RESUMEN
In the early stages of meiosis, maternal and paternal chromosomes pair with their homologous partner and recombine to ensure exchange of genetic information and proper segregation. These events can vary drastically between species and between males and females of the same species. In Drosophila, in contrast to females, males do not form synaptonemal complexes (SCs), do not recombine, and have no crossing over; yet, males are able to segregate their chromosomes properly. Here, we investigated the early steps of homolog pairing in Drosophila males. We found that homolog centromeres are not paired in germline stem cells (GSCs) and become paired in the mitotic region before meiotic entry, similarly to females. Surprisingly, male germline cells express SC proteins, which localize to centromeres and promote pairing. We further found that the SUN/KASH (LINC) complex and microtubules are required for homolog pairing as in females. Chromosome movements in males, however, are much slower than in females and we demonstrate that this slow dynamic is compensated in males by having longer cell cycles. In agreement, slowing down cell cycles was sufficient to rescue pairing-defective mutants in female meiosis. Our results demonstrate that although meiosis differs significantly between males and females, sex-specific cell cycle kinetics integrate similar molecular mechanisms to achieve proper centromere pairing.
Asunto(s)
Emparejamiento Cromosómico , Drosophila , Animales , Masculino , Femenino , Emparejamiento Cromosómico/genética , Drosophila/genética , Complejo Sinaptonémico , Centrómero/genética , Meiosis/genética , Cromosomas , Segregación Cromosómica/genéticaRESUMEN
PURPOSE: In women under the age of 40, primary ovarian insufficiency (POI) is a devastating diagnosis with significant prevalence of 1-4% (Rajkovic and Pangas, Semin Reprod Med. 35(3):231-40, 2017). POI is characterized by amenorrhea with elevated levels of follicle stimulating hormone (FSH) and reduced estrogen levels, mimicking the menopausal state. Genetic determinants account for just over 10% of POI cases, yet determining whether particular single nucleotide polymorphisms (SNPs) are pathogenic is challenging. METHODS: We performed exome sequencing on a cohort of women with POI. CRISPR mutagenesis was employed to create a mutation in a conserved amino acid in the nematode protein. Functional relevance was assessed by analysis of bivalents and aberrant DNA morphologies in diakinesis nuclei. RESULTS: We identified a nonsynonymous c.C1051G; p.R351G variant, in a conserved region of the MSH5 protein. Mutation of this conserved amino acid in the C. elegans homolog, msh-5, revealed defective crossover outcomes in the homozygous and hemizygous states. CONCLUSIONS: These studies further implicate MSH5 as a POI gene and c.C1051G; p.R351G variant as likely playing a functional role in mammalian meiosis. This approach also highlights the ability of model organisms, such as C. elegans, to rapidly and inexpensively identify alleles of interest for further studies in mammalian models.
Asunto(s)
Insuficiencia Ovárica Primaria , Alelos , Aminoácidos , Animales , Caenorhabditis elegans/genética , Proteínas de Ciclo Celular/genética , Femenino , Humanos , Mamíferos/genética , Insuficiencia Ovárica Primaria/genética , Insuficiencia Ovárica Primaria/patología , Secuenciación del ExomaRESUMEN
The sporophyte generation of the brown alga Ectocarpus sp. exhibits an unusual pattern of development compared with the majority of brown algae. The first cell division is symmetrical and the apical-basal axis is established late in development. In the immediate upright (imm) mutant, the initial cell undergoes an asymmetric division to immediately establish the apical-basal axis. We provide evidence which suggests that this phenotype corresponds to the ancestral state of the sporophyte. The IMM gene encodes a protein of unknown function that contains a repeated motif also found in the EsV-1-7 gene of the Ectocarpus virus EsV-1. Brown algae possess large families of EsV-1-7 domain genes but these genes are rare in other stramenopiles, suggesting that the expansion of this family might have been linked with the emergence of multicellular complexity. EsV-1-7 domain genes have a patchy distribution across eukaryotic supergroups and occur in several viral genomes, suggesting possible horizontal transfer during eukaryote evolution.
