The multiple roles of mps1 in Drosophila female meiosis.
Ontology highlight
ABSTRACT:
SUBMITTER: Gilliland WD
PROVIDER: S-EPMC1914070 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:The physical connections established by recombination are normally sufficient to ensure proper chromosome segregation during female Meiosis I. However, nonexchange chromosomes (such as the Muller F element or "dot" chromosome in D. melanogaster) can still segregate accurately because they remain connected by heterochromatic tethers. A recent study examined female meiosis in the closely related species D. melanogaster and D. simulans, and found a nearly twofold difference in the mean distance the obligately nonexchange dot chromosomes were separated during Prometaphase. That study proposed two speculative hypotheses for this difference, the first being the amount of heterochromatin in each species, and the second being the species' differing tolerance for common inversions in natural populations. We tested these hypotheses by examining female meiosis in 12 additional Drosophila species. While neither hypothesis had significant support, we did see 10-fold variation in dot chromosome sizes, and fivefold variation in the frequency of chromosomes out on the spindle, which were both significantly correlated with chromosome separation distances. In addition to demonstrating that heterochromatin abundance changes chromosome behavior, this implies that the duration of Prometaphase chromosome movements must be proportional to the size of the F element in these species. Additionally, we examined D. willistoni, a species that lacks a free dot chromosome. We observed that chromosomes still moved out on the meiotic spindle, and the F element was always positioned closest to the spindle poles. This result is consistent with models where one role of the dot chromosomes is to help organize the meiotic spindle.
| S-EPMC7202005 | biostudies-literature
Project description:Genetic screens for recessive alleles induce mutations, make the mutated chromosomes homozygous, and then assay those homozygotes for the phenotype of interest. When screening for genes required for female meiosis, the phenotype of interest has typically been nondisjunction from chromosome segregation errors. As this requires that mutant females be viable and fertile, any mutants that are lethal or sterile when homozygous cannot be recovered by this approach. To overcome these limitations, our lab has screened the VALIUM22 collection produced by the Harvard TRiP Project, which contains RNAi constructs targeting genes known to be expressed in the germline in a vector optimized for germline expression. By driving RNAi with GAL4 under control of a germline-specific promoter (nanos or mat-alpha4), we can test genes that would be lethal if knocked down in all cells, and by examining unfertilized metaphase-arrested mature oocytes, we can identify defects associated with genes whose knockdown results in sterility or causes other errors besides nondisjunction. We screened this collection to identify genes that disrupt either of two phenotypes when knocked down: the ability of meiotic chromosomes to congress to a single mass at the end of prometaphase, and the sequestration of Mps1-GFP to ooplasmic filaments in response to hypoxia. After screening >1450 lines of the collection, we obtained multiple hits for both phenotypes, identified novel meiotic phenotypes for genes that had been previously characterized in other processes, and identified the first phenotypes to be associated with several previously uncharacterized genes.
| S-EPMC10827134 | biostudies-literature
Project description:The model of Drosophila female meiosis I was recently revised by the discovery that chromosome congression precedes metaphase I arrest. Use of the prior framework to interpret data from meiotic mutants led to the conclusion that chromosome segregation errors (nondisjunction, NDJ) occurred when nonexchange chromosomes moved out on the spindle in a maloriented configuration and became trapped there at metaphase arrest. The discovery that congression returns nonexchange chromosomes to the metaphase plate invalidates this interpretation and raises the question of what events actually do lead to NDJ. To address this, we have assayed an allelic series of ald (mps1) meiotic mutants that complete congression at wild-type rates, but have widely varying NDJ rates in an otherwise isogenic background, as well as a nod mutant background that primarily undergoes loss of chromosome 4. Using genetic assays to measure NDJ rates, and FISH assays to measure chromosome malorientation rates in metaphase-arrested oocytes, shows that these two rates are highly correlated across ald mutants, suggesting that malorientation during congression commits these chromosomes to eventually nondisjoin. Likewise, the rate of chromosome loss observed in nod is similar to the rate at which these chromosomes fail to associate with the main chromosome mass. Together these results provide a proximal mechanism for how these meiotic mutants cause NDJ and chromosome loss and improve our understanding of how prometaphase chromosome congression relates to anaphase chromosome segregation.
