Complex Formation of RNA Silencing Proteins in the Perinuclear Region of Neurospora crassa

In Neurospora, genes not paired during meiosis are targeted by meiotic silencing by unpaired DNA (MSUD). Here, our bimolecular fluorescence complementation (BiFC) study suggests that RNA-directed RNA polymerase, Dicer, Argonaute, and others form a silencing complex in the perinuclear region, with intimate interactions among the majority of them. We have also shown that SAD-2 is likely the anchor for this assembly.     

A Quality Control Mechanism Linking Meiotic Success to Release of Ascospores

Eukaryotic organisms employ a variety of mechanisms during meiosis to assess and ensure the quality of their gametes. Defects or delays in successful meiotic recombination activate conserved mechanisms to delay the meiotic divisions, but many multicellular eukaryotes also induce cell death programs to eliminate gametes deemed to have failed during meiosis. It is generally thought that yeasts lack such mechanisms. Here, we show that in the fission yeast Schizosaccharomyces pombe, defects in meiotic recombination lead to the activation of a checkpoint that is linked to ascus wall endolysis – the process by which spores are released in response to nutritional cues for subsequent germination. Defects in meiotic recombination are sensed as unrepaired DNA damage through the canonical ATM and ATR DNA damage response kinases, and this information is communicated to the machinery that stimulates ascus wall breakdown. Viability of spores that undergo endolysis spontaneously is significantly higher than that seen upon chemical endolysis, demonstrating that this checkpoint contributes to a selective mechanism for the germination of high quality progeny. These results provide the first evidence for the existence of a checkpoint linking germination to meiosis and suggest that analysis solely based on artificial, enzymatic endolysis bypasses an important quality control mechanism in this organism and potentially other ascomycota, which are models widely used to study meiosis.     

Meiotic silencing in the homothallic fungus Gibberella zeae

The homothallic ascomycete fungus Gibberella zeae is an important pathogen on major cereal crops. The objective of this study was to determine whether meiotic silencing occurs in G. zeae. Cytological studies demonstrated that GFP and RFP-fusion proteins were not detected during meiosis, both in heterozygous outcrosses and homozygous selfings. The deletion of rsp-1, a homologue used for studies on meiotic silencing of Neurospora crassa, triggered abnormal ascospores from selfing, but outcrosses between the mutant and wild-type strain resulted in some ascospores with mutant phenotype (low occurrence of ascus dominance). When the ectopic mutants that carried an additional copy of rsp-1 were selfed, they primarily produced ascospores with normal shape but a few ascospores (0.23 %) were abnormal, in which both endogenous and ectopically integrated genes contained numerous point mutations. The ectopic mutants showed low occurrence of ascus dominance in outcrosses with strains that carried the wild-type allele. Approximately 10 % of ascospores were abnormal but all of the single-ascospore isolates produced normal-shaped ascospores from selfing. However, no ascus dominance was observed when the mutants were outcrossed with a sad-1 deletion mutant, which lacks the putative RNA-dependent RNA polymerase essential for meiotic silencing in N. crassa. All results were consistent with those generated from an additional gene, roa, required for ascospore morphogenesis. This study demonstrated that G. zeae possesses a functional meiotic silencing mechanism which is triggered by unpaired DNA, as in N. crassa.     

Meiosis Drives Extraordinary Genome Plasticity in the Haploid Fungal Plant Pathogen Mycosphaerella graminicola

Meiosis in the haploid plant-pathogenic fungus Mycosphaerella graminicola results in eight ascospores due to a mitotic division following the two meiotic divisions. The transient diploid phase allows for recombination among homologous chromosomes. However, some chromosomes of M. graminicola lack homologs and do not pair during meiosis. Because these chromosomes are not present universally in the genome of the organism they can be considered to be dispensable. To analyze the meiotic transmission of unequal chromosome numbers, two segregating populations were generated by crossing genetically unrelated parent isolates originating from Algeria and The Netherlands that had pathogenicity towards durum or bread wheat, respectively. Detailed genetic analyses of these progenies using high-density mapping (1793 DArT, 258 AFLP and 25 SSR markers) and graphical genotyping revealed that M. graminicola has up to eight dispensable chromosomes, the highest number reported in filamentous fungi. These chromosomes vary from 0.39 to 0.77 Mb in size, and represent up to 38% of the chromosomal complement. Chromosome numbers among progeny isolates varied widely, with some progeny missing up to three chromosomes, while other strains were disomic for one or more chromosomes. Between 15–20% of the progeny isolates lacked one or more chromosomes that were present in both parents. The two high-density maps showed no recombination of dispensable chromosomes and hence, their meiotic processing may require distributive disjunction, a phenomenon that is rarely observed in fungi. The maps also enabled the identification of individual twin isolates from a single ascus that shared the same missing or doubled chromosomes indicating that the chromosomal polymorphisms were mitotically stable and originated from nondisjunction during the second division and, less frequently, during the first division of fungal meiosis. High genome plasticity could be among the strategies enabling this versatile pathogen to quickly overcome adverse biotic and abiotic conditions in wheat fields.     