Asunto(s)
Proteínas Algáceas/genética , Phaeophyceae/genética , Proteínas Algáceas/antagonistas & inhibidores , Proteínas Algáceas/química , Secuencia de Aminoácidos , Clonación Molecular , Cisteína/química , Evolución Molecular , Perfilación de la Expresión Génica , Transferencia de Gen Horizontal , Modelos Genéticos , Familia de Multigenes , Mutación , Phaeophyceae/crecimiento & desarrollo , Phaeophyceae/virología , Filogenia , Interferencia de ARN , Homología de Secuencia de Aminoácido , Proteínas Virales/química , Proteínas Virales/genéticaRESUMEN
Meiotic crossovers (COs) generate genetic diversity and are critical for the correct completion of meiosis in most species. Their occurrence is tightly constrained but the mechanisms underlying this limitation remain poorly understood. Here we identified the conserved AAA-ATPase FIDGETIN-LIKE-1 (FIGL1) as a negative regulator of meiotic CO formation. We show that Arabidopsis FIGL1 limits CO formation genome-wide, that FIGL1 controls dynamics of the two conserved recombinases DMC1 and RAD51 and that FIGL1 hinders the interaction between homologous chromosomes, suggesting that FIGL1 counteracts DMC1/RAD51-mediated inter-homologue strand invasion to limit CO formation. Further, depleting both FIGL1 and the previously identified anti-CO helicase FANCM synergistically increases crossover frequency. Additionally, we showed that the effect of mutating FANCM on recombination is much lower in F1 hybrids contrasting from the phenotype of inbred lines, while figl1 mutation equally increases crossovers in both contexts. This shows that the modes of action of FIGL1 and FANCM are differently affected by genomic contexts. We propose that FIGL1 and FANCM represent two successive barriers to CO formation, one limiting strand invasion, the other disassembling D-loops to promote SDSA, which when both lifted, leads to a large increase of crossovers, without impairing meiotic progression.
Asunto(s)
Adenosina Trifosfatasas/genética , Proteínas de Arabidopsis/genética , Arabidopsis/genética , Intercambio Genético/genética , ADN Helicasas/genética , Meiosis/genética , ATPasas Asociadas con Actividades Celulares Diversas , Proteínas de Ciclo Celular/genética , Reparación del ADN/genética , Variación Genética/genética , Proteínas Asociadas a Microtúbulos , Recombinasa Rad51/genética , Rec A Recombinasas/genética , Recombinación GenéticaRESUMEN
Meiotic crossovers (COs) have two important roles, shuffling genetic information and ensuring proper chromosome segregation. Despite their importance and a large excess of precursors (i.e., DNA double-strand breaks, DSBs), the number of COs is tightly regulated, typically one to three per chromosome pair. The mechanisms ensuring that most DSBs are repaired as non-COs and the evolutionary forces imposing this constraint are poorly understood. Here we identified Topoisomerase3α (TOP3α) and the RECQ4 helicases--the Arabidopsis slow growth suppressor 1 (Sgs1)/Bloom syndrome protein (BLM) homologs--as major barriers to meiotic CO formation. First, the characterization of a specific TOP3α mutant allele revealed that, in addition to its role in DNA repair, this topoisomerase antagonizes CO formation. Further, we found that RECQ4A and RECQ4B constitute the strongest meiotic anti-CO activity identified to date, their concomitant depletion leading to a sixfold increase in CO frequency. In both top3α and recq4ab mutants, DSB number is unaffected, and extra COs arise from a normally minor pathway. Finally, both TOP3α and RECQ4A/B act independently of the previously identified anti-CO Fanconi anemia of complementation group M (FANCM) helicase. This finding shows that several parallel pathways actively limit CO formation and suggests that the RECQA/B and FANCM helicases prevent COs by processing different substrates. Despite a ninefold increase in CO frequency, chromosome segregation was unaffected. This finding supports the idea that CO number is restricted not because of mechanical constraints but likely because of the long-term costs of recombination. Furthermore, this work demonstrates how manipulating a few genes holds great promise for increasing recombination frequency in plant-breeding programs.
Asunto(s)
Proteínas de Arabidopsis/genética , Intercambio Genético , ADN Helicasas/genética , ADN-Topoisomerasas de Tipo I/genética , Meiosis/genética , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/clasificación , Proteínas de Arabidopsis/metabolismo , Roturas del ADN de Doble Cadena , ADN Helicasas/clasificación , ADN Helicasas/metabolismo , ADN-Topoisomerasas de Tipo I/metabolismo , Mutación , Filogenia , Plantas Modificadas Genéticamente , Recombinación GenéticaRESUMEN
The recombining regions of sex chromosomes (pseudoautosomal regions, PARs) are predicted to exhibit unusual features due to their being genetically linked to the nonrecombining, sex-determining region. This phenomenon is expected to occur in both diploid (XY, ZW) and haploid (UV) sexual systems, with slightly different consequences for UV sexual systems because of the absence of masking during the haploid phase (when sex is expressed) and because there is no homozygous sex in these systems. Despite a considerable amount of theoretical work on PAR genetics and evolution, these genomic regions have remained poorly characterized empirically. We show here that although the PARs of the U/V sex chromosomes of the brown alga Ectocarpus recombine at a similar rate to autosomal regions of the genome, they exhibit many genomic features typical of nonrecombining regions. The PARs were enriched in clusters of genes that are preferentially, and often exclusively, expressed during the sporophyte generation of the life cycle, and many of these genes appear to have evolved since the Ectocarpales diverged from other brown algal lineages. A modeling-based approach was used to investigate possible evolutionary mechanisms underlying this enrichment in sporophyte-biased genes. Our results are consistent with the evolution of the PAR in haploid systems being influenced by differential selection pressures in males and females acting on alleles that are advantageous during the sporophyte generation of the life cycle.
