Hybrid Dysgenesis in DROSOPHILA MELANOGASTER: The Genetics of Cytotype Determination in a Neutral Strain

The genetic determination of resistance to the sterility-producing genetic elements called P factors was studied in a strain characterized as neutral (Q) in the P-M system of hybrid dysgenesis. Sixteen lines were synthesized, representing all possible homozygous combinations of the three major chromosomes and differing maternal cytoplasms of an original resistant (Q) and susceptible (M) strain.—The results provide a detailed genetic analysis of the determination of cytotype (which mediates resistance or susceptibility to P factors) in the absence of the P-M dysgenic interaction. They extend the findings of Engels (1979) by providing specific information on both the location and relative magnitude of effect of cytotype-determining chromosomal factors and their interaction over time with maternally transmitted cytoplasm.—Cytotype was found to be primarily controlled by the genotype, but the maternal cytoplasm, under some circumstances, has an important short-term effect. Major cytotype-determining chromosomal factors are localized to the distal half of the X chromosome. However, there was also evidence for minor factors located on the major autosomes, particularly chromosome 3. Under certain circumstances, cytotypic switches in either direction can be produced in a single generation by the substitution of an X chromosome carrying a major cytotype determinant. This may provide an explanation of why reciprocal differences have sometimes been interpreted as direct effects of X-chromosome suppressors. However, slow but systematic changes of M to P cytotype were observed in five synthesized lines of mixed origin over twenty generations with no chromosomal substitution. Alternative explanations of these changes in terms of delayed effects of minor autosomal factors or of the transposability of cytotype determinants are discussed.     

Maternal inheritance of P cytotype in Drosophila melanogaster: a “pre-P cytotype” is strictly extra-chromosomally transmitted

In Drosophila melanogaster, transposition of the P element is under the control of a cellular state known as cytotype. The P cytotype represses P transposition whereas the M cytotype is permissive for transposition. In the long-term, the P cytotype is determined by chromosomal P elements but over a small number of generations it is maternally inherited. In order to analyse the nature of this maternal inheritance, we tested whether a maternal component can be transmitted without chromosomal P elements. We used a stable determinant of P cytotype, linked to the presence of two P elements at the tip of the X chromosome (IA site) in a genome devoid of other P elements. We measured P repression capacity using two different assays: gonadal dysgenic sterility (GD) and P-lacZ transgene repression. We show that zygotes derived from a P cytotype female (heterozygous for P (1A)/balancer devoid of P copies) and which inherit no chromosomal P elements from the mother, have, however, maternally received a P-type extra-chromosomal component: this component is insufficient to specify the P cytotype if the zygote formed does not carry chromosomal P elements but can promote P cytotype determination if regulatory P elements have been introduced paternally. We refer to this strictly extra-chromosomally inherited state as the “pre-P cytotype”. In addition, we show that a zygote that has the pre-P cytotype but which has not inherited any chromosomal P elements, does not transmit the pre-P cytotype to the following generation. The nature of the molecular determinants of the pre-P cytotype is discussed.     

Elements causing hybrid dysgenesis on the second chromosome ofDrosophila melanogaster

Summary Hybrid dysgenesis inDrosophila melanogaster is a syndrome of germline abnormalities including temperature-dependent gonadal dysgenesis (GD sterility), high rates of mutation and male recombination. In theP-M system, hybrid dysgenesis results from interaction between chromosomally-linked factors (P factors) and a particular extrachromosomal state refered to as theM cytotype. TheT007/Cy strain, shown by other authors to induce a high level of mutation and male recombination, is presently studied with respect to gonadal dysgenesis. TheP activity appears mainly linked with theT007 second chromosome and has been essentially mapped to a 0.6 centimorgan long interval, i.e. betweenhk andpr. On the other hand, 14 strains balanced for deficiencies on the left arm of the second chromosome are studied for their relative level ofM cytotype activity.In F1 females, inheriting the same maternal cytotype and the same paternalT007 chromosome, significant differences inGD sterility are found between flies receiving the maternal deficiency and those receiving the alternate non-deleted chromosome. This effect appears only when the chromosomes are deleted for a common region (37F5-38A7), suggesting the presence of elements intervening in the determinism ofGD sterility in this zone. As this region is included in the correspondinghk-pr interval (37C1-38B6), these results state the problem of the nature of the elements located in this interval and two hypotheses are discussed.     

