RESUMO
Spatially controlled release of sister chromatid cohesion during progression through the meiotic divisions is of paramount importance for error-free chromosome segregation during meiosis. Cohesion is mediated by the cohesin protein complex and cleavage of one of its subunits by the endoprotease separase removes cohesin first from chromosome arms during exit from meiosis I and later from the pericentromeric region during exit from meiosis II. At the onset of the meiotic divisions, cohesin has also been proposed to be present within the centromeric region for the unification of sister centromeres into a single functional entity, allowing bipolar orientation of paired homologs within the meiosis I spindle. Separase-mediated removal of centromeric cohesin during exit from meiosis I might explain sister centromere individualization which is essential for subsequent biorientation of sister centromeres during meiosis II. To characterize a potential involvement of separase in sister centromere individualization before meiosis II, we have studied meiosis in Drosophila melanogaster males where homologs are not paired in the canonical manner. Meiosis does not include meiotic recombination and synaptonemal complex formation in these males. Instead, an alternative homolog conjunction system keeps homologous chromosomes in pairs. Using independent strategies for spermatocyte-specific depletion of separase complex subunits in combination with time-lapse imaging, we demonstrate that separase is required for the inactivation of this alternative conjunction at anaphase I onset. Mutations that abolish alternative homolog conjunction therefore result in random segregation of univalents during meiosis I also after separase depletion. Interestingly, these univalents become bioriented during meiosis II, suggesting that sister centromere individualization before meiosis II does not require separase.
Assuntos
Centrômero/genética , Pareamento Cromossômico/genética , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Meiose/genética , Separase/genética , Animais , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas Cromossômicas não Histona/genética , Proteínas Cromossômicas não Histona/metabolismo , Segregação de Cromossomos/genética , Cinetocoros/fisiologia , Masculino , Interferência de RNA , RNA Interferente Pequeno/genética , CoesinasAssuntos
Segregação de Cromossomos , Drosophila , Animais , Cromossomos , Drosophila/genética , Masculino , Meiose/genética , SeparaseRESUMO
Cohesion between sister chromatids is mediated by cohesin and is essential for proper meiotic segregation of both sister chromatids and homologs. solo encodes a Drosophila meiosis-specific cohesion protein with no apparent sequence homology to cohesins that is required in male meiosis for centromere cohesion, proper orientation of sister centromeres and centromere enrichment of the cohesin subunit SMC1. In this study, we show that solo is involved in multiple aspects of meiosis in female Drosophila. Null mutations in solo caused the following phenotypes: 1) high frequencies of homolog and sister chromatid nondisjunction (NDJ) and sharply reduced frequencies of homolog exchange; 2) reduced transmission of a ring-X chromosome, an indicator of elevated frequencies of sister chromatid exchange (SCE); 3) premature loss of centromere pairing and cohesion during prophase I, as indicated by elevated foci counts of the centromere protein CID; 4) instability of the lateral elements (LE)s and central regions of synaptonemal complexes (SCs), as indicated by fragmented and spotty staining of the chromosome core/LE component SMC1 and the transverse filament protein C(3)G, respectively, at all stages of pachytene. SOLO and SMC1 are both enriched on centromeres throughout prophase I, co-align along the lateral elements of SCs and reciprocally co-immunoprecipitate from ovarian protein extracts. Our studies demonstrate that SOLO is closely associated with meiotic cohesin and required both for enrichment of cohesin on centromeres and stable assembly of cohesin into chromosome cores. These events underlie and are required for stable cohesion of centromeres, synapsis of homologous chromosomes, and a recombination mechanism that suppresses SCE to preferentially generate homolog crossovers (homolog bias). We propose that SOLO is a subunit of a specialized meiotic cohesin complex that mediates both centromeric and axial arm cohesion and promotes homolog bias as a component of chromosome cores.