Asunto(s)
Phaeophyceae/genética , Cromosomas Sexuales , Animales , Evolución Biológica , Evolución Molecular , Femenino , Sitios Genéticos , Haploidia , Masculino , Modelos Genéticos , Recombinación GenéticaRESUMEN
Germline cells produce gametes, which are specialized cells essential for sexual reproduction. Germline cells first amplify through several rounds of mitosis before switching to the meiotic program, which requires specific sets of proteins for DNA recombination, chromosome pairing, and segregation. Surprisingly, we previously found that some proteins of the synaptonemal complex, a prophase I meiotic structure, are already expressed and required in the mitotic region of Drosophila females. Here, to assess if additional meiotic genes were expressed earlier than expected, we isolated mitotic and meiotic cell populations to compare their RNA content. Our transcriptomic analysis reveals that all known meiosis I genes are already expressed in the mitotic region; however, only some of them are translated. As a case study, we focused on mei-W68, the Drosophila homolog of Spo11, to assess its expression at both the mRNA and protein levels and used different mutant alleles to assay for a premeiotic function. We could not detect any functional role for Mei-W68 during homologous chromosome pairing in dividing germ cells. Our study paves the way for further functional analysis of meiotic genes expressed in the mitotic region.
RESUMEN
Two distinct pathways for meiotic crossover formation coexist in most eukaryotes. The Arabidopsis SHOC1 protein is required for class I crossovers and shows sequence similarity with the XPF endonuclease family. Active XPF endonucleases form a heterodimer with ERCC1 proteins. Here, we show that PTD, an ERCC1-like protein, is required for class-I-interfering crossovers along with SHOC1, MSH4, MSH5, MER3 and MLH3. SHOC1 interacts with PTD in a two-hybrid assay, through its XPF-like nuclease-(HhH)(2) domain. We propose that a XPF-ERCC1-like heterodimer, represented by SHOC1 and PTD in Arabidopsis, involving Zip2 in Saccharomyces cerevisiae and C9orf84 in human, is required for formation of class I crossovers.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Intercambio Genético , Endodesoxirribonucleasas/metabolismo , Endonucleasas/metabolismo , Proteínas de Arabidopsis/genética , Proteínas Portadoras , Proteínas de Transporte de Catión , Intercambio Genético/genética , Proteínas de Unión al ADN/metabolismo , Dimerización , Endodesoxirribonucleasas/genética , Endonucleasas/genética , Epistasis Genética/genética , Evolución Molecular , Humanos , Meiosis , Mutación/genética , Filogenia , Unión Proteica , Proteínas , Saccharomyces cerevisiaeRESUMEN
Cytoplasmic linker-associated proteins (CLASPs) form a conserved family of microtubule-associated proteins (MAPs) that maintain microtubules in a growing state by promoting rescue while suppressing catastrophe.1 CLASP function involves an ordered array of tumor overexpressed gene (TOG) domains and binding to multiple protein partners via a conserved C-terminal domain (CTD).2,3 In migrating cells, CLASPs concentrate at the cortex near focal adhesions as part of cortical microtubule stabilization complexes (CMSCs), via binding of their CTD to the focal adhesion protein PHLDB2/LL5ß.4,5 Cortical CLASPs also stabilize a subset of microtubules, which stimulate focal adhesion turnover and generate a polarized microtubule network toward the leading edge of migrating cells. CLASPs are also recruited to the trans-Golgi network (TGN) via an interaction between their CTD and the Golgin protein GCC185.6 This allows microtubule growth toward the leading edge of migrating cells, which is required for Golgi organization, polarized intracellular transport, and cell motility.7 In dividing cells, CLASPs are essential at kinetochores for efficient chromosome segregation and anaphase spindle integrity.8,9 Both CENP-E and ASTRIN bind and target CLASPs to kinetochores,10,11 although the CLASP domain required for this interaction is not known. Despite its high evolutionary conservation, the CTD remains structurally uncharacterized. Here, we find that the CTD can be structurally modeled as a TOG domain. We identify a surface-exposed and conserved arginine residue essential for CLASP CTD interaction with partner proteins. Together, our results provide a structural mechanism by which the CLASP CTD directs diverse sub-cellular localizations throughout the cell cycle.
