X-Y Exchange and the Coevolution of the X and Y Rdna Arrays in Drosophila melanogaster

The nucleolus organizers on the X and Y chromosomes of Drosophila melanogaster are the sites of 200-250 tandemly repeated genes for ribosomal RNA. As there is no meiotic crossing over in male Drosophila, the X and Y chromosomal rDNA arrays should be evolutionarily independent, and therefore divergent. The rRNAs produced by X and Y are, however, very similar, if not identical. Molecular, genetic and cytological analyses of a series of X chromosome rDNA deletions (bb alleles) showed that they arose by unequal exchange through the nucleolus organizers of the X and Y chromosomes. Three separate exchange events generated compound X·Y ^L chromosomes carrying mainly Y-specific rDNA. This led to the hypothesis that X-Y exchange is responsible for the coevolution of X and Y chromosomal rDNA. We have tested and confirmed several of the predictions of this hypothesis: First, X· Y^L chromosomes must be found in wild populations. We have found such a chromosome. Second, the X·Y^L chromosome must lose the Y^L arm, and/or be at a selective disadvantage to normal X⁺ chromosomes, to retain the normal morphology of the X chromosome. Six of seventeen sublines founded from homozygous X·Y^Lbb stocks have become fixed for chromosomes with spontaneous loss of part or all of the appended Y^L. Third, rDNA variants on the X chromosome are expected to be clustered within the X⁺ nucleolus organizer, recently donated (" Y") forms being proximal, and X-specific forms distal. We present evidence for clustering of rRNA genes containing Type 1 insertions. Consequently, X-Y exchange is probably responsible for the coevolution of X and Y rDNA arrays.     

Ribosomal RNA Cistrons of X Chromosomes Clonally Derived from D. MELANOGASTER Laboratory Populations: Redundancy, Organization and Stability

To determine whether heterogeneity exists in the organization or redundancy of the rRNA cistrons of inbred populations of Drosophila melanogaster, we have derived a number of sublines from the strains Oregon R and Canton S. These two strains were chosen because our previous studies have demonstrated a difference in the competence of these strains to exhibit a "compensatory response" of the rDNA. In each subline, the X chromosomes are descended from a single maternal X, that is, each line is homozygous for a particular nucleolus organizer (NO). These derivative lines have been characterized in terms of rDNA content and organization, using quantitative liquid hybridizations and Southern blot analyses, respectively. Our studies reveal that both of the highly inbred parent populations contained a heterogeneous array of X chromosomal rDNA contents. Once isogenized, the rDNA redundancy of a given X chromosomal NO can be shown to remain stable for at least 20 generations in culture. We detect no restriction pattern heterogeneity among X chromosomes isolated from a given strain, despite relatively large differences in their rDNA contents. This leads us to suggest that there is no significant clustering of intervening sequence-bearing (ivs ⁺) genes within the rDNA loci of chromosomes from the populations examined. Furthermore, we conclude that apparent alterations in rDNA redundancy known as the compensatory response are not related to the heterogeneity of rDNA content within a population.     

Distribution of spacer length classes and the intervening sequence among different nucleolus organizers in Drosophila hydei

Drosophila hydei rRNA genes from different chromosomes and from different stocks have been studied by restriction enzyme analysis. In DNA from wild-type females, about half of the X chromosomal rRNA genes are interrupted by an intervening sequence within the 28S coding region. In contrast to D. melanogaster, the intervening sequences belong to a single size class of 6.0 kb. Although there are two nucleolus organizers on the Y chromosome, genes containing the intervening sequence seem to be restricted to the X chromosome. — As shown in four cloned rDNA fragments, the nontranscribed spacers differ in length by having varying numbers of a 242 base pair sequence located in tandem in the right section of the spacer. In genomic rDNA, the spacers also differ in length by a regular 0.25 kb interval. Spacers with between 5 and 15 subrepeats occur frequently within the X and Y chromosomal nucleolus organizers in different D. hydei stocks; shorter and longer spacers are also present but are relatively rare. — Although each genotype is characterized by different frequencies of some spacer classes, the prominent spacer length heterogeneity pattern is similar among the different nucleolus organizers and, therefore, seems to be conserved during evolution.This paper is dedicated to Professor Dr. W. Beermann on the occasion of his 60th birthday     