Assuntos
Proteínas de Ciclo Celular/genética , Proteínas Cromossômicas não Histona/genética , Proteínas de Drosophila/genética , Drosophila/genética , Meiose/genética , Animais , Proteínas de Ciclo Celular/metabolismo , Centrômero/genética , Proteínas Cromossômicas não Histona/metabolismo , Segregação de Cromossomos/genética , Cromossomos/genética , Proteínas de Drosophila/metabolismo , Feminino , Masculino , Recombinação Genética/genética , Troca de Cromátide Irmã/genética , Complexo Sinaptonêmico/genéticaRESUMO
Homologous pairing establishes the foundation for accurate reductional segregation during meiosis I in sexual organisms. This Commentary summarizes recent progress in our understanding of homologous pairing in meiosis, and will focus on the characteristics and mechanisms of specialized chromosome sites, called pairing centers (PCs), in Caenorhabditis elegans and Drosophila melanogaster. In C. elegans, each chromosome contains a single PC that stabilizes chromosome pairing and initiates synapsis of homologous chromosomes. Specific zinc-finger proteins recruited to PCs link chromosomes to nuclear envelope proteins--and through them to the microtubule cytoskeleton--thereby stimulating chromosome movements in early prophase, which are thought to be important for homolog sorting. This mechanism appears to be a variant of the 'telomere bouquet' process, in which telomeres cluster on the nuclear envelope, connect chromosomes through nuclear envelope proteins to the cytoskeleton and lead chromosome movements that promote homologous synapsis. In Drosophila males, which undergo meiosis without recombination, pairing of the largely non-homologous X and Y chromosomes occurs at specific repetitive sequences in the ribosomal DNA. Although no other clear examples of PC-based pairing mechanisms have been described, there is evidence for special roles of telomeres and centromeres in aspects of chromosome pairing, synapsis and segregation; these roles are in some cases similar to those of PCs.
Assuntos
Pareamento Cromossômico , Meiose/genética , Animais , Pareamento de Bases , Caenorhabditis elegans , Drosophila melanogaster , Feminino , Masculino , Recombinação GenéticaRESUMO
Drosophila males undergo meiosis without recombination or chiasmata but homologous chromosomes pair and disjoin regularly. The X-Y pair utilizes a specific repeated sequence within the heterochromatic ribosomal DNA blocks as a pairing site. No pairing sites have yet been identified for the autosomes. To search for such sites, we utilized probes targeting specific heterochromatic regions to assay heterochromatin pairing sequences and behavior in meiosis by fluorescence in situ hybridization (FISH). We found that the small fourth chromosome pairs at heterochromatic region 61 and associates with the X chromosome throughout prophase I. Homolog pairing of the fourth chromosome is disrupted when the homolog conjunction complex is perturbed by mutations in SNM or MNM. On the other hand, six tested heterochromatic regions of the major autosomes proved to be largely unpaired after early prophase I, suggesting that stable homolog pairing sites do not exist in heterochromatin of the major autosomes. Furthermore, FISH analysis revealed two distinct patterns of sister chromatid cohesion in heterochromatin: regions with stable cohesion and regions lacking cohesion. This suggests that meiotic sister chromatid cohesion is incomplete within heterochromatin and may occur at specific preferential sites.
Assuntos
Cromátides , Drosophila melanogaster/genética , Heterocromatina/química , Meiose , Espermatócitos/metabolismo , Cromossomo X/química , Cromossomo Y/química , Animais , Centrômero/química , Centrômero/genética , Cromátides/química , Cromátides/genética , Cromátides/metabolismo , Pareamento Cromossômico , Segregação de Cromossomos , Fluorescência , Heterocromatina/genética , Hibridização in Situ Fluorescente , Masculino , Sondas Moleculares/análise , Mutação , Espermatócitos/citologia , Cromossomo X/genética , Cromossomo Y/genéticaRESUMO
A wide variety of techniques have been utilized to determine the localization of various proteins from premeiotic through meiotic stages in Drosophila males. Live imaging has been instrumental in monitoring chromosome pairing and the localization of fusion proteins. Immunofluorescence has been a widely utilized technique to examine the localization and colocalization of the many proteins involved in meiosis. Recently, an immuno-FISH protocol was developed to observe the co-localization of DNA probes and proteins. In this chapter, detailed protocols outlining these three types of experiments are presented.