Sterility and Hypermutability in the P-M System of Hybrid Dysgenesis in DROSOPHILA MELANOGASTER

Inbred wild strains of Drosophila melanogaster derived from the central and eastern United States were used to make dysgenic hybrids in the P-M system. These strains possessed P elements and the P cytotype, the condition that represses P element transposition. Their hybrids were studied for the mutability of the P element insertion mutation, snw, and for the incidence of gonadal dysgenesis (GD) sterility. All the strains tested were able to induce hybrid dysgenesis by one or both of these assays; however, high levels of dysgenesis were rare. Sets of X chromosomes and autosomes from the inbred wild strains were more effective at inducing GD sterility than were sets of Y chromosomes and autosomes. In two separate analyses, GD sterility was positively correlated with snw mutability, suggesting a linear relationship. However, one strain appeared to induce too much GD sterility for its level of snw destabilization, indicating an uncoupling of these two manifestations of hybrid dysgenesis.     

Components of Hybrid Dysgenesis in a Wild Population of DROSOPHILA MELANOGASTER

Hybrid dysgenesis is a condition found in certain interstrain hybrids of Drosophila melanogaster caused by the interaction of chromosomal and cytoplasmic factors. Germ-line abnormalities, including sterility, high mutability and male recombination, appear in the affected individuals. There are at least two distinct systems of hybrid dysgenesis. We examined a Wisconsin wild population in two consecutive years to determine the distribution of the chromosomal P factor and the extrachromosomal M cytotype that together cause one kind of hybrid dysgenic sterility. The P factor was found to be very common in the population, with all three major chromosomes being polymorphic for it. This polymorphism was strongly correlated with variability for male recombination elements, suggesting that these two traits are part of the same system of hybrid dysgenesis. There was a slight tendency for the P factor to be lost in lines taken from this population and inbred in the laboratory for many generations. A large-scale search for the M cytotype, which causes susceptibility to the P factor, showed that it is present in the population at only very low frequencies. Further evidence that the population is mostly immune to the action of the P factor was our finding of a general lack of dysgenic sterility in the wild flies themselves. However, we were able to isolate several wild strains that consistently showed the M cytotype. In some cases, the frequency of the M cytotype could be maintained in these lines, but it could not usually be increased by artificial selection. Some possible consequences of hybrid dysgenesis for the evolutionary biology of Drosophila are suggested.     

Gonadal Dysgenesis Determinants in a Natural Population of DROSOPHILA MELANOGASTER

Three populations of Drosophila melanogaster from northern California were surveyed for the ability to produce and resist gonadal dysgenesis in the P-M system of hybrid dysgenesis. Males from all three populations produced low to moderate levels of gonadal dysgenesis in crosses to Oregon-R M females. Most females had the P cytotype, but the M cytotype occurred occasionally. The three populations could not be statistically differentiated from one another, but were easily distinguished from populations from Australia and Wisconsin on the basis of gonadal dysgenesis potential. The California populations had higher levels of M cytotype than did the Wisconsin population. Thirteen X chromosomes and 11 pairs of autosomes were extracted from one of the California populations, using a modification of the standard balancer chromosome technique to suppress hybrid dysgenesis during extraction. All lines produced strongly skewed sterility distributions in crosses to M-strain females, and mean levels of sterility were less than 50%. There was evidence of nonadditive interactions between the autosomes. Most extraction lines had the P cytotype, but M and intermediate cytotypes were observed. Some of the intermediate cytotypes were stable over time. Lines were tested at two different times after extraction. Some lines evolved higher sterility potential as they were kept in the laboratory, even in the presence of P cytotype. The results point out a number of deficiencies in current genetic and population genetic models of hybrid dysgenesis and imply that gonadal dysgenesis is unlikely to be an important evolutionary force in this population.     

