RESUMO
RanGTP is important for chromosome-dependent spindle assembly in Xenopus extracts. Here we report on experiments to determine the role of the Ran pathway on microtubule dynamics in Drosophila oocytes and embryos. Females expressing a dominant-negative form of Ran have fertility defects, suggesting that RanGTP is required for normal fertility. This is not, however, because of a defect in acentrosomal meiotic spindle assembly. Therefore, RanGTP does not appear to be essential or sufficient for the formation of the acentrosomal spindle. Instead, the most important function of the Ran pathway in spindle assembly appears to be in the tapering of microtubules at the spindle poles, which might be through regulation of proteins such as TACC and the HURP homolog, Mars. One consequence of this spindle organization defect is an increase in the nondisjunction of achiasmate chromosomes. However, the meiotic defects are not severe enough to cause the decreased fertility. Reductions in fertility occur because RanGTP has a role in microtubule assembly that is not directly nucleated by the chromosomes. This includes microtubules nucleated from the sperm aster, which are required for pronuclear fusion. We propose that following nuclear envelope breakdown, RanGTP is released from the nucleus and creates a cytoplasm that is activated for assembling microtubules, which is important for processes such as pronuclear fusion. Around the chromosomes, however, RanGTP might be redundant with other factors such as the chromosome passenger complex.
Assuntos
Proteínas de Drosophila/metabolismo , Drosophila/embriologia , Drosophila/enzimologia , Fuso Acromático/metabolismo , Proteína ran de Ligação ao GTP/metabolismo , Animais , Segregação de Cromossomos , Cromossomos de Insetos/genética , Drosophila/citologia , Drosophila/genética , Proteínas de Drosophila/genética , Feminino , Guanosina Trifosfato/metabolismo , Masculino , Microtúbulos/genética , Microtúbulos/metabolismo , Mitose , Oócitos/citologia , Oócitos/enzimologia , Oócitos/metabolismo , Fuso Acromático/genética , Proteína ran de Ligação ao GTP/genéticaRESUMO
At one end of each Caenorhabditis elegans chromosome is a locus required for meiotic crossing over. Recent studies have shown that these sites mediate chromosome pairing and synapsis during meiosis, and that each site contains binding sites for a non-canonical C2H2 zinc finger protein.
Assuntos
Proteínas de Caenorhabditis elegans/genética , Caenorhabditis elegans/genética , Pareamento Cromossômico , Troca Genética , Meiose , Animais , Sítios de Ligação , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Dedos de Zinco/genética , Dedos de Zinco/fisiologiaRESUMO
Chromosomes have multiple roles both in controlling the cell assembly and structure of the spindle and in determining chromosomal position on the spindle in many meiotic cells and in some types of mitotic cells. Moreover, functionally significant chromosome-microtubule interactions are not limited to the kinetochore but are also mediated by proteins localized along the arms of chromosomes. Finally, chromosomes also play a crucial role in control of the cell cycle.
Assuntos
Ciclo Celular , Cromossomos/fisiologia , Meiose , Anáfase , Animais , Cromossomos/ultraestrutura , Proteínas de Ligação a DNA/fisiologia , Cinesinas/fisiologia , Cinetocoros/fisiologia , Metáfase , Proteínas dos Microtúbulos/fisiologia , Microtúbulos/fisiologia , Microtúbulos/ultraestrutura , Mutação , Proteínas Nucleares/fisiologia , Fuso Acromático/fisiologia , Fuso Acromático/ultraestruturaRESUMO
Although in Saccharomyces cerevisiae the initiation of meiotic recombination, as indicated by double-strand break formation, appears to be functionally linked to the initiation of synapsis, meiotic chromosome synapsis in Drosophila females occurs in the absence of meiotic exchange. Electron microscopy of oocytes from females homozygous for either of two meiotic mutants (mei-W68 and mei-P22), which eliminate both meiotic crossing over and gene conversion, revealed normal synaptonemal complex formation. Thus, synapsis in Drosophila is independent of meiotic recombination, consistent with a model in which synapsis is required for the initiation of meiotic recombination. Furthermore, the basic processes of early meiosis may have different functional or temporal relations, or both, in yeast and Drosophila.
