Rapid Evolution of a Coadapted Gene Complex: Evidence from the Segregation Distorter (Sd) System of Meiotic Drive in Drosophila Melanogaster

Segregation Distorter (SD) is a system of meiotic drive found in natural populations of Drosophila melanogaster. Males heterozygous for an SD second chromosome and a normal homologue (SD(+)) produce predominantly SD-bearing sperm. The coadapted gene complex responsible for this transmission advantage spans the second chromosome centromere, consisting of three major and several minor interacting loci. To investigate the evolutionary history of this system, we surveyed levels of polymorphism and divergence at six genes that together encompass this pericentromeric region and span seven map units. Interestingly, there was no discernible divergence between SD and SD(+) chromosomes for any of these molecular markers. Furthermore, SD chromosomes harbored much less polymorphism than did SD(+) chromosomes. The results suggest that the SD system evolved recently, swept to appreciable frequencies worldwide, and carried with it the entire second chromosome centromeric region (roughly 10% of the genome). Despite its well-documented genetic complexity, this coadapted system appears to have evolved on a time scale that is much shorter than can be gauged using nucleotide substitution data. Finally, the large genomic region hitchhiking with SD indicates that a multilocus, epistatically selected system could affect the levels of DNA polymorphism observed in regions of reduced recombination.     

Transposition of the Responder Element (Rsp) of the Segregation Distorter System (Sd) to the X Chromosome in Drosophila Melanogaster

In order to test whether the meiotic drive system Segregation distorter (SD) can operate on the X chromosome to exclude it from functional sperm, we have transposed the Responder locus (Rsp) to this element. This was accomplished by inducing detachments of a compound-X chromosome in females carrying a Y chromosome bearing a Rsps allele. Six Responder-sensitive-bearing X chromosomes, with kappa values ranging from 0.90 to 1.00, were established as permanent lines. Two of these have been characterized more extensively with respect to various parameters affecting meiotic drive. SD males with a Responder-sensitive X chromosome produce almost exclusively male embryos, while those with a Rsp-Y chromosome produce almost exclusively female embryos. This provides a genetic system of great potential utility for the study of early sex-specific differentiation events as it allows the collection of large numbers of embryos of a given sex.     

X-Linked Elements Associated with Negative Segregation Distortion in the Sd System of Drosophila Melanogaster

Three elements, M(1), M(2) and M(3), found in a special X chromosome, supp-X(SD), modify the degree and direction of segregation distortion in the SD system of Drosophila melanogaster. The first element, M(1), is located between the y and the cv loci, probably close to the y locus. The second element, M(2), is located near the cv locus and the third element, M(3), is located between the y and the car loci. The M(1) element appears to cause a relatively small amount of reduction in the rate of recovery of the SD-72, but not the cn bw, chromosome from SD-72/ cn bw males, when raised at 27.5°. The M(2) and the M(3) elements cause considerable decrease in the recovery rate of the SD-72 chromosome, whereas they increase the recovery rate of the cn bw chromosome. The amount of decrease is nearly the same as the amount of increase for each element. Some type of ``switch' mechanism in the directions of distortion is suggested for each of these two elements and their effects appear to be approximately additive.     

A Possible Case of Negative Segregation Distortion in the Sd System of Drosophila Melanogaster

Models proposed to explain segregation distortion in Drosophila melanogaster are based partly upon the observation that, in the Sd heterozygous Rspi/Rsps male, the chromosome carrying the sensitive Rsps allele is distorted, but the chromosome carrying the insensitive Rspi allele is not. The results of the present study suggest that this may not always be the case. Under a certain genotypic condition, the chromosome carrying the Rsps allele can be transmitted to the progeny in frequencies of more than 0.5 (about 0.6), or correspondingly, the chromosome carrying the Rspi allele may be distorted with respect to the one with the Rsps allele. Thus, the relative sensitivity and insensitivity of the two Rsp alleles in a male are not absolute, but relative; and they may be reversed depending upon the residual genotype. If this is true, a major modification of the current models or a proposal of an entirely new model may become necessary to explain the mechanism of segregation distortion satisfactorily.     

Examination of a Mutable System Affecting the Components of the Segregation Distorter Meiotic Drive System of Drosophila Melanogaster

Segregation distortion in Drosophila melanogaster is the result of an interaction between the genetic elements Sd, a Rsp sensitive to Sd, and an array of modifiers, that results in the death of sperm carrying Rsp. A stock (designated M-5; cn bw) has been constructed which has the property of inducing the partial loss of sensitivity from previously sensitive cn bw chromosomes, the partial loss of distorting ability from SD chromosomes, and a concomitant acquisition of modifiers on the X chromosome and possibly also on the autosomes. By several criteria the changes exhibited under the influence of M-5; cn bw are characteristic of the transposable-element systems which produce hybrid dysgenesis. In the first place, the magnitude of these effects depends on the nature of the crosses performed. The analogy is further strengthened by the observation that the changes induced by M-5; cn bw share other stigmata of Drosophila transposable-element systems, including high sterility among the progeny of outcrosses, and the production of chromosomal rearrangements. The possible relationship of this system to the P, I and hobo transposable element systems is discussed, as well as its bearing on aspects of the Segregation Distorter phenomenon which have yet to be explained.     