Cohabitation of KP and full-length P elements in the genome of MR strains inducing P-M-like hybrid dysgenesis in Drosophila melanogaster

Summary P strains of Drosophila melanogaster are characterized by the presence of both full-length and deletion derivatives of the transposable element P in their genome, and by their ability to induce the syndrome of hybrid dysgenesis among the progeny of certain intra-strain crosses, when introduced through the male parents. In contrast, strains belonging to the M' class, and which were also found to bear P element-homologous sequences, lack this ability and this has been attributed to the presence in the genome of most of these strains of a distinct class of deletion derivatives termed KP, which can suppress the action of functional P factors. Here we demonstrate that KP elements are present, next to full-length ones, in the genome of at least three strains which induce P-M-like dysgenic symptoms, including GD sterility. KP elements form the majority of the P-homologous sequences in the strains MR-h12, 23.5/CyL ⁴ and the latter's derivative 23.5 ^*/Cy. While the first one is a genuine P strain and the second one depicts a strong P cytotype, the third is a genuine M' strain. The hybrid dysgenesis induced by the two 23.5 MRF strains seems to be due, not primarily to the P elements, but to the action of hobo elements.     

EVOLUTIONARY STEPS AND TRANSPOSABLE ELEMENTS IN DROSOPHILA MELANOGASTER: THE MISSING RP TYPE OBTAINED BY GENETIC TRANSFORMATION

The I-R and P-M hybrid dysgenesis systems in Drosophila melanogaster have been interpreted as due to recent invasions of the genome by the I and P mobile genetic elements. Temporal and geographical surveys have never shown individuals harboring P sequences but devoid of active I elements. We describe here the successful genetic transformation by autonomous P elements of embryos initially devoid of active I elements and any P sequences. The results demonstrate that P elements may invade the genome of Drosophila melanogaster in the absence of active I elements. Using gel blotting, in situ hybridization techniques, and genetic experiments, we have monitored the behavior of newly introduced P elements in several transformed lines over 30 generations. The switch of cytotype from M to P occurred very slowly and the number of P copies simultaneously increased to about 25. These RP lines possess the properties required to induce P-M hybrid dysgenesis but totally retain the R cellular state. Consequently, this new mobile element combination presents a strong reciprocal post-zygotic isolation with IM strains due to both P-M and I-R hybrid dysgenesis systems. This genomic incompatibility could be considered as a first step toward speciation in Drosophila populations.     

An Analysis of Male-Recombination Elements in a Natural Population of DROSOPHILA MELANOGASTER in South Texas

Genomes from a group of Drosophila melanogaster collected from a natural population at San Benito, South Texas, in March of 1975 were analyzed for the presence of male-recombination elements. All three autosomes and both sex chromosomes were examined, with emphasis placed on the two major autosomes, the second and third chromosomes. In samples of 16 second and 16 third chromosomes, at least half, but not all, of each were found to carry male-recombination elements. It is suggested, although the data are not conclusive, that some of the fourth, X, and Y chromosomes might also be associated with male-recombination elements.—When a male-recombination element, or elements, was located in the second chromosome, relatively more male recombination was induced in the second than in the third chromosome. This situation was reversed when the element(s) was located in the third chromosome.—Distortion of transmission frequency, one of the characteristics of previously studied second chromosome lines associated with male recombination, was confirmed for these second chromosomes that carried male-recombination elements. Similar, but less pronounced, distortion was observed for the third chromosome lines that carried male-recombination elements.     

Distribution of heterochromatin on the mitotic chromosomes of Musca domestica L. in relation to the activity of male-determining factors

In the housefly, male sex is determined by a dominant factor, M, located either on the Y, on the X, or on any of the five autosomes. M factors on autosome I and on fragments of the Y chromosome show incomplete expressivity, whereas M factors on the other autosomes are fully expressive. To test whether these differences might be caused by heterochromatin-dependent position effects, we studied the distribution of heterochromatin on the mitotic chromosomes by C-banding and by fluorescence in situ hybridization of DNA fragments amplified from microdissected mitotic chromosomes. Our results show a correlation between the chromosomal position of M and the strength of its male-determining activity: weakly masculinizing M factors are exclusively located on chromosomes with extensive heterochromatic regions, i.e., on autosome I and on the Y chromosome. The Y is known to contain at least two copies of the M factor, which ensures a strong masculinizing effect despite the heterochromatic environment. The heterochromatic regions of the sex chromosomes consist of repetitive sequences that are unique to the X and the Y, whereas their euchromatic parts contain sequences that are ubiquitously found in the euchromatin of all chromosomes of the complement. Received: 20 February 1998; in revised form: 11 May 1998 / Accepted: 23 May 1998     