Assuntos
Cromossomos/fisiologia , Drosophila melanogaster/fisiologia , Meiose , Recombinação Genética , Complexo Sinaptonêmico/fisiologia , Animais , Cromossomos/genética , Cromossomos/ultraestrutura , Troca Genética , Drosophila melanogaster/genética , Feminino , Conversão Gênica , Mutação , Oócitos/fisiologia , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/fisiologia , Troca de Cromátide IrmãRESUMO
Using an antibody against the phosphorylated form of His2Av (gamma-His2Av), we have described the time course for the series of events leading from the formation of a double-strand break (DSB) to a crossover in Drosophila female meiotic prophase. MEI-P22 is required for DSB formation and localizes to chromosomes prior to gamma-His2Av foci. Drosophila females, however, are among the group of organisms where synaptonemal complex (SC) formation is not dependent on DSBs. In the absence of two SC proteins, C(3)G and C(2)M, the number of DSBs in oocytes is significantly reduced. This is consistent with the appearance of SC protein staining prior to gamma-His2Av foci. However, SC formation is incomplete or absent in the neighboring nurse cells, and gamma-His2Av foci appear with the same kinetics as in oocytes and do not depend on SC proteins. Thus, competence for DSB formation in nurse cells occurs with a specific timing that is independent of the SC, whereas in the oocytes, some SC proteins may have a regulatory role to counteract the effects of a negative regulator of DSB formation. The SC is not sufficient for DSB formation, however, since DSBs were absent from the heterochromatin even though SC formation occurs in these regions. All gamma-His2Av foci disappear before the end of prophase, presumably as repair is completed and crossovers are formed. However, oocytes in early prophase exhibit a slower response to X-ray-induced DSBs compared to those in the late pachytene stage. Assuming all DSBs appear as gamma-His2Av foci, there is at least a 3:1 ratio of noncrossover to crossover products. From a comparison of the frequency of gamma-His2Av foci and crossovers, it appears that Drosophila females have only a weak mechanism to ensure a crossover in the presence of a low number of DSBs.
Assuntos
Quebras de DNA de Cadeia Dupla , Reparo do DNA/fisiologia , Drosophila melanogaster/genética , Meiose/fisiologia , Animais , Animais Geneticamente Modificados , Proteínas Cromossômicas não Histona , Troca Genética/fisiologia , Quebras de DNA de Cadeia Dupla/efeitos da radiação , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Drosophila melanogaster/efeitos da radiação , Feminino , Heterocromatina/fisiologia , Histonas/genética , Histonas/metabolismo , Meiose/efeitos da radiação , Prófase Meiótica I/fisiologia , Mutação , Oócitos/citologia , Oócitos/efeitos da radiação , Estágio Paquíteno/efeitos da radiação , Fosforilação , Complexo Sinaptonêmico/fisiologia , Fatores de Tempo , Raios XRESUMO
In the oocytes of many species, bipolar spindles form in the absence of centrosomes. Drosophila melanogaster oocyte chromosomes have a major role in nucleating microtubules, which precedes the bundling and assembly of these microtubules into a bipolar spindle. Here we present evidence that a region similar to the anaphase central spindle functions to organize acentrosomal spindles. Subito mutants are characterized by the formation of tripolar or monopolar spindles and nondisjunction of homologous chromosomes at meiosis I. Subito encodes a kinesinlike protein and associates with the meiotic central spindle, consistent with its classification in the Kinesin 6/MKLP1 family. This class of proteins is known to be required for cytokinesis, but our results suggest a new function in spindle formation. The meiotic central spindle appears during prometaphase and includes passenger complex proteins such as AurB and Incenp. Unlike mitotic cells, the passenger proteins do not associate with centromeres before anaphase. In the absence of Subito, central spindle formation is defective and AurB and Incenp fail to properly localize. We propose that Subito is required for establishing and/or maintaining the central spindle in Drosophila oocytes, and this substitutes for the role of centrosomes in organizing the bipolar spindle.