| S-EPMC3567735 | biostudies-literature
Project description:Silencing of transposons in the Drosophila ovary relies on three Piwi family proteins--Piwi, Aubergine (Aub), and Ago3--acting in concert with their small RNA guides, the Piwi-interacting RNAs (piRNAs). Aub and Ago3 are found in the germ cell cytoplasm, where they function in the ping-pong cycle to consume transposon mRNAs. The nuclear Piwi protein is required for transposon silencing in both germ and somatic follicle cells, yet the precise mechanisms by which Piwi acts remain largely unclear. We investigated the role of Piwi by combining cell type-specific knockdowns with measurements of steady-state transposon mRNA levels, nascent RNA synthesis, chromatin state, and small RNA abundance. In somatic cells, Piwi loss led to concerted effects on nascent transcripts and transposon mRNAs, indicating that Piwi acts through transcriptional gene silencing (TGS). In germ cells, Piwi loss showed disproportionate impacts on steady-state RNA levels, indicating that it also exerts an effect on post-transcriptional gene silencing (PTGS). Piwi knockdown affected levels of germ cell piRNAs presumably bound to Aub and Ago3, perhaps explaining its post-transcriptional impacts. Overall, our results indicate that Piwi plays multiple roles in the piRNA pathway, in part enforcing transposon repression through effects on local chromatin states and transcription but also participating in germ cell piRNA biogenesis.
| S-EPMC3589557 | biostudies-literature
Project description:In this study we provide evidence that the spindle matrix protein Skeletor in Drosophila interacts with the human ASCIZ (also known as ATMIN and ZNF822) ortholog, Digitor/dASCIZ. This interaction was first detected in a yeast two-hybrid screen and subsequently confirmed by pull-down assays. We also confirm a previously documented function of Digitor/dASCIZ as a regulator of Dynein light chain/Cut up expression. Using transgenic expression of a mCitrine-labeled Digitor construct, we show that Digitor/dASCIZ is a nuclear protein that is localized to interband and developmental puff chromosomal regions during interphase but redistributes to the spindle region during mitosis. Its mitotic localization and physical interaction with Skeletor suggest the possibility that Digitor/dASCIZ plays a direct role in mitotic progression as a member of the spindle matrix complex. Furthermore, we have characterized a P-element insertion that is likely to be a true null Digitor/dASCIZ allele resulting in complete pupal lethality when homozygous, indicating that Digitor/dASCIZ is an essential gene. Phenotypic analysis of the mutant provided evidence that Digitor/dASCIZ plays critical roles in regulation of metamorphosis and organogenesis as well as in the DNA damage response. In the Digitor/dASCIZ null mutant larvae there was greatly elevated levels of ?H2Av, indicating accumulation of DNA double-strand breaks. Furthermore, reduced levels of Digitor/dASCIZ decreased the resistance to paraquat-induced oxidative stress resulting in increased mortality in a stress test paradigm. We show that an early developmental consequence of the absence of Digitor/dASCIZ is reduced third instar larval brain size although overall larval development appeared otherwise normal at this stage. While Digitor/dASCIZ mutant larvae initiate pupation, all mutant pupae failed to eclose and exhibited various defects in metamorphosis such as impaired differentiation, incomplete disc eversion, and faulty apoptosis. Altogether we provide evidence that Digitor/dASCIZ is a nuclear protein that performs multiple roles in Drosophila larval and pupal development.
| S-EPMC5115829 | biostudies-literature
Project description:Spindle formation in female meiosis differs from mitosis in many animals, as it takes place independently of centrosomes, and the molecular requirements of this pathway remain to be understood. Here, we report two crucial roles of Incenp, an essential subunit of the chromosomal passenger complex (the Aurora B complex), in centrosome-independent spindle formation in Drosophila female meiosis. First, the initial assembly of spindle microtubules is drastically delayed in an incenp mutant. This clearly demonstrates, for the first time, a crucial role for Incenp in chromosome-driven spindle microtubule assembly in living oocytes. Additionally, Incenp is necessary to stabilise the equatorial region of the metaphase I spindle, in contrast to mitosis, where the equivalent function becomes prominent after anaphase onset. Our analysis suggests that Subito, a kinesin-6 protein, cooperates with Incenp for this latter function, but not in microtubule assembly. We propose that the two functions of Incenp are part of the mechanisms that compensate for the lack of centrosomes during meiotic spindle formation.