Hybrid Dysgenesis in Drosophila melanogaster: Evidence from Sterility and Southern Hybridization Tests That P Cytotype Is Not Maintained in the Absence of Chromosomal P Factors

A two-generation crossing program was used to replace the entire chromosome complement of P strains by M strain chromosomes, the maternal contribution being from the P strain. The cytotype of chromosomally substituted females was indistinguishable from M strain cytotype, judged by the sterility of offspring from the cross of such females to P strain males. In addition, following replacement of the chromosomes, the level of DNA homologous to the P factor was sufficiently low to be explicable by low levels of P factor transposition. These results are consistent with immediate chromosomal control for the switching from P to M cytotype. However, the reverse chromosome substitution, replacing all chromosomes of an M strain with P chromosomes, did not usually lead to immediate change of cytotype properties, showing that there is a true maternal effect in the M to P direction. The absence of true maternal inheritance for P cytotype argues against models of P factor repression which depend on autonomous replication of a nonchromosomal element. The repression could still be explained by nonchromosomal copies of the P factor, provided that these are replenished from chromosomal P factors. A model is put forward in which P cytotype is due to the presence of circular P factors carrying a P factor target sequence, leading to preferential transposition of chromosomal P factors to nonchromosomal target sites.     

An Excess of Gene Expression Divergence on the X Chromosome in Drosophila Embryos: Implications for the Faster-X Hypothesis

The X chromosome is present as a single copy in the heterogametic sex, and this hemizygosity is expected to drive unusual patterns of evolution on the X relative to the autosomes. For example, the hemizgosity of the X may lead to a lower chromosomal effective population size compared to the autosomes, suggesting that the X might be more strongly affected by genetic drift. However, the X may also experience stronger positive selection than the autosomes, because recessive beneficial mutations will be more visible to selection on the X where they will spend less time being masked by the dominant, less beneficial allele—a proposal known as the faster-X hypothesis. Thus, empirical studies demonstrating increased genetic divergence on the X chromosome could be indicative of either adaptive or non-adaptive evolution. We measured gene expression in Drosophila species and in D. melanogaster inbred strains for both embryos and adults. In the embryos we found that expression divergence is on average more than 20% higher for genes on the X chromosome relative to the autosomes; but in contrast, in the inbred strains, gene expression variation is significantly lower on the X chromosome. Furthermore, expression divergence of genes on Muller''s D element is significantly greater along the branch leading to the obscura sub-group, in which this element segregates as a neo-X chromosome. In the adults, divergence is greatest on the X chromosome for males, but not for females, yet in both sexes inbred strains harbour the lowest level of gene expression variation on the X chromosome. We consider different explanations for our results and conclude that they are most consistent within the framework of the faster-X hypothesis.     

Fundamentally different repetitive element composition of sex chromosomes in Rumex acetosa

Background and AimsDioecious species with well-established sex chromosomes are rare in the plant kingdom. Most sex chromosomes increase in size but no comprehensive analysis of the kind of sequences that drive this expansion has been presented. Here we analyse sex chromosome structure in common sorrel (Rumex acetosa), a dioecious plant with XY₁Y₂ sex determination, and we provide the first chromosome-specific repeatome analysis for a plant species possessing sex chromosomes.MethodsWe flow-sorted and separately sequenced sex chromosomes and autosomes in R. acetosa using the two-dimensional fluorescence in situ hybridization in suspension (FISHIS) method and Illumina sequencing. We identified and quantified individual repeats using RepeatExplorer, Tandem Repeat Finder and the Tandem Repeats Analysis Program. We employed fluorescence in situ hybridization (FISH) to analyse the chromosomal localization of satellites and transposons.Key ResultsWe identified a number of novel satellites, which have, in a fashion similar to previously known satellites, significantly expanded on the Y chromosome but not as much on the X or on autosomes. Additionally, the size increase of Y chromosomes is caused by non-long terminal repeat (LTR) and LTR retrotransposons, while only the latter contribute to the enlargement of the X chromosome. However, the X chromosome is populated by different LTR retrotransposon lineages than those on Y chromosomes.ConclusionsThe X and Y chromosomes have significantly diverged in terms of repeat composition. The lack of recombination probably contributed to the expansion of diverse satellites and microsatellites and faster fixation of newly inserted transposable elements (TEs) on the Y chromosomes. In addition, the X and Y chromosomes, despite similar total counts of TEs, differ significantly in the representation of individual TE lineages, which indicates that transposons proliferate preferentially in either the paternal or the maternal lineage.     