Assuntos
Centrossomo/fisiologia , Proteínas de Drosophila/fisiologia , Drosophila melanogaster/fisiologia , Cinesinas/fisiologia , Meiose/fisiologia , Fuso Acromático/fisiologia , Sequência de Aminoácidos , Animais , Drosophila melanogaster/ultraestrutura , Feminino , Dados de Sequência Molecular , Oócitos/fisiologia , Homologia de Sequência de Aminoácidos , Fuso Acromático/ultraestruturaRESUMO
We have isolated and characterized 76 duplications of chromosome I in the genome of Caenorhabditis elegans. The region studied is the 20 map unit left half of the chromosome. Sixty-two duplications were induced with gamma radiation and 14 arose spontaneously. The latter class was apparently the result of spontaneous breaks within the parental duplication. The majority of duplications behave as if they are free. Three duplications are attached to identifiable sequences from other chromosomes. The duplication breakpoints have been mapped by complementation analysis relative to genes on chromosome I. Nineteen duplication breakpoints and seven deficiency breakpoints divide the left half of the chromosome into 24 regions. We have studied the relationship between duplication size and segregational stability. While size is an important determinant of mitotic stability, it is not the only one. We observed clear exceptions to a size-stability correlation. In addition to size, duplication stability may be influenced by specific sequences or chromosome structure. The majority of the duplications were stable enough to be powerful tools for gene mapping. Therefore the duplications described here will be useful in the genetic characterization of chromosome I and the techniques we have developed can be adapted to other regions of the genome.
Assuntos
Caenorhabditis/genética , Cromossomos , Família Multigênica , Mutação , Animais , Caenorhabditis/efeitos da radiação , Deleção Cromossômica , Mapeamento Cromossômico , Feminino , Raios gama , Genótipo , Masculino , Meiose , Mitose , Recombinação GenéticaRESUMO
Previous studies have shown that isolated portions of Caenorhabditis elegans chromosomes are not equally capable of meiotic exchange. These results led to the proposal that a homolog recognition region (HRR), defined as the region containing those sequences enabling homologous chromosomes to pair and recombine, is localized near one end of each chromosome. Using translocations and duplications we have localized the chromosome I HRR to the right end. Whereas the other half of chromosome I did not confer any ability for homologs to pair and recombine, deficiencies in this region dominantly suppressed recombination to the middle of the chromosome. These deletions may have disrupted pairing mechanisms that are secondary to and require an HRR. Thus, the processes of pairing and recombination appear to utilize at least two chromosomal elements, the HRR and other pairing sites. For example, terminal sequences from other chromosomes increase the ability of free duplications to recombine with their normal homologs, suggesting that telomere-associated sequences, homologous or nonhomologous, play a role in facilitating meiotic exchange. Recombination can also initiate at internal sites separated from the HRR by chromosome rearrangement, such as deletions of the unc-54 region of chromosome I. When crossing over was suppressed in a region of chromosome I, compensatory increases were observed in other regions. Thus, the presence of the HRR enabled recombination to occur but did not determine the distribution of the crossover events. It seems most likely that there are multiple initiation sites for recombination once homolog recognition has been achieved.
Assuntos
Caenorhabditis elegans/genética , Meiose/genética , Recombinação Genética , Animais , Mecanismo Genético de Compensação de Dose , Heterozigoto , Família Multigênica , Translocação Genética , Cromossomo XRESUMO
In the nematode Caenorhabditis elegans, recombination suppression in translocation heterozygotes is severe and extensive. We have examined the meiotic properties of two translocations involving chromosome I, szT1(I;X) and hT1(I;V). No recombination was observed in either of these translocation heterozygotes along the left (let-362-unc-13) 17 map units of chromosome I. Using half-translocations as free duplications, we mapped the breakpoints of szT1 and hT1. The boundaries of crossover suppression coincided with the physical breakpoints. We propose that DNA sequences at the right end of chromosome I facilitate pairing and recombination. We use the data from translocations of other chromosomes to map the location of pairing sites on four other chromosomes. hT1 and szT1 differed markedly in their effect on recombination adjacent to the crossover suppressed region. hT1 had no effect on recombination in the adjacent interval. In contrast, the 0.8 map unit interval immediately adjacent to the szT1(I;X) breakpoint on chromosome I increased to 2.5 map units in translocation heterozygotes. This increase occurs in a chromosomal interval which can be expanded by treatment with radiation. These results are consistent with the suggestion that the szT1(I) breakpoint is in a region of DNA in which meiotic recombination is suppressed relative to the genomic average. We propose that DNA sequences disrupted by the szT1 translocation are responsible for determining the frequency of meiotic recombination in the vicinity of the breakpoint.