Inviability of hybrids between D. melanogaster and D. simulans results from the absence of simulans X not the presence of simulans Y chromosome

Interspecific crosses between D. melanogaster and D. simulans or its sibling species result in unisexual inviability of the hybrids. Mostly, crosses of D. melanogaster females X D. simulans males produce hybrid females. On the other hand, only hybrid males are viable in the reciprocal crosses. A classical question is the cause of the unisexual hybrid inviability on the chromosomal level. Is it due to the absence of a D. simulans X chromosome or is it due to the presence of a D. simulans Y chromosome? A lack of adequate chromosomal rearrangements available in D. simulans has made it difficult to answer this question. However, it has been assumed that the lethality results from the absence of the D. simulans X rather than the presence of the D. simulans Y. Recently I synthesized the first D. simulans compound-XY chromosome that consists of almost the entire X and Y chromosomes. Males carrying the compound-XY and no free Y chromosome are fertile. By utilizing the compound-XY chromosome, the viability of hybrids with various constitutions of cytoplasm and sex chromosomes has been examined. The results consistently demonstrate that the absence of a D. simulans X chromosome in hybrid genome, and not the presence of the Y chromosome, is a determinant of the hybrid inviability.     

Factors Affecting Recognition and Disjunction of Chromosomes at Distributive Pairing in Female DROSOPHILA MELANOGASTER. I. Total Length VS. Arm Length

The behavior of a compound metacentric fourth chromosome (see PDF) has been examined to determine whether arm length or total length is the basis for recognition in distributive pairing. Recognition was judged by the frequency with which the (see PDF) nondisjoined from a series of X duplications (Dp), ranging in size from ≤ 0.3 to > 4 times the size of a single fourth chromosome. Dp, (see PDF) nondisjunction was measured in the absence and in the presence of a competitor, a compound metacentric X. In both situations, total length and not arm length, was found to confer the characteristic recognition property to the (see PDF). A comparison of Dp, (see PDF) nondisjunction curves for both the noncompetitive and competitive situations with analogous Dp, 4 curves previously obtained, show the Dp, (see PDF) curves to be similar in shape to those obtained earlier but displaced one unit to the right, corresponding precisely to the difference in size between the (see PDF) and the 4. Rules governing chromosome recognition for acrocentrics were found completely applicable to metacentrics; disjunctive behavior of metacentrics differed from that of acrocentrics in that two arms conferred on a chromosome the capacity to act as the intermediate of a trivalent when size no longer warranted this attribute. This capacity, itself, is size-dependent.     

Check of Gene Number during the Process of rDNA Magnification

THE multiple sequences of rDNA (DNA complementary to ribosomal RNA) of the Drosophila genome are localized at the bobbed locus, located in the X chromosome, position 66 and in the short arm of the Y chromosome^1,2. Wild bobbed (bb⁺) is that locus which, without a partner, gives rise to a normal phenotype. That locus which in similar conditions is incapable of giving rise to a normal phenotype is called a bobbed mutation (bb) and contains fewer genes for rRNA. The number of genes for rRNA in different individuals can vary considerably. One mechanism for rDNA variation is unequal crossing over³. Another mechanism, described by Tartof⁴, becomes apparent when individual flies, carrying only one bobbed locus, are constructed and only if such a locus is on the X chromosome; that is, if one constructs Xbb⁺/O males (and also Xbb/O males) or Xbb⁺/X_NO- females. Such individuals show a higher rDNA content than expected from the analysis of the same locus in Xbb⁺/Xbb⁺ females or in Xbb⁺/Ybb⁺ males. The increase of rDNA in this case is not inheritable⁴.     