Cytotype regulation by telomeric P elements in Drosophila melanogaster: interactions with P elements from M' strains

P strains of Drosophila are distinguished from M strains by having P elements in their genomes and also by having the P cytotype, a maternally inherited condition that strongly represses P-element-induced hybrid dysgenesis. The P cytotype is associated with P elements inserted near the left telomere of the X chromosome. Repression by the telomeric P elements TP5 and TP6 is significantly enhanced when these elements are crossed into M′ strains, which, like P strains, carry P elements, but have little or no ability to repress dysgenesis. The telomeric and M′ P elements must coexist in females for this enhanced repression ability to develop. However, once established, it is transmitted maternally to the immediate offspring independently of the telomeric P elements themselves. Females that carry a telomeric P element but that do not carry M′ P elements may also transmit an ability to repress dysgenesis to their offspring independently of the telomeric P element. Cytotype regulation therefore involves a maternally transmissible product of telomeric P elements that can interact synergistically with products from paternally inherited M′ P elements. This synergism between TP and M′ P elements also appears to persist for at least one generation after the TP has been removed from the genotype.     

Hybrid Dysgenesis in DROSOPHILA MELANOGASTER: Nature and Inheritance of P Element Regulation

The genetic determination of the control of resistance or susceptibility to germ line changes mediated by P elements was studied in two strains and in derivatives of crosses between them. One strain, characterized as true M, completely lacked P elements. The second strain, pseudo-M (M'), carried a number of P elements, but these did not have the potential to induce the gonadal sterility that is associated with P-M hybrid dysgenesis. Individuals from the true M strain were invariably unable to suppress P factor activity (i.e., all daughters of outcrosses of M females and P males were sterile). In contrast, individuals from the M' strain showed variable degrees of suppression that were manifested in a wide range of gonadal sterility frequencies in standard tests. This continuous distribution pattern was reproducible for more than 25 generations.--The results of the genetic analysis indicate that a strain with a variable degree of suppression of gonadal dysgenesis is not necessarily in a transient state between the extreme conditions of P and M cytotype. A large variance in the ability to suppress gonadal dysgenesis with a mean value intermediate between the extremes of P and M cytotype may be a relatively stable strain characteristic. No reciprocal cross effect was observed in the suppression of sterility of F1 females from M X M' matings. Thus, the existence of M' strains indicates a Mendelian component in P element regulation and suggests that cytotype, which has an extrachromosomal aspect, may be only one of perhaps several mechanisms involved in regulation. Analysis of the effects of individual chromosomes from the M' strain showed that each chromosome contributed to the reduction of gonadal dysgenesis in the progeny of test matings. The results are consistent with a one-component titration model for P element regulation.     

High Mutability in Male Hybrids of DROSOPHILA MELANOGASTER

The frequencies of sex-linked lethal mutations arising in hybrid male offspring from various crosses and in nonhybrid controls were determined. The hybrids were produced by crossing representative strains of the P-M system of hybrid dysgenesis in all possible combinations. Males from the cross of P males x M females had a mutation rate about 15 times higher than that of nonhybrid males from the P strain. Genetically identical males from the reciprocal cross had a mutation rate 3 to 4 times that of the nonhybrids. For crosses involving a Q strain, a significant increase in the mutation rate was detected in males produced by matings of Q males with M females. No increase was observed in genetically identical males from the reciprocal mating. Crosses between P and Q strains gave male hybrids with mutation rates not different from those of nonhybrids. Many of the lethals that occurred in hybrids from the cross of P males x M females appeared to be unstable; fewer lethals that arose in hybrids from the cross of Q males x M females were unstable. The relationship between P and Q strains is discussed with respect to a model of mutation induction in dysgenic hybrids.     