Assuntos
Caenorhabditis/genética , Translocação Genética , Animais , Caenorhabditis/citologia , Mapeamento Cromossômico , Troca Genética , Transtornos do Desenvolvimento Sexual , Genes Letais , Teste de Complementação Genética , Ligação Genética , Heterozigoto , Masculino , Meiose , Recombinação Genética , Cromossomo XRESUMO
The mei-218 gene product is required for both meiotic crossing over and for the production of recombination modules, suggesting that these organelles are required for meiotic exchange. In this study the null phenotype of mei-218 was defined through the analysis of three preexisting and five new alleles. Consistent with previous studies, in homozygous mei-218 mutants meiotic crossing over is reduced to < 10% of normal levels. A molecular analysis of mei-218 was initiated with the isolation and mapping of lethal mutations and genome rearrangements in the region containing mei-218, polytene interval 15E on the X chromosome. This high resolution genetic map was aligned with a physical map constructed from cosmid and P1 clones by genetically mapping restriction fragment length polymorphisms and localizing rearrangement breakpoints. Within a region of 65 kb, we have identified seven transcription units, including mei-218 and the Minute(1)15D gene, which encodes ribosomal protein S5. The mei-218 mutant phenotype has been rescued by germline transformation with both a genomic fragment and a cDNA under the control of the hsp83 promoter. The mei-218 gene is predicted to produce an 1186-amino acid protein that has no significant similarities to any known proteins.
Assuntos
Proteínas de Ciclo Celular/genética , Proteínas de Drosophila , Drosophila melanogaster/genética , Meiose , Recombinação Genética , Alelos , Sequência de Aminoácidos , Animais , Sequência de Bases , Mapeamento Cromossômico , Clonagem Molecular , DNA , DNA Complementar , Feminino , Deleção de Genes , Genoma , Humanos , Masculino , Dados de Sequência Molecular , Família Multigênica , Fenótipo , Homologia de Sequência de AminoácidosRESUMO
We have investigated the chromosomal region around unc-60 V, a gene affecting muscle structure, in the nematode Caenorhabditis elegans. The region studied covers 3 map units and lies at the left end of linkage group (LG) V. Compared to the region around dpy-11 (at the center of LGV), the unc-60 region has relatively few visible genes per map unit. We found the same to be true for essential genes. By screening simultaneously for recessive lethals closely linked to either dpy-11 or unc-60, we recovered ethyl methanesulfonate-induced mutations in 10 essential genes near dpy-11 but in only two genes near unc-60. Four deficiency breakpoints were mapped to the unc-60 region. Using recombination and deficiency mapping we established the following gene order: let-336, unc-34, let-326, unc-60, emb-29, let-426. Regarding unc-60 itself, we compared the effect of ten alleles (including five isolated during this study) on hermaphrodite mobility and fecundity. We used intragenic mapping to position eight of these alleles. The results show that these alleles are not distributed uniformly within the gene, but map to two groups approximately 0.012 map unit apart.
Assuntos
Caenorhabditis elegans/genética , Genes de Helmintos , Alelos , Animais , Mapeamento Cromossômico , Genes Letais , Genes Recessivos , Supressão GenéticaRESUMO
We present the cloning and characterization of mei-P26, a novel P-element-induced exchange-defective female meiotic mutant in Drosophila melanogaster. Meiotic exchange in females homozygous for mei-P26(1) is reduced in a polar fashion, such that distal chromosomal regions are the most severely affected. Additional alleles generated by duplication of the P element reveal that mei-P26 is also necessary for germline differentiation in both females and males. To further assess the role of mei-P26 in germline differentiation, we tested double mutant combinations of mei-P26 and bag-of-marbles (bam), a gene necessary for the control of germline differentiation and proliferation in both sexes. A null mutation at the bam locus was found to act as a dominant enhancer of mei-P26 in both males and females. Interestingly, meiotic exchange in mei-P26(1); bam(Delta)(86)/+ females is also severely decreased in comparison to mei-P26(1) homozygotes, indicating that bam affects the meiotic phenotype as well. These data suggest that the pathways controlling germline differentiation and meiotic exchange are related and that factors involved in the mitotic divisions of the germline may regulate meiotic recombination.