Assuntos
Proteínas Cromossômicas não Histona/metabolismo , Pareamento Cromossômico/fisiologia , Análise Citogenética/métodos , Drosophila/citologia , Drosophila/genética , Espermatócitos/citologia , Animais , Sobrevivência Celular/fisiologia , Proteínas Cromossômicas não Histona/análise , Mapeamento Cromossômico/métodos , Masculino , Modelos Biológicos , Espermatócitos/metabolismoRESUMO
In most eukaryotes, segregation of homologous chromosomes during meiosis is dependent on crossovers that occur while the homologs are intimately paired during early prophase. Crossovers generate homolog connectors known as chiasmata that are stabilized by cohesion between sister-chromatid arms. In Drosophila males, homologs pair and segregate without recombining or forming chiasmata. Stable pairing of homologs is dependent on two proteins, SNM and MNM, that associate with chromosomes throughout meiosis I until their removal at anaphase I. SNM and MNM localize to the rDNA region of the X-Y pair, which contains 240-bp repeats that have previously been shown to function as cis-acting chromosome pairing/segregation sites. Here we show that heterochromatic mini-X chromosomes lacking native rDNA but carrying transgenic 240-bp repeat arrays segregate preferentially from full-length sex chromosomes and from each other. Mini-X pairs do not form autonomous bivalents but do associate at high frequency with the X-Y bivalent to form trivalents and quadrivalents. Both disjunction of mini-X pairs and multivalent formation are dependent on the presence of SNM and MNM. These results imply that 240-bp repeats function to mediate association of sex chromosomes with SNM and MNM.
Assuntos
Proteínas de Ciclo Celular/genética , Pareamento Cromossômico , DNA Ribossômico/genética , Proteínas de Drosophila/genética , Drosophila/genética , Meiose , Cromossomo X , Animais , Feminino , Masculino , Sequências Repetitivas de Ácido NucleicoRESUMO
Homologous chromosomes must pair and establish stable connections during prophase I of meiosis to segregate reliably from each other at anaphase I. In most organisms, the stable connections, called chiasmata, arise from crossovers. In Drosophila males, homologs pair and segregate without crossing over. Chiasmata are replaced by a homolog conjunction complex that includes the Stromalin in Meiosis (SNM) and Modifier of Mdg4 in Meiosis (MNM) proteins. MNM is one of 31 alternative splice products of mod(mdg4), all of which share a common 402-amino-acid N terminus and differ at their C termini. Previous data demonstrated that an MNM-specific exon is required for homolog conjunction, but did not address whether the N-terminal common region, which includes a BTB domain that can mediate coalescence of protein-DNA complexes, is also required. Here we describe a mutation in the common region of mod(mdg4), Z3-3401, that causes qualitatively similar phenotypes as the MNM-specific alleles but disrupts X-Y segregation much more drastically than autosomal segregation. The mutant MNM protein in Z3-3401 is expressed throughout prophase I in spermatocytes but the protein is confined to the cytoplasm, suggesting that the Z3-3401 mutation disrupts a signal required for nuclear localization or retention. Z3-3401 fails to complement a large battery of lethal and semilethal alleles in the common region for meiotic nondisjunction, including an allele containing an amino acid substitution at a conserved residue in the BTB/POZ domain, consistent with a general requirement for the mod(mdg4) common region in homolog segregation.
Assuntos
Segregação de Cromossomos/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Meiose , Fatores de Transcrição/metabolismo , Alelos , Processamento Alternativo , Sequência de Aminoácidos , Substituição de Aminoácidos , Animais , Instabilidade Cromossômica/genética , Quebra Cromossômica , Pareamento Cromossômico/genética , Proteínas de Drosophila/química , Proteínas de Drosophila/genética , Drosophila melanogaster/citologia , Teste de Complementação Genética , Masculino , Dados de Sequência Molecular , Não Disjunção Genética , Sinais de Localização Nuclear/metabolismo , Fenótipo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Fatores de Transcrição/química , Fatores de Transcrição/genética , Cromossomo X/genética , Cromossomo Y/genéticaRESUMO
Rad51 is a crucial enzyme in DNA repair, mediating the strand invasion and strand exchange steps of homologous recombination (HR). Mutations in the Drosophila Rad51 gene (spn-A) disrupt somatic as well as meiotic double-strand break (DSB) repair, similar to fungal Rad51 genes. However, the sterility of spn-A mutant females prevented a thorough analysis of the role of Rad51 in meiosis. In this study, we generated transgenic animals that express spn-A dsRNA under control of an inducible promoter, and examined the effects of inhibiting expression of spn-A on DNA repair, meiotic recombination and meiotic chromosome pairing and segregation. We found that depletion of spn-A mRNA had no effect on the viability of non-mutagen-treated transgenic animals but greatly reduced the survival of larvae that were exposed to the radiomimetic drug MMS, in agreement with the MMS and X-ray sensitivity of spn-A mutant animals. We also found that increases in dose of spn-A gene enhanced larval resistance to MMS exposure, suggesting that at high damage levels, Rad51 protein levels may be limiting for DNA repair. spn-A RNAi strongly stimulated X-X nondisjunction and decreased recombination along the X in female meiosis, consistent with a requirement of Rad51 in meiotic recombination. However, neither RNAi directed against the spn-A mRNA nor homozygosity for a spn-A null mutation had any effect on male fertility or on X-Y segregation in male meiosis, indicating that Rad51 likely plays no role in male meiotic chromosome pairing. Our results support a central role for Rad51 in HR in both somatic and meiotic DSB repair, but indicate that Rad51 in Drosophila is dispensable for meiotic chromosome pairing. Our results also provide the first demonstration that RNAi can be used to inhibit the functions of meiotic genes in Drosophila.