Evolutionary breakpoint regions and chromosomal remodeling in Harttia (Siluriformes: Loricariidae) species diversification

The Neotropical armored catfish genus Harttia presents a wide variation of chromosomal rearrangements among its representatives. Studies indicate that translocation and Robertsonian rearrangements have triggered the karyotype evolution in the genus, including differentiation of sex chromosome systems. However, few studies used powerful tools, such as comparative whole chromosome painting, to clarify this highly diversified scenario. Here, we isolated probes from the X₁ (a 5S rDNA carrier) and the X₂ (a 45S rDNA carrier) chromosomes of Harttia punctata, which displays an X₁X₁X₂X₂/X₁X₂Y multiple sex chromosome system. Those probes were applied in other Harttia species to evidence homeologous chromosome blocks. The resulting data reinforce that translocation events played a role in the origin of the X₁X₂Y sex chromosome system in H. punctata. The repositioning of homologous chromosomal blocks carrying rDNA sites among ten Harttia species has also been demonstrated. Anchored to phylogenetic data it was possible to evidence some events of the karyotype diversification of the studied species and to prove an independent origin for the two types of multiple sex chromosomes, XX/XY₁Y₂ and X₁X₁X₂X₂/X₁X₂Y, that occur in Harttia species. The results point to evolutionary breakpoint regions in the genomes within or adjacent to rDNA sites that were widely reused in Harttia chromosome remodeling.     

Regulation of ribosomal RNA cistron number in a strain of Neurospora crassa with a duplication of the nucleolus organizer region

Some progeny from a cross of the translocation mutant T(VL→IVL)AR33 with wild-type Neurospora crassa are double nucleolus organizer (DNO) strains, usually displaying two distinct nucleolus organizer regions. The DNO strain is sterile but displays the same growth response as normal laboratory strains of Neurospora. We used DNA-DNA hybridization techniques to quantify the number of rRNA cistrons in the DNO mutant and its vegetative progeny. Comparisons of the rate of hybridization of genomic DNA from the parental AR33 strain and from the DNO strain showed that hybridization was more rapid for the DNO strain than for the parental strain. Successive vegetative progeny of the DNO strain displayed hybridization rates intermediate to those of the original DNO strain and the parental single nucleolus strain, indicating that the number of rRNA cistrons had decreased during vegetative propagation. Estimates of rRNA cistron number obtained from comparisons of the amount of single copy DNA and rDNA hybridized to genomic DNO and AR33 DNA at saturation indicate that the parental AR33 strain contains 225 copies of the rRNA repeat unit, while the DNO strain has approx. 440 copies. The number of rRNA cistrons decreases gradually in the successive vegetative progeny, approximating the parental haploid value by the eleventh vegetative transfer.     

Factors Influencing Disproportionate Replication of the Ribosomal RNA Cistrons in DROSOPHILA MELANOGASTER

This report describes studies of the compensatory response employing D. melanogaster stocks that bear cloned-X chromosomes derived from laboratory populations of strains Oregon R and Canton S. We find that modification of the autosomal background in either the female or the (see PDF) male parent influences the expression of the compensatory response by X chromosomes derived from the Canton S population, whereas Oregon R isolates are unresponsive to these effects. We have also studied compensatory replication in X/O larvae produced from cloned-X derivatives of both Canton S and Oregon R. Canton S larval compensation exceeds that of the adult, whereas in Oregon R the converse is true. We have concluded that both X chromosomal and autosomal factors affect the expression and magnitude of the compensatory response.     

Genetically Induced Mitotic Exchange in the Heterochromatin of DROSOPHILA MELANOGASTER

Multiple copies of the 18S and 28S ribosomal RNA cistrons are present in both the X and Y chromosomes of Drosophila melanogaster. Data are presented here that identify a locus, Rex, that causes exchange-like events between duplicated ribosomal complexes at the ends of an attached-XY chromosome. Rex: (1) is close to or in the basal heterochromatin of the X chromosome; (2) is semidominant and (its effect) is temperature sensitive; (3) acts maternally; and (4) affects behavior of paternally derived attached-XY chromosomes shortly after fertilization. Though, at this point, the existence of Rex is known only from its effects on behavior of a particular compound chromosome, it presents intriguing possibilities for understanding regulation of chromosome behavior and organization of the ribosomal cistrons.     