Assuntos
Proteínas de Transporte/genética , Proteínas de Transporte/fisiologia , Proteínas de Drosophila , Drosophila/genética , Células Germinativas/citologia , Células Germinativas/fisiologia , Proteínas de Insetos/genética , Proteínas de Insetos/fisiologia , Meiose/genética , Alelos , Animais , Divisão Celular , Clonagem Molecular , Drosophila/citologia , Drosophila/fisiologia , Elementos Facilitadores Genéticos/genética , Feminino , Genes Dominantes , Infertilidade/genética , Masculino , Modelos Genéticos , Mutagênese , Não Disjunção Genética , Fenótipo , Plasmídeos/genética , Recombinação Genética , Transcrição Gênica , Transformação Genética , Cromossomo X/genética , Dedos de ZincoRESUMO
We have isolated two alleles of a previously unidentified meiotic recombination gene, mei-217. Genetic analysis of these mutants shows that mei-217 is a typical "precondition" gene. The phenotypes of the mutants are meiosis specific. The strongest allele has <10% of the normal level of crossing over, and the residual events are distributed abnormally. We have used double mutant analysis to position mei-217 in the meiotic recombination pathway. In general, mutations causing defects in the initiation of meiotic recombination are epistatic to mutations in mei-41 and spnB. These two mutations, however, are epistatic to mei-217, suggesting that recombination is initiated normally in mei-217 mutants. It is likely that mei-217 mutants are able to make Holliday junction intermediates but are defective in the production of crossovers. These phenotypes are most similar to mutants of the mei-218 gene. This is striking because mei-217 and mei-218 are part of the same transcription unit and are most likely produced from a dicistronic message.
Assuntos
Proteínas de Ciclo Celular/genética , Proteínas de Drosophila , Drosophila/genética , Meiose/genética , Recombinação Genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Troca Genética , DNA Complementar , Ligação Genética , Dados de Sequência Molecular , Fenótipo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transcrição Gênica , Cromossomo XRESUMO
Meiotic recombination and DNA repair are mediated by overlapping sets of genes. In the yeast Saccharomyces cerevisiae, many genes required to repair DNA double-strand breaks are also required for meiotic recombination. In contrast, mutations in genes required for nucleotide excision repair (NER) have no detectable effects on meiotic recombination in S. cerevisiae. The Drosophila melanogaster mei-9 gene is unique among known recombination genes in that it is required for both meiotic recombination and NER. We have analyzed the mei-9 gene at the molecular level and found that it encodes a homologue of the S. cerevisiae excision repair protein Rad1, the probable homologue of mammalian XPF/ERCC4. Hence, the predominant process of meiotic recombination in Drosophila proceeds through a pathway that is at least partially distinct from that of S. cerevisiae, in that it requires an NER protein. The biochemical properties of the Rad1 protein allow us to explain the observation that mei-9 mutants suppress reciprocal exchange without suppressing the frequency of gene conversion.
Assuntos
Reparo do DNA/genética , Proteínas de Ligação a DNA , Proteínas de Drosophila , Drosophila melanogaster/genética , Genes de Insetos , Proteínas Nucleares , Proteínas/genética , Recombinação Genética/genética , Saccharomyces cerevisiae/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Proteínas de Transporte/genética , Primers do DNA , Enzimas Reparadoras do DNA , Elementos de DNA Transponíveis , Drosophila melanogaster/efeitos dos fármacos , Endonucleases/genética , Proteínas Fúngicas/genética , Genes Fúngicos , Meiose , Metanossulfonato de Metila/farmacologia , Dados de Sequência Molecular , Mutagênese , Reação em Cadeia da Polimerase , Biossíntese de Proteínas , Proteínas/química , Receptores de Esteroides/metabolismo , Proteínas de Saccharomyces cerevisiae , Homologia de Sequência de AminoácidosRESUMO
The segregation of homologous chromosomes from one another is the essence of meiosis. In many organisms, accurate segregation is ensured by the formation of chiasmata resulting from crossing over. Drosophila melanogaster females use this type of recombination-based system, but they also have mechanisms for segregating achiasmate chromosomes with high fidelity. We describe a P-element mutagenesis and screen in a sensitized genetic background to detect mutations that impair meiotic chromosome pairing, recombination, or segregation. Our screen identified two new recombination-deficient mutations: mei-P22, which fully eliminates meiotic recombination, and mei-P26, which decreases meiotic exchange by 70% in a polar fashion. We also recovered an unusual allele of the ncd gene, whose wild-type product is required for proper structure and function of the meiotic spindle. However, the screen yielded primarily mutants specifically defective in the segregation of achiasmate chromosomes. Although most of these are alleles of previously undescribed genes, five were in the known genes alphaTubulin67C, CycE, push, and Trl. The five mutations in known genes produce novel phenotypes for those genes.