Assuntos
Reparo do DNA , Proteínas de Ligação a DNA/fisiologia , Proteínas de Drosophila/fisiologia , Drosophila melanogaster/enzimologia , Meiose/genética , Recombinação Genética , Animais , Animais Geneticamente Modificados , Pareamento Cromossômico , Segregação de Cromossomos , Troca Genética , Dano ao DNA , Drosophila melanogaster/metabolismo , Feminino , Homozigoto , Larva , Masculino , Mutação , RNA Interferente Pequeno/farmacologia , Rad51 Recombinase , Tolerância a Radiação/genéticaRESUMO
Assembly of the synaptonemal complex (SC) in Drosophila depends on two independent pathways defined by the chromosome axis proteins C(2)M and ORD. Because C(2)M encodes a Kleisin-like protein and ORD is required for sister-chromatid cohesion, we tested the hypothesis that these two SC assembly pathways depend on two cohesin complexes. Through single- and double-mutant analysis to study the mitotic cohesion proteins Stromalin (SA) and Nipped-B (SCC2) in meiosis, we provide evidence that there are at least two meiosis-specific cohesin complexes. One complex depends on C(2)M, SA, and Nipped-B. Despite the presence of mitotic cohesins SA and Nipped-B, this pathway has only a minor role in meiotic sister-centromere cohesion and is primarily required for homolog interactions. C(2)M is continuously incorporated into pachytene chromosomes even though SC assembly is complete. In contrast, the second complex, which depends on meiosis-specific proteins SOLO, SUNN, and ORD is required for sister-chromatid cohesion, localizes to the centromeres and is not incorporated during prophase. Our results show that the two cohesin complexes have unique functions and are regulated differently. Multiple cohesin complexes may provide the diversity of activities required by the meiotic cell. For example, a dynamic complex may allow the chromosomes to regulate meiotic recombination, and a stable complex may be required for sister-chromatid cohesion.
Assuntos
Proteínas de Ciclo Celular/genética , Proteínas Cromossômicas não Histona/genética , Drosophila melanogaster/genética , Regulação da Expressão Gênica , Proteínas de Insetos/genética , Complexo Sinaptonêmico/genética , Animais , Proteínas de Ciclo Celular/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Pareamento Cromossômico , Segregação de Cromossomos , Drosophila melanogaster/metabolismo , Feminino , Proteínas de Insetos/metabolismo , CoesinasRESUMO
Pairing of homologous chromosomes is an essential feature of meiosis, acting to promote high levels of recombination and to ensure segregation of homologs. However, homologous pairing also occurs in somatic cells, most regularly in Dipterans such as Drosophila, but also to a lesser extent in other organisms, and it is not known how mitotic and meiotic pairing relate to each other. In this article, I summarize results of recent molecular studies of pairing in both mitosis and meiosis, focusing especially on studies using fluorescent in situ hybridization (FISH) and GFP-tagging of single loci, which have allowed investigators to assay the pairing status of chromosomes directly. These approaches have permitted the demonstration that pairing occurs throughout the cell cycle in mitotic cells in Drosophila, and that the transition from mitotic to meiotic pairing in spermatogenesis is accompanied by a dramatic increase in pairing frequency. Similar approaches in mammals, plants and fungi have established that with few exceptions, chromosomes enter meiosis unpaired and that chromosome movements involving the telomeric, and sometimes centromeric, regions often precede the onset of meiotic pairing. The possible roles of proteins involved in homologous recombination, synapsis and sister chromatid cohesion in homolog pairing are discussed with an emphasis on those for which mutant phenotypes have permitted an assessment of effects on homolog pairing. Finally, I consider the question of the distribution and identity of chromosomal pairing sites, using recent data to evaluate possible relationships between pairing sites and other chromosomal sites, such as centromeres, telomeres, promoters and heterochromatin. I cite evidence that may point to a relationship between matrix attachment sites and homologous pairing sites.