Arabidopsis thaliana WAPL Is Essential for the Prophase Removal of Cohesin during Meiosis

Sister chromatid cohesion, which is mediated by the cohesin complex, is essential for the proper segregation of chromosomes in mitosis and meiosis. The establishment of stable sister chromatid cohesion occurs during DNA replication and involves acetylation of the complex by the acetyltransferase CTF7. In higher eukaryotes, the majority of cohesin complexes are removed from chromosomes during prophase. Studies in fly and human have shown that this process involves the WAPL mediated opening of the cohesin ring at the junction between the SMC3 ATPase domain and the N-terminal domain of cohesin''s α-kleisin subunit. We report here the isolation and detailed characterization of WAPL in Arabidopsis thaliana. We show that Arabidopsis contains two WAPL genes, which share overlapping functions. Plants in which both WAPL genes contain T-DNA insertions show relatively normal growth and development but exhibit a significant reduction in male and female fertility. The removal of cohesin from chromosomes during meiotic prophase is blocked in Atwapl mutants resulting in chromosome bridges, broken chromosomes and uneven chromosome segregation. In contrast, while subtle mitotic alterations are observed in some somatic cells, cohesin complexes appear to be removed normally. Finally, we show that mutations in AtWAPL suppress the lethality associated with inactivation of AtCTF7. Taken together our results demonstrate that WAPL plays a critical role in meiosis and raises the possibility that mechanisms involved in the prophase removal of cohesin may vary between mitosis and meiosis in plants.     

Preferential Occurrence of Nonsister Spores in Two-Spored Asci of SACCHAROMYCES CEREVISIAE: Evidence for Regulation of Spore-Wall Formation by the Spindle Pole Body

Yeast cells subjected to a reversible thermal arrest of meiosis yielded progressively fewer spores per ascus as the arrest was extended. Dissection of two-spored asci by a newly developed method that prevents selection of false asci revealed that the spores were not a random sample of the haploid meiotic products. Most, if not all, pairs of spores contain nonsister products of the reductional division. Electron microscopic examination of the meiotic cells revealed the cytological basis for this bias. All four spindle pole bodies (SPBs) present at the second meiotic division normally gain a structural modification (the outer plaque) upon which the initiation of the prospore wall occurs. In the formation of a two-spored ascus, only one spindle pole body on each meiosis II spindle was so modified. These observations suggest that the morphogenesis of spores is regulated at meiosis II by limiting the number of SPBs gaining the outer plaque. The enhancement of spore yield upon addition of fresh medium suggests that this morphogenetic regulation responds more directly to nutrient deprivation arising during the thermal arrest, rather than to elevated temperature per se.     

Meiotic self-pairing of the Psalidodon (Characiformes,Characidae) iso-B chromosome: A successful perpetuation mechanism

B chromosomes are non-essential additional genomic elements present in several animal and plant species. In fishes, species of the genus Psalidodon (Characiformes, Characidae) harbor great karyotype diversity, and multiple populations carry different types of non-essential B chromosomes. This study analyzed how the dispensable supernumerary B chromosome of Psalidodon paranae behaves during meiosis to overcome checkpoints and express its own meiosis-specific genes. We visualized the synaptonemal complexes of P. paranae individuals with zero, one, or two B chromosomes using immunodetection with anti-medaka SYCP3 antibody and fluorescence in situ hybridization with a (CA)₁₅ microsatellite probe. Our results showed that B chromosomes self-pair in cells containing only one B chromosome. In cells with two identical B chromosomes, these elements remain as separate synaptonemal complexes or close self-paired elements in the nucleus territory. Overall, we reveal that B chromosomes can escape meiotic silencing of unsynapsed chromatin through a self-pairing process, allowing expression of their own genes to facilitate regular meiosis resulting in fertile individuals. This behavior, also seen in other congeneric species, might be related to their maintenance throughout the evolutionary history of Psalidodon.     