Asunto(s)
Proteínas Asociadas a Microtúbulos , Microtúbulos , Proteínas Asociadas a Microtúbulos/metabolismo , Microtúbulos/metabolismo , Movimiento Celular , Cinetocoros/metabolismo , Red trans-Golgi/metabolismoRESUMEN
During cell division, chromosome segregation is orchestrated by a microtubule-based spindle. Interaction between spindle microtubules and kinetochores is central to the bi-orientation of chromosomes. Initially dynamic to allow spindle assembly and kinetochore attachments, which is essential for chromosome alignment, microtubules are eventually stabilized for efficient segregation of sister chromatids and homologous chromosomes during mitosis and meiosis I, respectively. Therefore, the precise control of microtubule dynamics is of utmost importance during mitosis and meiosis. Here, we study the assembly and role of a kinetochore module, comprised of the kinase BUB-1, the two redundant CENP-F orthologs HCP-1/2, and the CLASP family member CLS-2 (hereafter termed the BHC module), in the control of microtubule dynamics in Caenorhabditis elegans oocytes. Using a combination of in vivo structure-function analyses of BHC components and in vitro microtubule-based assays, we show that BHC components stabilize microtubules, which is essential for meiotic spindle formation and accurate chromosome segregation. Overall, our results show that BUB-1 and HCP-1/2 do not only act as targeting components for CLS-2 at kinetochores, but also synergistically control kinetochore-microtubule dynamics by promoting microtubule pause. Together, our results suggest that BUB-1 and HCP-1/2 actively participate in the control of kinetochore-microtubule dynamics in the context of an intact BHC module to promote spindle assembly and accurate chromosome segregation in meiosis.
Asunto(s)
Proteínas de Caenorhabditis elegans , Huso Acromático , Animales , Huso Acromático/genética , Microtúbulos , Meiosis , Cinetocoros , Caenorhabditis elegans/genética , Segregación Cromosómica , Mitosis , Proteínas Asociadas a Microtúbulos/genética , Proteínas de Caenorhabditis elegans/genéticaRESUMEN
Crossovers (COs) are essential for the completion of meiosis in most species and lead to new allelic combinations in gametes. Two pathways of meiotic crossover formation have been distinguished. Class I COs, which are the major class of CO in budding yeast, mammals, Caenorhabditis elegans, and Arabidopsis, depend on a group of proteins called ZMM and rely on specific DNA structure intermediates that are processed to form COs. We identified a novel gene, SHOC1, involved in meiosis in Arabidopsis. Shoc1 mutants showed a striking reduction in the number of COs produced, a similar phenotype to the previously described Arabidopsis zmm mutants. The early steps of recombination, revealed by DMC1 foci, and completion of synapsis are not affected in shoc1 mutants. Double mutant analysis showed that SHOC1 acts in the same pathway as AtMSH5, a conserved member of the ZMM group. SHOC1 is thus a novel gene required for class I CO formation in Arabidopsis. Sequence similarity studies detected putative SHOC1 homologs in a large range of eukaryotes including human. SHOC1 appears to be related to the XPF endonuclease protein family, which suggests that it is directly involved in the maturation of DNA intermediates that lead to COs.
Asunto(s)
Proteínas de Arabidopsis/genética , Proteínas de Caenorhabditis elegans/genética , Proteínas Fúngicas/genética , Secuencia de Aminoácidos , Animales , Arabidopsis/citología , Arabidopsis/genética , Proteínas de Arabidopsis/química , Caenorhabditis elegans/citología , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/química , Secuencia Conservada , Intercambio Genético , ADN Complementario/genética , Fertilidad/genética , Proteínas Fúngicas/química , Humanos , Mamíferos , Meiosis , Datos de Secuencia Molecular , Saccharomycetales/citología , Saccharomycetales/genética , Alineación de SecuenciaRESUMEN
Meiosis is a key event in the manufacturing of an oocyte. During this process, the oocyte creates a set of unique chromosomes by recombining paternal and maternal copies of homologous chromosomes, and by eliminating one set of chromosomes to become haploid. While meiosis is conserved among sexually reproducing eukaryotes, there is a bewildering diversity of strategies among species, and sometimes within sexes of the same species, to achieve proper segregation of chromosomes. Here, we review the very first steps of meiosis in females, when the maternal and paternal copies of each homologous chromosomes have to move, find each other and pair. We explore the similarities and differences observed in C. elegans, Drosophila, zebrafish and mouse females.