Assuntos
Pareamento Cromossômico , Meiose/genética , Mitose/genética , Animais , Centrômero/genética , Centrômero/metabolismo , Drosophila/genética , Humanos , Matriz Nuclear/genética , Matriz Nuclear/metabolismo , Recombinação Genética , Telômero/genética , Telômero/metabolismoRESUMO
Regular meiotic chromosome segregation requires sister centromeres to mono-orient (orient to the same pole) during the first meiotic division (meiosis I) when homologous chromosomes segregate, and to bi-orient (orient to opposite poles) during the second meiotic division (meiosis II) when sister chromatids segregate. Both orientation patterns require cohesion between sister centromeres, which is established during meiotic DNA replication and persists until anaphase of meiosis II. Meiotic cohesion is mediated by a conserved four-protein complex called cohesin that includes two structural maintenance of chromosomes (SMC) subunits (SMC1 and SMC3) and two non-SMC subunits. In Drosophila melanogaster, however, the meiotic cohesion apparatus has not been fully characterized and the non-SMC subunits have not been identified. We have identified a novel Drosophila gene called sisters unbound (sunn), which is required for stable sister chromatid cohesion throughout meiosis. sunn mutations disrupt centromere cohesion during prophase I and cause high frequencies of non-disjunction (NDJ) at both meiotic divisions in both sexes. SUNN co-localizes at centromeres with the cohesion proteins SMC1 and SOLO in both sexes and is necessary for the recruitment of both proteins to centromeres. Although SUNN lacks sequence homology to cohesins, bioinformatic analysis indicates that SUNN may be a structural homolog of the non-SMC cohesin subunit stromalin (SA), suggesting that SUNN may serve as a meiosis-specific cohesin subunit. In conclusion, our data show that SUNN is an essential meiosis-specific Drosophila cohesion protein.
Assuntos
Centrômero/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/citologia , Drosophila melanogaster/metabolismo , Meiose , Animais , Proteínas de Ciclo Celular/química , Cromátides/metabolismo , Proteínas Cromossômicas não Histona/química , Segregação de Cromossomos , Proteínas de Drosophila/química , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Feminino , Genes de Insetos , Loci Gênicos , Masculino , Prófase Meiótica I , Mutação/genética , Não Disjunção Genética , Oócitos/citologia , Oócitos/metabolismo , Ligação Proteica , Homologia de Sequência de Aminoácidos , Cromossomos Sexuais , Espermatozoides/citologia , Espermatozoides/metabolismo , CoesinasRESUMO
Meiosis entails sorting and separating both homologous and sister chromatids. The mechanisms for connecting sister chromatids and homologs during meiosis are highly conserved and include specialized forms of the cohesin complex and a tightly regulated homolog synapsis/recombination pathway designed to yield regular crossovers between homologous chromatids. Drosophila male meiosis is of special interest because it dispenses with large segments of the standard meiotic script, particularly recombination, synapsis and the associated structures. Instead, Drosophila relies on a unique protein complex composed of at least two novel proteins, SNM and MNM, to provide stable connections between homologs during meiosis I. Sister chromatid cohesion in Drosophila is mediated by cohesins, ring-shaped complexes that entrap sister chromatids. However, unlike other eukaryotes Drosophila does not rely on the highly conserved Rec8 cohesin in meiosis, but instead utilizes two novel cohesion proteins, ORD and SOLO, which interact with the SMC1/3 cohesin components in providing meiotic cohesion.