| S-EPMC2536564 | biostudies-literature
Project description:During the extended prophase of Drosophila gametogenesis, spermatocytes undergo robust gene transcription and store many transcripts in the cytoplasm in a repressed state, until translational activation of select mRNAs in later steps of spermatogenesis. Here, we characterize the Drosophila Doublefault (Dbf) protein as a C2H2 zinc-finger protein, primarily expressed in testes, that is required for normal meiotic division and spermiogenesis. Loss of Dbf causes premature centriole disengagement and affects spindle structure, chromosome segregation and cytokinesis. We show that Dbf interacts with the RNA-binding protein Syncrip/hnRNPQ, a key regulator of localized translation in Drosophila We propose that the pleiotropic effects of dbf loss-of-function mutants are associated with the requirement of dbf function for translation of specific transcripts in spermatocytes. In agreement with this hypothesis, Dbf protein binds cyclin B mRNA and is essential for translation of cyclin B in mature spermatocytes.
| S-EPMC6899030 | biostudies-literature
Project description:Meiosis, like mitosis depends on the activity of the mitotic Cyclin dependent kinase, Cdk1 and its cyclin partners. Here we examine the specific requirements for the three mitotic cyclins, A, B and B3 in meiosis of Drosophila melanogaster We find that all three cyclins contribute redundantly to nuclear envelope breakdown, though Cyclin A appears to make the most important individual contribution. Cyclin A is also required for bi-orientation of homologues in meiosis I. Cyclin B3, as previously reported, is required for anaphase progression in meiosis I and in meiosis II. We find that it also plays a redundant role, with Cyclin A, in preventing DNA replication during meiosis. Cyclin B is required for maintenance of the metaphase I arrest in mature oocytes, for spindle organization and for timely progression through the 2(nd) meiotic division. It is also essential for polar body formation at the completion of meiosis. With the exception of its redundant role in meiotic maturation, Cyclin B appears to function independently of Cyclins A and B3 through most of meiosis. We conclude that the 3 mitotic Cyclin-Cdk complexes have distinct and overlapping functions in Drosophila female meiosis.
| S-EPMC5100870 | biostudies-other
Project description:Heterochromatic homology ensures the segregation of achiasmate chromosomes during meiosis I in Drosophila melanogaster females, perhaps as a consequence of the heterochromatic threads that connect achiasmate homologs during prometaphase I. Here, we ask how these threads, and other possible heterochromatic entanglements, are resolved prior to anaphase I. We show that the knockdown of Topoisomerase II (Top2) by RNAi in the later stages of meiosis results in a specific defect in the separation of heterochromatic regions after spindle assembly. In Top2 RNAi-expressing oocytes, heterochromatic regions of both achiasmate and chiasmate chromosomes often failed to separate during prometaphase I and metaphase I. Heterochromatic regions were stretched into long, abnormal projections with centromeres localizing near the tips of the projections in some oocytes. Despite these anomalies, we observed bipolar spindles in most Top2 RNAi-expressing oocytes, although the obligately achiasmate 4th chromosomes exhibited a near complete failure to move toward the spindle poles during prometaphase I. Both achiasmate and chiasmate chromosomes displayed defects in biorientation. Given that euchromatic regions separate much earlier in prophase, no defects were expected or observed in the ability of euchromatic regions to separate during late prophase upon knockdown of Top2 at mid-prophase. Finally, embryos from Top2 RNAi-expressing females frequently failed to initiate mitotic divisions. These data suggest both that Topoisomerase II is involved in the resolution of heterochromatic DNA entanglements during meiosis I and that these entanglements must be resolved in order to complete meiosis.
| S-EPMC4207608 | biostudies-literature
Project description:A key feature of meiosis is the step-wise removal of cohesin, the protein complex holding sister chromatids together, first from arms in meiosis I and then from the centromere region in meiosis II. Centromeric cohesin is protected by Sgo2 from Separase-mediated cleavage, in order to maintain sister chromatids together until their separation in meiosis II. Failures in step-wise cohesin removal result in aneuploid gametes, preventing the generation of healthy embryos. Here, we report that kinase activities of Bub1 and Mps1 are required for Sgo2 localisation to the centromere region. Mps1 inhibitor-treated oocytes are defective in centromeric cohesin protection, whereas oocytes devoid of Bub1 kinase activity, which cannot phosphorylate H2A at T121, are not perturbed in cohesin protection as long as Mps1 is functional. Mps1 and Bub1 kinase activities localise Sgo2 in meiosis I preferentially to the centromere and pericentromere respectively, indicating that Sgo2 at the centromere is required for protection.In meiosis I centromeric cohesin is protected by Sgo2 from Separase-mediated cleavage ensuring that sister chromatids are kept together until their separation in meiosis II. Here the authors demonstrate that Bub1 and Mps1 kinase activities are required for Sgo2 localisation to the centromere region.
| S-EPMC5612927 | biostudies-literature