The Inbreeding Effective Population Number and the Expected Homozygosity for an X-Linked Locus

Assuming random mating and discrete nonoverlapping generations, the inbreeding effective population number, (see PDF), is calculated for an X-linked locus. For large populations, the result agrees with the variance effective population number. As an application, the maintenance of genetic variability by the joint action of mutation and random drift is investigated. It is shown that, if every allele mutates at rate u to new types, then the probabilities of identity in state (and hence the expected homozygosity of females) converge to the approximate value (see PDF) at the approximate asymptotic rate (see PDF).     

Indirect evidence of alteration in the expression of the rDNA genes in interspecific hybrids betweenDrosophila melanogaster andDrosophila simulans

Crosses betweenDrosophila melanogaster females andD. simulans males produce viable hybrid females, while males are lethal. These males are rescued if they carry theD. simulans Lhr gene. This paper reports that females of the wild-typeD. melanogaster population Staket do not produce viable hybrid males when crossed withD. simulans Lhr males, a phenomenon which we designate as the Staket phenotype. The agent responsible for this phenomenon was found to be the StaketX chromosome (X ^mel ,Stk). Analysis of the Staket phenotype showed that it is suppressed by extra copies ofD. melanogaster rDNA genes and that theX ^mel ,Stk chromosome manifests a weak bobbed phenotype inD. melanogaster X ^mel ,Stk/0 males. The numbers of functional rDNA genes inX ^mel ,Stk andX ^mel ,y w (control) chromosomes were found not to differ significantly. Thus a reduction in rDNA gene number cannot account for the weak bobbedX ^mel ,Stk phenotype let alone the Staket phenotype. The rRNA precursor molecules transcribed from theX ^mel ,Stk rDNA genes seem to be correctly processed in both intraspecific (melanogaster) and interspecific (melanogaster-simulans) conditions. It is therefore suggested that theX ^mel ,Stk rDNA genes are inefficiently transcribed in themelanogaster-simulans hybrids.     

The ribosomal DNA of Calliphora erythrocephala: The cistron classes of total genomic DNA

The composition of the genome set of ribosomal DNA cistrons in Calliphora erythrocephala (a Dipteran fly) has been analyzed. In contrast to previously cloned fragments of the rDNA (see Beckingham & White, 1980), the great majority of the rDNA cistrons do not contain introns in the 28 S β coding region. In the strain of flies studied, however, most cistrons fall into two discrete length classes that are present in approximately equal amounts in the genome. These results from distinct size variants of the non-transcribed spacer in the cistron population.The major genome class of intron-containing (intron⁺) rDNA cistrons was found to constitute approximately 5% of all cistrons and to contain introns of 6·1 × 10³ base-pairs. Interestingly, the intron⁺ cistrons were shown to be clustered within the rDNA and to contain a different population of non-transcribed spacer/external transcribed spacer (NTS + ETS) regions to that seen amongst the intron^? cistrons. The implications of these findings in relation to the mechanisms that maintain homogeneity within tandemly repeated gene sets are discussed.Some evidence for the existence of intron sequence DNA outside the rDNA is presented.     

Pleiotropic effects associated with the deletion of heterochromatin surrounding rDNA on the X chromosome of Drosophila