RESUMO
Sister chromatid cohesion is essential to maintain stable connections between homologues and sister chromatids during meiosis and to establish correct centromere orientation patterns on the meiosis I and II spindles. However, the meiotic cohesion apparatus in Drosophila melanogaster remains largely uncharacterized. We describe a novel protein, sisters on the loose (SOLO), which is essential for meiotic cohesion in Drosophila. In solo mutants, sister centromeres separate before prometaphase I, disrupting meiosis I centromere orientation and causing nondisjunction of both homologous and sister chromatids. Centromeric foci of the cohesin protein SMC1 are absent in solo mutants at all meiotic stages. SOLO and SMC1 colocalize to meiotic centromeres from early prophase I until anaphase II in wild-type males, but both proteins disappear prematurely at anaphase I in mutants for mei-S332, which encodes the Drosophila homologue of the cohesin protector protein shugoshin. The solo mutant phenotypes and the localization patterns of SOLO and SMC1 indicate that they function together to maintain sister chromatid cohesion in Drosophila meiosis.
Assuntos
Proteínas de Ciclo Celular/metabolismo , Centrômero/metabolismo , Cromátides/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Pareamento Cromossômico/fisiologia , Proteínas de Drosophila/metabolismo , Anáfase/fisiologia , Animais , Proteínas de Ciclo Celular/genética , Centrômero/genética , Cromátides/genética , Proteínas Cromossômicas não Histona/genética , Proteínas de Drosophila/genética , Drosophila melanogaster , Feminino , Masculino , Mutação , Fuso AcromáticoRESUMO
In Drosophila males, homologous chromosomes segregate by an unusual process involving physical connections not dependent on recombination. We have identified two meiotic proteins specifically required for this process. Stromalin in Meiosis (SNM) is a divergent member of the SCC3/SA/STAG family of cohesin proteins, and Modifier of Mdg4 in Meiosis (MNM) is one of many BTB-domain proteins expressed from the mod(mdg4) locus. SNM and MNM colocalize along with a repetitive rDNA sequence known to function as an X-Y pairing site to nucleolar foci during meiotic prophase and to a compact structure associated with the X-Y bivalent during prometaphase I and metaphase I. Additionally, MNM localizes to autosomal foci throughout meiosis I. These proteins are mutually dependent for their colocalization, and at least MNM requires the function of teflon, another meiotic gene. SNM and MNM do not colocalize with SMC1, suggesting that the homolog conjunction mechanism is independent of cohesin.
Assuntos
Proteínas de Ciclo Celular/fisiologia , Segregação de Cromossomos/fisiologia , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Meiose/fisiologia , Fatores de Transcrição/genética , Animais , Proteínas de Ciclo Celular/genética , Divisão do Núcleo Celular/genética , Proteínas Cromossômicas não Histona/genética , Aberrações Cromossômicas , Pareamento Cromossômico/genética , Segregação de Cromossomos/genética , Cromossomos/genética , Cromossomos/metabolismo , DNA Ribossômico/genética , Dípteros/genética , Proteínas de Drosophila/fisiologia , Drosophila melanogaster/fisiologia , Éxons/genética , Feminino , Proteínas Fúngicas/genética , Masculino , Meiose/genética , Dados de Sequência Molecular , Mutação/genética , Não Disjunção Genética , Proteínas Nucleares/genética , Fenótipo , Filogenia , Isoformas de Proteínas/genética , Cromossomos Sexuais/genética , Fatores de Transcrição/fisiologia , CoesinasRESUMO
Among proteins involved in homologous recombination, Rad51 is an essential enzyme in DNA repair and recombination. However, little is known about its role in cell cycle regulation and apoptosis. To examine the function of Drosophila Rad51 (DmRad51) in cell cycle regulation and apoptosis, DmRad51 protein was overexpressed using a heat shock-inducible promoter or the UAS-GAL4 binary expression system. We observed that ubiquitous expression of DmRad51 protein in flies carrying hsp26- Rad51 or UAS- Rad51 transgenes was lethal. Induction of DmRad51--more specifically in eye or wing imaginal discs--caused tissue-specific cell death in the domains of DmRad51 expression. Cell death was due to apoptosis, as shown by staining with the TdT-mediated dUTP nick-end labeling assay. Immunocytochemistry revealed that cells expressing DmRad51 colocalized with apoptotic cells. In addition, the phenotypes caused by the overexpression of DmRad51 were similar to those caused by ectopic expression of Reaper, a proapoptotic protein, and were partially suppressed by the coexpression of p35, an antiapoptotic protein. Using an antiphosphohistone H3 antibody, we also observed that the overexpression of DmRad51 protein disrupted normal cell cycle progression in eye imaginal discs. Taken together, these results show that ectopically expressed DmRad51 disrupts cell cycle regulation and induces apoptosis.