Kyoichi Sawamura

CHARACTERIZATION OF A REPRODUCTIVE ISOLATION GENE, ZYGOTIC HYBRID RESCUE, OF DROSOPHILA MELANOGASTER BY USING MINICHROMOSOMES

Hybrids carrying the wild type allele of zygotic hybrid rescue (zhr) of Drosophila melanogaster and the maternal cytoplasm of its sibling species (D. simulans, D. mauritiana or D. sechellia) are embryonic lethal, irrespective of their sex. The zhr gene has been localized in the X heterochromatin, slightly distal to the In(1)sc8 breakpoint, a location where highly repetitive satellite DNA exists. A free duplication minichromosome, Dp(1;f)1162, which is derived from the In(1)sc8 chromosome by a massive interstitial deletion has a significant, but not complete, effect on the hybrid lethality. In order to examine the organization of zhr cytogenetically, we synthesized four deletions from Dp(1;f)1162 by γ-ray irradiation. Three of them, Dp(1;f)1162S2, Dp(1;f)1162S3, and Dp(l;f)1162S5, showed a hybrid lethal effect not significantly different from Dp(1;f)1162, but one, Dp(1;f)1162S4, showed a significantly weaker hybrid lethal effect. Interestingly, hybrids were completely lethal when two copies of Dp(1;f)1162S4 were present. That the lethal effect is proportional to the number of the minichromosomes supports the model that a class of repeated sequences around the Dp(1;f)1162S4 breakpoint may be involved in the zhr gene activity. In the future, the mini-chromosome could be useful in the cloning and characterization of this reproductive isolation gene.

Biology of reproductive isolation in Drosophila: toward a better understanding of speciation

Abstract Mechanisms of premating and postmating isolation between Drosophila melanogaster and its sibling species, D. simulans, D. mauritiana, and D. sechellia, are reviewed here. As D. melanogaster has been used as a “model organism” in a variety of studies, we can apply much knowledge about this species to interspecific hybridizations. Mating success between species is low because of species specificity of pheromonal and acoustic signals and the receivers, although other factors are also involved. On the other hand, genetic incompatibility between species results in inviability and sterility in F1 and backcross hybrids. Studies of reproductive isolation between these Drosophila species will shed light on animal speciation and the origin of biodiversity.

Introgression of Drosophila Simulans Nuclear Pore Protein 160 in Drosophila Melanogaster Alone Does Not Cause Inviability but Does Cause Female Sterility

We have been analyzing genes for reproductive isolation by replacing Drosophila melanogaster genes with homologs from Drosophila simulans by interspecific backcrossing. Among the introgressions established, we found that a segment of the left arm of chromosome 2, Int(2L)S, carried recessive genes for hybrid sterility and inviability. That nuclear pore protein 160 (Nup160) in the introgression region is involved in hybrid inviability, as suggested by others, was confirmed by the present analysis. Male hybrids carrying an X chromosome of D. melanogaster were not rescued by the Lethal hybrid rescue (Lhr) mutation when the D. simulans Nup160 allele was made homozygous or hemizygous. Furthermore, we uniquely found that Nup160 is also responsible for hybrid sterility. Females were sterile when D. simulans Nup160 was made homozygous or hemizygous in the D. melanogaster genetic background. Genetic analyses indicated that the D. simulans Nup160 introgression into D. melanogaster was sufficient to cause female sterility but that other autosomal genes of D. simulans were also necessary to cause lethality. The involvement of Nup160 in hybrid inviability and female sterility was confirmed by transgene experiment.

Genetics of hybrid inviability and sterility in Drosophila: dissection of introgression of D. simulans genes in D. melanogaster genome

Interspecific crosses between Drosophila melanogaster and Drosophila simulans usually produce sterile unisexual hybrids. The barrier preventing genetic analysis of hybrid inviability and sterility has been taken away by the discovery of a D. simulans strain which produces fertile female hybrids. D. simulans genes in the cytological locations of 21A1 to 22C1-23B1 and 30F3-31C5 to 36A2-7 have been introgressed into the D. melanogaster genetic background by consecutive backcrosses. Flies heterozygous for the introgression are fertile, while homozygotes are sterile both in females and males. The genes responsible for the sterility have been mapped in the introgression. The male sterility is caused by the synergistic effect of multiple genes, while the female sterility genes have been localized to a 170 kb region (32D2 to 32E4) containing 20 open reading frames. Thus, the female sterility might be attributed to a single gene with a large effect. We have also found that the Lethal hybrid rescue mutation which prevents the inviability of male hybrids from the cross of D. melanogaster females and D. simulans males cannot rescue those carrying the introgression, suggesting that D. simulans genes maybe non-functional in this hybrid genotype. The genes responsible for the inviability have not been separated from the female sterility genes by recombination.

Chromatin Evolution and Molecular Drive in Speciation

Are there biological generalities that underlie hybrid sterility or inviability? Recently, around a dozen “speciation genes” have been identified mainly in Drosophila, and the biological functions of these genes are revealing molecular generalities. Major cases of hybrid sterility and inviability seem to result from chromatin evolution and molecular drive in speciation. Repetitive satellite DNAs within heterochromatin, especially at centromeres, evolve rapidly through molecular drive mechanisms (both meiotic and centromeric). Chromatin-binding proteins, therefore, must also evolve rapidly to maintain binding capability. As a result, chromatin binding proteins may not be able to interact with chromosomes from another species in a hybrid, causing hybrid sterility and inviability.