In Drosophila melanogaster X chromosome heterochromatin (Xh) constitutes the proximal 40% of the X chromosome DNA and contains a number of genetic elements with homologous sites on the Y chromosome, one of which is well defined, namely, the bobbed locus, the repetitive structural locus for the 18S and 28S rRNAs. This report presents the localisation of specific repeated DNA sequences within Xh and the employment of this sequence map in constructing new chromosomes to analyse the nature of the heterochromatin surrounding the rDNA region. Repeated sequences were located relative to inversion breakpoints which differentiate Xh cytogenetically. When the rDNA region was manipulated to be in a position in the chromosome so that it was without the Xh which normally surrounds it, the following obser-vations were made, (i) The rDNA region of Xh is intrinsically hetero-chromatic, remaining genetically active and yet possessing major heterochromatic properties even in the absence of the flanking heterochromatin regions, (ii) The size of the deletion removing the portion of Xh normally located distal to the rDNA region affected the dominance relationship between the X and Y nucleolar organizers (activity/endoreduplication assayed in male salivary glands). The X rDNA without any flanking heterochromatin was dominant over Y rDNA while the presence of some Xh allowed both the X and Y rDNA to be utilized, (iii) Enhancement of the position effect variegation on the white locus was demonstrated to occur as a result of the Xh deletions generated. EMS mutagenesis studies argue that the regions of Xh flanking the rDNA region contain no vital loci despite the fact that they strongly effect gene expression in some genotypes. This is consistent with early studies using X-ray mutagenesis (Lindsley et al., 1960). The pleiotropic effects of deleting specific regions of Xh is discussed in relation to the possible influence of heterochromatin on the organisation of the functional interphase nucleus.     

Sex Chromosome Translocations in the Evolution of Reproductive Isolation

Haldane's rule states that in organisms with differentiated sex chromosomes, hybrid sterility or inviability is generally expressed more frequently in the heterogametic sex. This observation has been variously explained as due to either genic or chromosomal imbalance. The fixation probabilities and mean times to fixation of sex-chromosome translocations of the type necessary to explain Haldane's rule on the basis of chromosomal imbalance have been estimated in small populations of Drosophila melanogaster. The fixation probability of an X chromosome carrying the long arm of the Y(X·Y^L) is approximately 30% greater than expected under the assumption of no selection. No fitness differences associated with the attached Y^L segment were detected. The fixation probability of a deficient Y chromosome is 300% greater than expected when the X chromosome contains the deleted portion of the Y. It is suggested that sex-chromosome translocations may play a role in the establishment of reproductive isolation.     

UNDER-REPLICATION OF RIBOSOMAL CISTRONS IN POLYTENE CHROMOSOMES OF RHYNCHOSCIARA

DNA preparations obtained from several tissues of Rhynchosciara americana and two related species, R. milleri and R. papaveroi, were hybridized to R. americana rRNA. The percentage of hybridization was found to be higher in tissues with low polyteny than in tissues with high polyteny, suggesting a relationship between the amount of rDNA and the tissue polyteny. This could be explained by under-replication of ribosomal cistrons in polytene cells, such as those from the salivary gland. Only slight tissue-dependent changes in the percentages of hybridization can be observed in heterologous hybridization using Xenopus laevis rRNA. The possibility that these experiments could not detect differences in the amount of ribosomal cistrons among tissues is discussed. The female:male ratio for the percentages of hybridization in the salivary gland of R. americana agrees with the results obtained by in situ hybridization experiments (16, 17) which have shown that the rRNA cistrons are distributed among chromosomes other than chromosome X.     

Differentiation and evolutionary relationships in Erythrinus erythrinus (Characiformes, Erythrinidae): comparative chromosome mapping of repetitive sequences

Erythrinus erythrinus presents extensive karyotypic diversity, with four karyomorphs (A–D) differing in the number of chromosomes, karyotype structure or sex chromosomes systems. Karyomorph A has 2n = 54 chromosomes in males and females without heteromorphic sex chromosomes, while karyomorph C has 2n = 52 chromosomes in females and 2n = 51 chromosomes in males, due a X₁X₁X₂X₂/X₁X₂Y sex chromosome system. Three allopatric populations of the karyomorph A and one population of the karyomorph C were now in deep investigated by molecular cytogenetic analyses, using repetitive DNAs as probes. The results reinforced the relatedness among populations of the karyomorph A, despite their large geographic distribution. Karyomorph C, however, showed a remarkably difference in the genomic constitution, especially concerning the amount and distribution of the 5S rDNA and Rex3 sequences on chromosomes. In addition, although karyomorphs C and D share several features, exclusive chromosomal markers show the derivative evolutionary pathway between them. Thus, besides the classical chromosomal rearrangements, the repetitive DNAs were useful tools to reveal the biodiversity, relatedness and differentiation of this fish group. The chromosomal set strongly corroborates that E. erythrinus corresponds to a species complex instead of a single biological entity.     