Hybrid Sterility Locus on Chromosome X Controls Meiotic Recombination Rate in Mouse

Meiotic recombination safeguards proper segregation of homologous chromosomes into gametes, affects genetic variation within species, and contributes to meiotic chromosome recognition, pairing and synapsis. The Prdm9 gene has a dual role, it controls meiotic recombination by determining the genomic position of crossover hotspots and, in infertile hybrids of house mouse subspecies Mus m. musculus (Mmm) and Mus m. domesticus (Mmd), it further functions as the major hybrid sterility gene. In the latter role Prdm9 interacts with the hybrid sterility X 2 (Hstx2) genomic locus on Chromosome X (Chr X) by a still unknown mechanism. Here we investigated the meiotic recombination rate at the genome-wide level and its possible relation to hybrid sterility. Using immunofluorescence microscopy we quantified the foci of MLH1 DNA mismatch repair protein, the cytological counterparts of reciprocal crossovers, in a panel of inter-subspecific chromosome substitution strains. Two autosomes, Chr 7 and Chr 11, significantly modified the meiotic recombination rate, yet the strongest modifier, designated meiotic recombination 1, Meir1, emerged in the 4.7 Mb Hstx2 genomic locus on Chr X. The male-limited transgressive effect of Meir1 on recombination rate parallels the male-limited transgressive role of Hstx2 in hybrid male sterility. Thus, both genetic factors, the Prdm9 gene and the Hstx2/Meir1 genomic locus, indicate a link between meiotic recombination and hybrid sterility. A strong female-specific modifier of meiotic recombination rate with the effect opposite to Meir1 was localized on Chr X, distally to Meir1. Mapping Meir1 to a narrow candidate interval on Chr X is an important first step towards positional cloning of the respective gene(s) responsible for variation in the global recombination rate between closely related mouse subspecies.     

The Evolutionary History of DROSOPHILA BUZZATII. Xii. the Genetic Basis of Sterility in Hybrids between D. BUZZATII and Its Sibling D. SERIDO from Argentina

The genetic basis of hybrid sterility has been investigated in backcross segmental hybrids between two sibling species, Drosophila buzzatii and D. serido. Asynapsis of homologous bands in hybrid polytene chromosomes has been used to identify the D. serido chromosome segments introgressed into the D. buzzatti genome. All the investigated chromosomes contain male sterility factors. For autosomes, sterility is produced when an introgressed D. serido chromosome segment, or combination of segments, reaches a minimum size. On the other hand, any introgressed X chromosome segment from D. serido, irrespective of its size, produces either male hybrid sterility or inviability.     

DNA REPLICATION PATTERNS AND CHROMOSOMAL PROTEIN SYNTHESIS IN OPOSSUM LYMPHOCYTES IN VITRO

DNA replication patterns were determined in the autosomes and sex chromosomes of phytohemagglutinin-stimulated lymphocytes from the opossum (Didelphis virginiana) by employing thymidine-³H labeling and high-resolution radioautography. Opossum chromosomes are desirable experimental material due to their large size, low number (2n = 22), and morphologically distinct sex chromosomes. The autosomes in both sexes began DNA synthesis synchronously and terminated replication asynchronously. One female X chromosome synthesized DNA throughout most of the S phase. Its homologue, however, began replication approximately 3.5 hr later. The two X's terminated DNA synthesis synchronously, slightly later than the autosomes. This form of late replication, in which one X chromosome begins DNA synthesis later than its homologue but completes replication at the same time as its homologue, is apparently unique in the opossum. The male X synthesized DNA throughout S while the Y chromosome exhibited late-replicating characteristics. The two sex chromosomes completed synthesis synchronously, slightly later than the autosomes. Grain counts were performed on all chromosomes to analyze trends in labeling intensity at hourly intervals of S. By analyzing the percent of labeled mitotic figures on radioautographs at various intervals after introduction of arginine-³H, chromosomal protein synthesis was found not to be restricted to any portion of interphase but to increase throughout S and into G₂.