Potential gene flow in natural populations of the Drosophila ananassae species cluster inferred from a nuclear mitochondrial pseudogene

A pseudogene with 94% similarity to mitochondrial cytochrome c oxidase subunit I (COI) was identified and localized to chromosome 4 of Drosophila ananassae. Because this chromosome is believed to have reduced recombination, its history can be traced using the pseudo-COI sequence. Pseudo-COI sequences were obtained from 27 iso-female lines of six taxa belonging to the D. ananassae species cluster in which reproductive isolation is incomplete. The phylogenetic network constructed from seven recognized haplotypes (#0-#6) indicated that different taxa inhabiting the same geographic area share the haplotypes: #1 from Papua New Guinean populations of D. ananassae and pallidosa-like-Wau; #2 from Papua New Guinean populations of D. ananassae, pallidosa-like, and papuensis-like; and #4 from South Pacific populations of D. ananassae and D. pallidosa. Taxon-K has a unique haplotype (#6), and 18 mutation steps separate it from the closest haplotype, #2. We discuss the possibility of chromosome 4 introgression beyond taxon boundaries.

The Origin of Reproductive Isolation: Biological Mechanisms of Genetic Incompatibility

The origin of species has been the mystery of mysteries since Charles Darwin’s era. According to the biological species concept advocated by the founders of the Modern Synthesis, the question ‘how new species evolve’ can be substituted by a more answerable question ‘how reproductive isolation is established between populations’. Mechanisms preventing gene exchange between species are various, and most of them are thought to be the result, not the cause, of genetic diversity. Some disturb species recognition systems (prezygotic isolation), and others terminate generations of mixed genomes (postzygotic isolation). Examples of the latter whose biological mechanisms have recently been documented well (tumorigenesis, sexual reversion, irregular dosage compensation, nucleolar dominance, homeotic transformation, spermatogenic defects, mitotic defects, and maternal/zygotic transition failure in interspecific hybrids) are reviewed here. Gene interactions between loci from different species play important roles. As has been shown in developmental biology, transcriptional regulation may be a key factor. Genetic incompatibility in hybrids may be the manifestation of improper transcriptional regulation resulting from the coevolution of DNA-binding proteins and their binding sites.

Copulatory Courtship Behavior and Sine Song as a Mate Recognition Cue in Drosophila lini and Its Sibling Species

Most Drosophila species sing species-specific pulse songs during their “precopulatory courtship.” Three sibling species of the Drosophila montium species subgroup performed “copulatory courtship”: males generated courtship songs by vibrating either wing only after mounting and during copulation. In these three species, strong sexual isolation was detected between D. ohnishii and D. lini and between D. ohnishii and D. ogumai, but not between D. lini and D. ogumai. Female showed strong repelling behavior when they were mounted by a heterospecific male in the species combinations including D. ohnishii, resulting in failure of the copulation attempt of the male. Acoustic analyses of courtship songs revealed that the pulse song was irregular, without any species-specific parameters, but that the frequency of the sine song was different among the three species in accordance with the modes of sexual isolation between them; it was significantly lower in D. ohnishii (mean ± SE = 193.0 ± 1.7 Hz) but higher in D. lini (253.4 ± 2.7 Hz) and D. ogumai (246.7 ± 5.3 Hz). We suggest that this difference in the sine song frequency is a sexual signal in the Specific Mate Recognition System (SMRS) among these three Drosophila species.

Evolutionary Relationships in the Drosophila ananassae Species Cluster Based on Introns of Multiple Nuclear Loci

The Drosophila ananassae species cluster includes D. ananassae, D. pallidosa, D. parapallidosa, and the cryptic species “pallidosa-like”, “pallidosa-like Wau” and “papuensis-like” Some of the taxa are sympatric in the South Pacific, Papua New Guinea, and Southeast Asia, and gene flow between different taxa has been suspected for a handful of genes. In the present analysis, we examined DNA sequences of introns in four loci: alpha actinin (Actn) on XL, white (w) on XR, CG7785 on 2L, and zinc ion transmembrane transporter 63C (ZnT63C) on 2R. Phylogenetic trees (neighbor-joining and haplotype network) were inconsistent among these loci. Some haplotypes shared between taxa were found for w, CG7785, and ZnT63C, suggesting recent gene flow. However, no haplotypes were shared, for example, between D. ananassae and D. pallidosa for CG7785, which is close to the proximal breakpoint of In(2L)D. This suggests that taxon-specific inversions prevent gene flow, as predicted by the chromosomal speciation hypothesis.

Genetic decay of balancer chromosomes in Drosophila melanogaster

Theoretical considerations predict that balancer chromosomes in Drosophila melanogaster should accumulate numerous deleterious mutations with time. We counted the number of recessive lethal mutations on two balancer chromosomes from the In(2LR)SM1/In(2LR)Pm strain maintained in our lab, after making the balancers heterozygous with deficiencies from second-chromosome Kyoto Deficiency kit strains. We detected 10 recessive lethal mutations in the balancer In(2LR)Pm, which is consistent with the mutation rate estimated previously. However, we detected only three mutations, a significantly smaller number, in the balancer In(2LR)SM1, although this may be an artifact. In conclusion, we observed genetic decay over an estimable timescale by using balancers with historical records. Thus, balancers of any strain may have accumulated many unidentified recessive lethal mutations.