of the B⁴, in the presence of a normal chromosome 4, was not frequently observed. Sectoring in endosperm tissues, after appropriate crosses, presumably indicated either late replication of this chromosome and loss during endosperm development, and/or inactivation of the Su locus which is near the breakage point of the translocation with the B chromosome. Reduced vigor of the plants carrying one or two B⁴ chromosomes was interpreted as an effect of the duplication. There are indications that hyperploidy for this specific region may affect the kernel size and weight.This work was supported by C.N.R., N.A.T.O. and Indiana University funds.     

Demonstration of replication patterns in the last premeiotic S-phase of male Chinese hamsters after BrdU pulse labeling

Chromosome replication in the last premeiotic S-phase of male mammals has been previously studied by [³H]thymidine autoradiography and by a 5-bromodeoxyuridine (BrdU)/Giemsa technique. We used a recently developed BrdU-antibody technique (BAT) in this study. The following conclusions were drawn: (1) The replication patterns observed are similar to that of somatic cells. (2) The heterochromatin starts replication in early S-phase. (3) The euchromatic part of the X chromosome of the male Chinese hamster replicates together with the autosomes and therefore behaves isocyclicly and not allocyclicly as hitherto assumed. Hence, genetic inactivity of the X chromosome may be brought about by a mechanism different from that in somatic cells.by P.B. Moens     

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.     

Asynchronous replication of constitutive heterochromatin on X chromosomes in female Mus dunni

Euchromatin DNA of one X chromosome in mammalian females, which becomes facultatively heterochromatinized, is known to replicate asynchronously late in S phase compared to its active homologue. In the females of a pygmy mouse species Mus dunni, which has prominent segment of constitutive heterochromatin as the short arm of its submetacentric X chromosome, we have observed asynchronous replication of c-heterochromatin arm as well, predominant number of cells showing the segment associated with the facultatively heterochromatic X to be terminating later. The preferential later termination of replication of the c-heterochromatic arm on the lyonized X appears to be due to the influence of facultative heterochromatin on the adjacent constitutive heterochromatin.     

Dauer der DNS-Replikation von Eu- und Heterochromatin bei Microtus agrestis

The duration of DNA replication of eu- and heterochromatin in kidney epithelial cell cultures of female Microtus agrestis was determined with combined H³-thymidine pulse labelling and cytophotometric determination of Feulgen DNA. The average duration of the total cell cycle was 23.3 hrs, with a G1 period of 14.6 hrs, S period of 5 hrs, G2 period of 2.7 hrs, and mitosis of 1 hr. The replication time of eu- and heterochromatin was determined by the frequency of the different labelling patterns after pulse labelling. The time sequence of the labelling patterns was ascertained by DNA measurements. During the S period, euchromatin replicates at first alone for 3 hrs (60% of the length of S) and 1 hr (19.3%) together with heterochromatin. During the last hour (20.7%), only heterochromatic regions replicate. The sex chromatin part of the one X chromosome starts synthesis 20 minutes (7.3% of S) before the remainder of the heterochromatic X material and ends 30 minutes (9.7% of S) prior to the termination of the S period. Replication of euchromatin takes about 80% of the duration of the total S period, whereas that of heterochromatin takes only 40%.

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