Assuntos
Elementos de DNA Transponíveis/genética , Drosophila melanogaster/genética , Genes de Insetos , Meiose/genética , Animais , Cromossomos/genética , DNA/genética , Feminino , Heterocromatina , Masculino , Metáfase , Mutação , Não Disjunção Genética , Fenótipo , Recombinação Genética , Projetos de Pesquisa , Cromossomo X/genéticaRESUMO
In Drosophila females, the majority of recombination events do not become crossovers and those that do occur are nonrandomly distributed. Furthermore, a group of Drosophila mutants specifically reduce crossing over, suggesting that crossovers depend on different gene products than noncrossovers. In mei-218 mutants, crossing over is reduced by approximately 90% while noncrossovers and the initiation of recombination remain unchanged. Importantly, the residual crossovers have a more random distribution than wild-type. It has been proposed that mei-218 has a role in establishing the crossover distribution by determining which recombination sites become crossovers. Surprisingly, a diverse group of genes, including those required for double strand break (DSB) formation or repair, have an effect on crossover distribution. Not all of these mutants, however, have a crossover-specific defect like mei-218 and it is not understood why some crossover-defective mutants alter the distribution of crossovers. Intragenic recombination experiments suggest that mei-218 is required for a molecular transition of the recombination intermediate late in the DSB repair pathway. We propose that the changes in crossover distribution in some crossover-defective mutants are a secondary consequence of the crossover reductions. This may be the activation of a regulatory system that ensures at least one crossover per chromosome, and which compensates for an absence of crossovers by attempting to generate them at random locations.
Assuntos
Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Troca Genética/genética , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Mutação/genética , Alelos , Animais , Troca Genética/efeitos da radiação , Dano ao DNA/genética , Reparo do DNA/genética , Drosophila melanogaster/metabolismo , Drosophila melanogaster/efeitos da radiação , Feminino , Genótipo , Masculino , Meiose/genética , Complexo Sinaptonêmico/metabolismo , Raios XRESUMO
Duplications in Caenorhabditis elegans spontaneously delete at frequencies ranging from 10(-4) to 10(-5). We have analyzed the structure and mitotic stability of 33 deleted duplications resulting from spontaneous breakage events. (i) Breakage usually occurred at a variety of sites; that is, there were no hot spots for breakage. An exception was the spontaneous breakage of the X chromosome into which hDp14 was inserted. These breaks were close to or at the site of the chromosome I insertion; therefore, the insertion created a type of fragile site. (ii) Spontaneous duplications often had complex structures. In some cases, their structures were most simply resolved by proposing that the progenitor duplication was a ring chromosome with a superimposed inversion. Most of the proposed ring chromosomes were mitotically unstable, suggesting that ring structures increase the frequency of chromosome loss. (iii) Clusters of spontaneous deletion events were rarely observed, suggesting that the majority of spontaneous breakage events probably occurred during meiosis. (iv) A minority of the spontaneous breakage events were associated with linkage to an autosome. Like free duplications of chromosome I, these linked duplications tended to segregate from the X chromosome in males. (v) Three meiotic mutants, him-3, him-6, and him-8, had no effect on somatic loss of the duplications but did reduce the frequency of breakage events. Given the conclusion that chromosome breakage is a meiotic event, these data are consistent with the function of the three meiotic genes being restricted to meiosis.