Asunto(s)
Cromosomas/metabolismo , Meiosis/inmunología , Animales , Caenorhabditis elegansRESUMEN
Three amino acid loop extension homeodomain transcription factors (TALE HD TFs) act as life cycle regulators in green algae and land plants. In mosses these regulators are required for the deployment of the sporophyte developmental program. We demonstrate that mutations in either of two TALE HD TF genes, OUROBOROS or SAMSARA, in the brown alga Ectocarpus result in conversion of the sporophyte generation into a gametophyte. The OUROBOROS and SAMSARA proteins heterodimerise in a similar manner to TALE HD TF life cycle regulators in the green lineage. These observations demonstrate that TALE-HD-TF-based life cycle regulation systems have an extremely ancient origin, and that these systems have been independently recruited to regulate sporophyte developmental programs in at least two different complex multicellular eukaryotic supergroups, Archaeplastida and Chromalveolata.
Asunto(s)
Embryophyta/crecimiento & desarrollo , Embryophyta/metabolismo , Proteínas de Homeodominio/metabolismo , Phaeophyceae/crecimiento & desarrollo , Phaeophyceae/metabolismo , Proteínas de Plantas/metabolismo , Secuencia de Aminoácidos , Embryophyta/genética , Evolución Molecular , Regulación de la Expresión Génica de las Plantas , Proteínas de Homeodominio/química , Proteínas de Homeodominio/genética , Mutación/genética , Phaeophyceae/genética , Fenotipo , Unión Proteica , Dominios Proteicos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factores de Transcripción/química , Factores de Transcripción/genéticaRESUMEN
Double-strand breaks (DSBs) are among the most deleterious lesions DNA can endure. Yet, DSBs are programmed at the onset of meiosis, and are required to facilitate appropriate reduction of ploidy in daughter cells. Repair of these breaks is tightly controlled to favor homologous recombination (HR)-the only repair pathway that can form crossovers. However, little is known about how the activities of alternative repair pathways are regulated at these stages. We discovered an unexpected synthetic interaction between the DSB machinery and strand-exchange proteins. Depleting the Caenorhabditis elegans DSB-promoting factors HIM-5 and DSB-2 suppresses the formation of chromosome fusions that arise in the absence of RAD-51 or other strand-exchange mediators. Our investigations reveal that nonhomologous and theta-mediated end joining (c-NHEJ and TMEJ, respectively) and single strand annealing (SSA) function redundantly to repair DSBs when HR is compromised, and that HIM-5 influences the utilization of TMEJ and SSA.
Asunto(s)
Proteínas de Caenorhabditis elegans/metabolismo , Roturas del ADN de Doble Cadena , Reparación del ADN/genética , Meiosis/genética , Animales , Caenorhabditis elegans/citología , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Recombinación Homóloga , MutaciónRESUMEN
Meiosis is the cell division that reshuffles genetic information between generations. Recently, much progress has been made in understanding this process; in particular, the identification and functional analysis of more than 80 plant genes involved in meiosis have dramatically deepened our knowledge of this peculiar cell division. In this review, we provide an overview of advancements in the understanding of all aspects of plant meiosis, including recombination, chromosome synapsis, cell cycle control, chromosome distribution, and the challenge of polyploidy.
Asunto(s)
Genes de Plantas , Meiosis , Plantas/genética , Recombinación Genética , Ciclo Celular , Emparejamiento Cromosómico , Cromosomas de las Plantas , PoliploidíaRESUMEN
The life cycle of an organism is one of its fundamental features, influencing many aspects of its biology. The brown algae exhibit a diverse range of life cycles indicating that transitions between life cycle types may have been key adaptive events in the evolution of this group. Life cycle mutants, identified in the model organism Ectocarpus, are providing information about how life cycle progression is regulated at the molecular level in brown algae. We explore some of the implications of the phenotypes of the life cycle mutants described to date and draw comparisons with recent insights into life cycle regulation in the green lineage. Given the importance of coordinating growth and development with life cycle progression, we suggest that the co-option of ancient life cycle regulators to control key developmental events may be a common feature in diverse groups of multicellular eukaryotes.