Structure,molecular evolution and maintenance of copy number of extended repeated structures in the X-heterochromatin of Drosophila melanogaster

The 60 kb repeats located in the distal heterochromatin of the X chromosome of Drosophila melanogaster were cloned in overlapping cosmids. These regions, designated as SCLRs, comprised the following types of repeated elements Stellate genes, which are known to be involved in spermatogenesis; copia-like retrotransposons; LINE elements, including amplified Type rDNA insertions; and rDNA fragments. The following steps in SCLR formation were hypothesized: insertion of mobile elements into the rDNA and Stellate gene clusters: internal tandem duplication events; recombination between the rDNA cluster and Stellate tandem repeat; and amplification of the whole SCLR structure. There are about nine SCLR copies per haploid genome, but there is approximately a twofold variation in copy number between fly stocks. The SCLR copy number differences between closely related stocks are suggested to be the result of unequal sister chromatid exchange (USCE). The restricted variation in SCLR copy number between unrelated stocks and the absence of chromosomes free of SCLRs suggests that natural selection is active in copy number maintenance.     

Polymorphisms in bovine immune genes and their associations with somatic cell count and milk production in dairy cattle

Background

Chromosome rearrangements are an important part of the speciation process in many taxa. The study of chromosome evolution in bivalves is hampered by the absence of clear chromosomal banding patterns and the similarity in both chromosome size and morphology. For this reason, obtaining good chromosome markers is essential for reliable karyotypic comparisons. To begin this task, the chromosomes of the mussels Brachidontes puniceus and B. rodriguezi were studied by means of fluorochrome staining and fluorescent in situ hybridization (FISH).

Results

Brachidontes puniceus and B. rodriguezi both have 2n = 32 chromosomes but differing karyotype composition. Vertebrate-type telomeric sequences appear at both ends of every single chromosome. B. puniceus presents a single terminal major rRNA gene cluster on a chromosome pair while B. rodriguezi shows two. Both mussels present two 5S rDNA and two core histone gene clusters intercalary located on the long arms of two chromosome pairs. Double and triple-FISH experiments demonstrated that one of the 5S rDNA and one of the major rDNA clusters appear on the same chromosome pair in B. rodriguezi but not in B. puniceus. On the other hand, the second 5S rDNA cluster is located in one of the chromosome pairs also bearing one of the core histone gene clusters in the two mussel species.

Conclusion

Knowledge of the chromosomal distribution of these sequences in the two species of Brachidontes is a first step in the understanding of the role of chromosome changes on bivalve evolution.     

Molecular differentiation of the homomorphic sex chromosomes inMegaselia scalaris (Diptera) detected by random DNA probes

Randomly cloned DNA fragments and a poly-(GATA) containing sequence were used as probes to identify sex chromosomal inheritance and to detect differences at the molecular level between the homomorphic X and Y in the phorid fly,Megaselia scalaris. Restriction fragment length differences between males and females and between two laboratory stocks of different geographic origin were used to differentiate between sex chromosomal and autosomal origin of the respective fragments. Five random probes detected X and Y chromosomal DNA loci and two others recognized autosomal DNA loci. One random probe and the poly(GATA) probe hybridized with both sex chromosomal and autosomal restriction fragments. Most of the Y chromosomal restriction fragments were conserved in length between the two stocks while most of the X chromosomal and autosomal fragments showed length polymorphism. It was concluded, therefore, that the Y chromosome contains a conserved segment in which crossover is suppressed and restriction site differences have accumulated relative to the X. These chromosomes, therefore, conform to a theoretically expected early stage of sex chromosome evolution.