The Effect of Novel Chromosome Position and Variable Dose on the Genetic Behavior of the Responder (Rsp) Element of the Segregation Distorter (Sd) System of Drosophila Melanogaster

In the Segregation distorter (SD) system of meiotic drive, a minimum of two trans-acting elements [Sd and E(SD)] act in concert to cause a certain probability of dysfunction for sperm carrying a sensitive allele at the Responder (Rsp) target locus. By employing a number of insertional translocations of autosomal material into the long arm of the Y chromosome, Rsp can be mapped as the most proximal locus in the 2R heterochromatin as defined both by cytology and lethal complementation tests. Several of these insertional translocations result in the transposition of Rsp to the Y chromosome, where its sensitivity remains virtually unaltered. This argues that Rsp is separable from the second chromosome centromere, that its behavior does not depend on its gross chromosomal position, and that meiotic pairing of the chromosomes carrying the various SD elements is not a prerequisite for sperm dysfunction. Several other translocations apparently leave both resulting chromosomes at least partially sensitive to SD action, suggesting that Rsp is a large subdivisible genetic element. This view is compatible with observations published elsewhere that suggest that Rsp is a cytologically large region of highly repetitive AT-rich DNA. The availability of Y-linked copies of Rsp also allows the construction of SD males carrying two independently segregating Rsp alleles; this in turn allows the production of sperm with zero, one or two Rsp copies from the same male. Examination of the relative recovery proportions of progeny arising from these gametes suggests that sperm with two Rsp copies survive at much lower frequencies than would be predicted if each Rsp acted independently in causing sperm dysfunction. Possible explanations for such behavior are discussed.     

Negative Segregation Distortion in the Sd System of Drosophila Melanogaster: A Challenge to the Concept of Differential Sensitivity of Rsp Alleles

Current models of segregation distortion based on previous experimental results predict that, in the Sd heterozygous Rspi/Rsps male, the chromosome carrying the sensitive Rsps allele is distorted or transmitted in a frequency smaller than that of the expected Mendelian 0.5 relative to the chromosome carrying the insensitive Rspi allele. The present study presents a case where this does not occur, that is, when the genotype of the males is supp-X(SD)/Y; Sd E(SD)Rspi M(SD)+/Sd+ E(SD)+ Rsps M(SD)+ where supp-X(SD) is an X chromosome carrying a strong suppressor or suppressors of SD activity and SD+ E(SD)+ Rsps M(SD)+ is the standard cn bw chromosome. Following the "inseminated female transfer" procedure, young males of the above genotype carrying the standard-X instead of the supp-X(SD) chromosome show k values for the SD chromosome (frequencies of the SD chromosome recovered among progeny) of about 0.75, but with the supp-X(SD) chromosome, the k values are reduced to 0.36-0.41. Several possibilities other than the mechanism of segregation distortion to explain the reduced k values are ruled out. The occurrence of "negative segregation distortion" is clearly demonstrated, where the chromosome carrying the Rspi allele is distorted but the chromosome with the Rsps allele is not. This result requires a major modification of the current models or even a new model for the mechanism of segregation distortion to accommodate Rsp allele sensitivity or insensitivity. The present study also shows that males of the genotype, Sd Rspss M(SD)+/Sd+ Rspss M(SD), are almost completely sterile, but their fertility is considerably increased when SD activity is suppressed by the presence of the supp-X(SD) chromosome. This result suggests that the amount of the Sd product is not limited with respect to the interacting sites available, that is, the amount is large enough to interact with both of the Rspss alleles.     

Segregation Distortion in Drosophila Melanogaster: Genomic Organization of Responder Sequences

The heterochromatic Responder (Rsp) locus of Drosophila melanogaster is the target of the two distorter loci Sd and E(SD). Rsp is located in a specific heterochromatic region of the second chromosome and is made up of AT-rich satellite sequences whose abundance is related to its sensitivity to the distorter chromosomes. Here we report that a cluster of Rsp sequences is also located in the third chromosome. The third-chromosome cluster has the same flanking sequences as the clone originally used to identify the Rsp elements, and one of the flanking sequences is a rearranged 412 retrotrsansposon. The presence of a second, unlinked Rsp-sequence cluster makes re-interpretation necessary for some earlier experiments in which segregation of the third chromosome had not been followed and raises interesing possibilities for the origin of the Rsp locus.     