Stage-Specific Expression Profiling of Drosophila Spermatogenesis Suggests that Meiotic Sex Chromosome Inactivation Drives Genomic Relocation of Testis-Expressed Genes

In Drosophila, genes expressed in males tend to accumulate on autosomes and are underrepresented on the X chromosome. In particular, genes expressed in testis have been observed to frequently relocate from the X chromosome to the autosomes. The inactivation of X-linked genes during male meiosis (i.e., meiotic sex chromosome inactivation—MSCI) was first proposed to explain male sterility caused by X-autosomal translocation in Drosophila, and more recently it was suggested that MSCI might provide the conditions under which selection would favor the accumulation of testis-expressed genes on autosomes. In order to investigate the impact of MSCI on Drosophila testis-expressed genes, we performed a global gene expression analysis of the three major phases of D. melanogaster spermatogenesis: mitosis, meiosis, and post-meiosis. First, we found evidence supporting the existence of MSCI by comparing the expression levels of X- and autosome-linked genes, finding the former to be significantly reduced in meiosis. Second, we observed that the paucity of X-linked testis-expressed genes was restricted to those genes highly expressed in meiosis. Third, we found that autosomal genes relocated through retroposition from the X chromosome were more often highly expressed in meiosis in contrast to their X-linked parents. These results suggest MSCI as a general mechanism affecting the evolution of some testis-expressed genes.     

Expression of Epitope-Tagged SYN3 Cohesin Proteins Can Disrupt Meiosis in Arabidopsis

a-kleisins are core components of meiotic and mitotic cohesin complexes. Arabidopsis contains genes encoding four a-kleisins. SYN1, a REC8 ortholog, is essential for meiosis, while SYN2 and SYN4 appear to be SCCI orthologs and function in mitosis. SYN3 is enriched in the nucleolus of meiotic and mitotic cells and is essential for megagametogenesis. It was recently shown that expression of SYN3-RNAi constructs in buds cause changes in meiotic gene expression that result in meiotic alterations. In this report we show that expression of SYN3 from the 35S promoter with either a c-terminal Myc or FAST tag causes a reduction in SYN1 mRNA levels that results in al- terations in sister chromatid cohesion, homologous chromosome synapsis female meiosis. and synaptonemal complex formation during both male and     

Fungal chromosomes

There are now four well-established methods to examine the chromosomes of filamentous fungi: mapping genes to linkage groups by recombination analyses, light-microscopic observation of chromosomes in meiotic divisions, electron-microscopic observation of the synaptonemal complexes between homologous chromosomes in prophase of meiosis, and separation of chromosomes as individual bands by pulsed field gel electrophoresis. These techniques and their contributions are described in brief with special reference toNeurospora. A fifth technique will be used more and more in characterizing chromosomes at the molecular level as DNA sequencing is completed for a limited number of the fungi. However, only the molecular studies of chromosome structures as they relate to centromeres, telomeres or nucleolus organizer regions are discussed, as is the potential usefulness of DNA sequencing to identify the junctions of chromosome rearrangements.     

The enigmatic meiotic dense body and its newly discovered component,SCML1, are dispensable for fertility and gametogenesis in mice

Meiosis is a critical phase in the life cycle of sexually reproducing organisms. Chromosome numbers are halved during meiosis, which requires meiosis-specific modification of chromosome behaviour. Furthermore, suppression of transposons is particularly important during meiosis to allow the transmission of undamaged genomic information between generations. Correspondingly, specialized genome defence mechanisms and nuclear structures characterize the germ line during meiosis. Survival of mammalian spermatocytes requires that the sex chromosomes form a distinct silenced chromatin domain, called the sex body. An enigmatic spherical DNA-negative structure, called the meiotic dense body, forms in association with the sex body. The dense body contains small non-coding RNAs including microRNAs and PIWI-associated RNAs. These observations gave rise to speculations that the dense body may be involved in sex body formation and or small non-coding RNA functions, e.g. the silencing of transposons. Nevertheless, the function of the dense body has remained mysterious because no protein essential for dense body formation has been reported yet. We discovered that the polycomb-related sex comb on midleg-like 1 (SCML1) is a meiosis-specific protein and is an essential component of the meiotic dense body. Despite abolished dense body formation, Scml1-deficient mice are fertile and proficient in sex body formation, transposon silencing and in timely progression through meiosis and gametogenesis. Thus, we conclude that dense body formation is not an essential component of the gametogenetic program in the mammalian germ line.