Assuntos
Caenorhabditis elegans/genética , Animais , Deleção Cromossômica , Feminino , Genes de Helmintos , Ligação Genética , Marcadores Genéticos , Masculino , Meiose/genética , Mitose/genética , Mosaicismo , Família Multigênica , Mutação , Cromossomos em Anel , Cromossomo XRESUMO
Meiotic recombination requires the action of several gene products in both Saccharomyces cerevisiae and Drosophila melanogaster. Genetic studies in D. melanogaster have shown that the mei-W68 gene is required for all meiotic gene conversion and crossing-over. We cloned mei-W68 using a new genetic mapping method in which P elements are used to promote crossing-over at their insertion sites. This resulted in the high-resolution mapping of mei-W68 to a <18-kb region that contains a homolog of the S. cerevisiae spo11 gene. Molecular analysis of several mutants confirmed that mei-W68 encodes an spo11 homolog. Spo11 and MEI-W68 are members of a family of proteins similar to a novel type II topoisomerase. On the basis of this and other lines of evidence, Spo11 has been proposed to be the enzymatic activity that creates the double-strand breaks needed to initiate meiotic recombination. This raises the possibility that recombination in Drosophila is also initiated by double-strand breaks. Although these homologous genes are required absolutely for recombination in both species, their roles differ in other respects. In contrast to spo11, mei-W68 is not required for synaptonemal complex formation and does have a mitotic role.
Assuntos
Drosophila melanogaster/genética , Genes de Insetos , Meiose/genética , Recombinação Genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Mapeamento Cromossômico , Clonagem Molecular , Cruzamentos Genéticos , Troca Genética , Primers do DNA/genética , DNA Topoisomerases Tipo I/genética , DNA Complementar/genética , Feminino , Proteínas de Insetos/genética , Masculino , Dados de Sequência Molecular , Mutação , Reação em Cadeia da Polimerase , Saccharomyces cerevisiae/genética , Especificidade da Espécie , Complexo Sinaptonêmico/genéticaRESUMO
Essential genes have been identified in the 1.5 map unit (m.u.) dpy-14-unc-29 region of chromosome 1 in Caenorhabditis elegans. Previous work defined nine genes with visible mutant phenotypes and nine genes with lethal mutant phenotypes. In this study, we have identified an additional 28 essential genes with 97 lethal mutations. The mutations were mapped using eleven duplication breakpoints, eight deficiencies and three-factor recombination experiments. Genes required for the early stages of development were common, with 24 of the 37 essential genes having mutant phenotypes arresting at an early larval stage. Most mutants of a gene have the same time of arrest; only four of the 20 essential genes with multiple alleles have alleles with different phenotypes. From the analysis of complementing alleles of let-389, alleles with the same time-of-arrest phenotype were classified as either hypomorphic or amorphic. Mutants of let-605, let-534 and unc-37 have both uncoordinated and lethal phenotypes, suggesting that these genes are required for the coordination of movement and for viability. The physical and genetic maps in the dpy-14 region were linked by positioning two N2/BO polymorphisms with respect to duplications in the region, and by localizing the right breakpoint of the deficiency hDf8 on the physical map. Using cross-species hybridization to C. briggsae, ten regions of homology have been identified, eight of which are known to be coding regions, based on Northern analysis and/or the isolation of cDNA clones.
Assuntos
Caenorhabditis/genética , Alelos , Animais , Southern Blotting , Mapeamento Cromossômico , Cromossomos , Cosmídeos , Genes Letais , Teste de Complementação Genética , Família Multigênica , Mutação , Fenótipo , Polimorfismo Genético , Mapeamento por RestriçãoRESUMO
A screen was conducted for lethal mutations in the nematode Caenorhabditis elegans in a strain containing the mutator mut-4 (st700)I to examine the nature of mutator-induced lethal mutations within two large chromosomal regions comprising a total of 49 map units (linkage group IV (right) and linkage group V (left)). The genetic analysis of 28 lethal mutations has revealed that the mutator locus mut-4(st700)I causes both putative single-gene mutations and deficiencies. We have identified lethal mutations in three different genes, in addition to seven deficiencies. There is a mutational hot spot on linkage group V (left) around the lin-40 locus. Six mutations appear to be alleles of lin-40. In addition, 5 of 7 deficiencies have breakpoints at or very near lin-40. All seven deficiencies delete the left-most known gene on linkage group V (left) and thus appear to delete the tip of the chromosome. This is in contrast to gamma ray and formaldehyde induced deficiencies, which infrequently delete the closest known gene to the tip of a chromosome.