Stability of Tandem Repeats in the Drosophila Melanogaster HSR-Omega Nuclear RNA

The Drosophila melanogaster Hsr-omega locus produces a nuclear RNA containing >5 kb of tandem repeat sequences. These repeats are unique to Hsr-omega and show concerted evolution similar to that seen with classical satellite DNAs. In D. melanogaster the monomer is ~280 bp. Sequences of 191/2 monomers differ by 8 +/- 5% (mean +/- SD), when all pairwise comparisons are considered. Differences are single nucleotide substitutions and 1-3 nucleotide deletions/insertions. Changes appear to be randomly distributed over the repeat unit. Outer repeats do not show the decrease in monomer homogeneity that might be expected if homogeneity is maintained by recombination. However, just outside the last complete repeat at each end, there are a few fragments of sequence similar to the monomer. The sequences in these flanking regions are not those predicted for sequences decaying in the absence of recombination. Instead, the fragmentation of the sequence homology suggests that flanking regions have undergone more severe disruptions, possibly during an insertion or amplification event. Hsr-omega alleles differing in the number of repeats are detected and appear to be stable over a few thousand generations; however, both increases and decreases in repeat numbers have been observed. The new alleles appear to be as stable as their predecessors. No alleles of less than ~5 kb nor more than ~16 kb of repeats were seen in any stocks examined. The evidence that there is a limit on the minimum number of repeats is consistent with the suggestion that these repeats are important in the function of the unusual Hsr-omega nuclear RNA.     

Further Characterization of Genetic Elements Associated with the Segregation Distorter Phenomenon in DROSOPHILA MELANOGASTER

Segregation Distorter, SD, associated with the second chromosome of Drosophila melanogaster, is known to cause sperm bearing the non-SD homologue to dysfunction in heterozygous males. In earlier studies, using different, independently derived, SD chromosomes, three major loci were identified as contributing to the distortion of segregation ratios in males. In this study the genetic components of the SD-5 chromosome have been the subjects of further investigation, and our findings offer the following information. Crossover analysis confirms the mapping of the Sd locus to a position distal to but closely linked with the genetic marker pr. Spontaneous and radiation-induced recombinational analyses and deficiency studies provide firm support to the notion that the Rsp (Responder) locus lies within the proximal heterochromatin of chromosome 2, between the genetic markers lt and rl and most likely in the heterochromatin of the right arm. The major focus of this study, however, has been on providing a better definition of the genetic properties of the Enhancer of SD [E(SD)]. Our findings place this locus within the region of the two most proximal essential genes in the heterochromatin of the left arm of chromosome 2. Moreover, our analysis reveals a probable association of the E(SD) locus with a meiotic drive independent of that caused by Sd.     

Partial Revertants of the Transposable Element-Associated Suppressible Allele White-Apricot in Drosophila Melanogaster: Structures and Responsiveness to Genetic Modifiers

The eye color phenotype of white-apricot (wa), a mutant allele of the white locus caused by the insertion of the transposable element copia into a small intron, is suppressed by the extragenic suppressor suppressor-of-white-apricot (su(wa] and enhanced by the extragenic enhancers suppressor-of-forked su(f] and Enhancer-of-white-apricot (E(wa]. Derivatives of wa have been analyzed molecularly and genetically in order to correlate the structure of these derivatives with their response to modifiers. Derivatives in which the copia element is replaced precisely by a solo long terminal repeat (sLTR) were generated in vitro and returned to the germline by P-element mediated transformation; flies carrying this allele within a P transposon show a nearly wild-type phenotype and no response to either su(f) or su(wa). In addition, eleven partial phenotypic revertants of wa were analyzed. Of these, one appears to be a duplication of a large region which includes wa, three are new alleles of su(wa), two are sLTR derivatives whose properties confirm results obtained using transformation, and five are secondary insertions into the copia element within wa. One of these, waR84h, differs from wa by the insertion of the most 3' 83 nucleotides of the I factor. The five insertion derivatives show a variety of phenotypes and modes of interaction with su[f) and su(wa). The eye pigmentation of waR84h is affected by su(f) and E(wa), but not su(wa). These results demonstrate that copia (as opposed to the interruption of white sequences) is essential for the wa phenotype and its response to genetic modifiers, and that there are multiple mechanisms for the alteration of the wa phenotype by modifiers.     

The Genetic Analysis of Achiasmate Segregation in Drosophila Melanogaster. III. the Wild-Type Product of the Axs Gene Is Required for the Meiotic Segregation of Achiasmate Homologs

The regular segregation of achiasmate chromosomes in Drosophila melanogaster females is ensured by two distinct segregational systems. The segregation of achiasmate homologs is assured by the maintenance of heterochromatic pairing; while the segregation of heterologous chromosomes is ensured by a separate mechanism that may not require physical association. Axs(D) (Aberrant X segregation) is a dominant mutation that specifically impairs the segregation of achiasmate homologs; heterologous achiasmate segregations are not affected. As a result, achiasmate homologs frequently participate in heterologous segregations at meiosis I. We report the isolation of two intragenic revertants of the Axs(D) mutation (Axs(r2) and Axs(r3)) that exhibit a recessive meiotic phenotype identical to that observed in Axs(D)/Axs(D) females. A third revertant (Axs(r1)) exhibits no meiotic phenotype as a homozygote, but a meiotic defect is observed in Axs(r1)/Axs(r2) females. Therefore mutations at the Axs(D) locus define a gene necessary and specific for homologous achiasmate segregation during meiosis. We also characterize the interactions of mutations at the Axs locus with two other meiotic mutations (ald and ncd). Finally, we propose a model in which Axs(+) is required for the normal separation of paired achiasmate homologs. In the absence of Axs(+) function, the homologs are often unable to separate from each other and behave as a single segregational unit that is free to segregate from heterologous chromosomes.     

Mutations to the piRNA Pathway Component Aubergine Enhance Meiotic Drive of Segregation Distorter in Drosophila melanogaster

Diploid sexual reproduction involves segregation of allelic pairs, ensuring equal representation of genotypes in the gamete pool. Some genes, however, are able to “cheat” the system by promoting their own transmission. The Segregation distorter (Sd) locus in Drosophila melanogaster males is one of the best-studied examples of this type of phenomenon. In this system the presence of Sd on one copy of chromosome 2 results in dysfunction of the non–Sd-bearing (Sd⁺) sperm and almost exclusive transmission of Sd to the next generation. The mechanism by which Sd wreaks such selective havoc has remained elusive. However, its effect requires a target locus on chromosome 2 known as Responder (Rsp). The Rsp locus comprises repeated copies of a satellite DNA sequence and Rsp copy number correlates with sensitivity to Sd. Under distorting conditions during spermatogenesis, nuclei with chromosomes containing greater than several hundred Rsp repeats fail to condense chromatin and are eliminated. Recently, Rsp sequences were found as small RNAs in association with Argonaute family proteins Aubergine (Aub) and Argonaute3 (AGO3). These proteins are involved in a germline-specific RNAi mechanism known as the Piwi-interacting RNA (piRNA) pathway, which specifically suppresses transposon activation in the germline. Here, we evaluate the role of piRNAs in segregation distortion by testing the effects of mutations to piRNA pathway components on distortion. Further, we specifically targeted mutations to the aub locus of a Segregation Distorter (SD) chromosome, using ends-out homologous recombination. The data herein demonstrate that mutations to piRNA pathway components act as enhancers of SD.     

Characterization of the Mus308 Gene in Drosophila Melanogaster

Among the available mutagen-sensitive mutations in Drosophila, those at the mus308 locus are unique in conferring hypersensitivity to DNA cross-linking agents but not to monofunctional agents. Those mutations are also associated with an elevated frequency of chromosomal aberrations, altered DNA metabolism and the modification of a deoxyribonuclease. This spectrum of phenotypes is shared with selected mammalian mutations including Fanconi anemia in humans. In anticipation of the molecular characterization of the mus308 gene, it has been localized cytogenetically to 87C9-87D1,2 on the right arm of chromosome three. Nine new mutant alleles of the gene have been generated by X-ray mutagenesis and one was recovered following hybrid dysgenesis. Characterization of these new alleles has uncovered additional phenotypes of mutations at this locus. Homozygous mus308 flies that have survived moderate mutagen treatment exhibit an altered wing position that is correlated with reduced flight ability and an altered mitochondrial morphology. In addition, observations of elevated embryo mortality are potentially explained by an aberrant distribution of nuclear material in early embryos which is similar to that seen in the mutant giant nuclei.     

Genetic Evidence That the Sans Fille Locus Is Involved in Drosophila Sex Determination

Females homozygous for sans fille1621 (= fs(1)1621) have an abnormal germ line. Instead of producing eggs, the germ-line cells proliferate forming ovarian tumors or excessive numbers of nurse cells. The Sex-lethal gene product(s) regulate the branch point of the dosage compensation and sex determination pathways in the soma. The role of Sex-lethal in the germ line is not clear but the germ line of females homozygous for female sterile Sex-lethal alleles or germ-line clones of loss-of-function alleles are characterized by ovarian tumors. Females heterozygous for sans fille1621 or Sex-lethal are phenotypically wild type with respect to viability and fertility but females trans-heterozygous for sans fille1621 and Sex-lethal show ovarian tumors, somatic sexual transformations, and greatly reduced viability.