Josef Loidl

Disjunction of Homologous Chromosomes in Meiosis I Depends on Proteolytic Cleavage of the Meiotic Cohesin Rec8 by Separin

It has been proposed but never proven that cohesion between sister chromatids distal to chiasmata is responsible for holding homologous chromosomes together while spindles attempt to pull them toward opposite poles during metaphase of meiosis I. Meanwhile, the mechanism by which disjunction of homologs is triggered at the onset of anaphase I has remained a complete mystery. In yeast, cohesion between sister chromatid arms during meiosis depends on a meiosis-specific cohesin subunit called Rec8, whose mitotic equivalent, Sccl, is cleaved at the metaphase to anaphase transition by an endopeptidase called separin. We show here that cleavage of Rec8 by separin at one of two different sites is necessary for the resolution of chiasmata and the disjunction of homologous chromosomes during meiosis.

Cohesin ensures bipolar attachment of microtubules to sister centromeres and resists their precocious separation

The multisubunit protein complex cohesin is required to establish cohesion between sister chromatids during S phase and to maintain it during G2 and M phases. Cohesin is essential for mitosis, and even partial defects cause very high rates of chromosome loss. In budding yeast, cohesin associates with specific sites which are distributed along the entire length of a chromosome but are more dense in the vicinity of the centromere. Real-time imaging of individual centromeres tagged with green fluorescent protein suggests that cohesin bound to centromeres is important for bipolar attachment to microtubules. This cohesin is, however, incapable of resisting the consequent force, which leads to sister centromere splitting and chromosome stretching. Meanwhile, cohesin bound to sequences flanking the centromeres prevents sister chromatids from completely unzipping and is required to pull back together sister centromeres that have already split. Cohesin therefore has a central role in generating a dynamic tension between microtubules and sister chromatid cohesion at centromeres, which lasts until chromosome segregation is finally promoted by separin-dependent cleavage of the cohesin subunit Scc1p.

The Aurora B Kinase AIR-2 Regulates Kinetochores during Mitosis and Is Required for Separation of Homologous Chromosomes during Meiosis

BACKGROUND Mitotic chromosome segregation depends on bi-orientation and capture of sister kinetochores by microtubules emanating from opposite spindle poles and the near synchronous loss of sister chromatid cohesion. During meiosis I, in contrast, sister kinetochores orient to the same pole, and homologous kinetochores are captured by microtubules emanating from opposite spindle poles. Additionally, mechanisms exist that prevent complete loss of cohesion during meiosis I. These features ensure that homologs separate during meiosis I and sister chromatids remain together until meiosis II. The mechanisms responsible for orienting kinetochores in mitosis and for causing asynchronous loss of cohesion during meiosis are not well understood. RESULTS During mitosis in C. elegans, aurora B kinase, AIR-2, is not required for sister chromatid separation, but it is required for chromosome segregation. Condensin recruitment during metaphase requires AIR-2; however, condensin functions during prometaphase, independent of AIR-2. During metaphase, AIR-2 promotes chromosome congression to the metaphase plate, perhaps by inhibiting attachment of chromatids to both spindle poles. During meiosis in AIR-2-depleted oocytes, congression of bivalents appears normal, but segregation fails. Localization of AIR-2 on meiotic bivalents suggests this kinase promotes separation of homologs by promoting the loss of cohesion distal to the single chiasma. Inactivation of the phosphatase that antagonizes AIR-2 causes premature separation of chromatids during meiosis I, in a separase-dependent reaction. CONCLUSIONS Aurora B functions to resolve chiasmata during meiosis I and to regulate kinetochore function during mitosis. Condensin mediates chromosome condensation during prophase, and condensin-independent pathways contribute to chromosome condensation during metaphase.

A model for heterochromatin dispersion and the evolution of C-band patterns

Taking many observations from many species an attempt was made to find general regularities and rules governing the karyo-type position of C-bands in order to provide clues to the mechanisms of cytological and molecular evolution of patterns, A model is presented to explain the identified rules.

Genetic and cytological characterization of the recombination protein RAD-51 in Caenorhabditis elegans.

We investigated the role of Caenorhabditis elegans rad-51 during meiotic prophase. We showed that rad-51 mutant worms are viable, have no defects in meiotic homology recognition and synapsis but exhibit abnormal chromosomal morphology and univalent formation at diakinesis. During meiosis RAD-51 becomes localized to distinct foci in nuclei of the transition zone of the gonad and is most abundant in nuclei at late zygotene/early pachytene. Foci then gradually disappear from chromosomes and no foci are observed in late pachytene. RAD-51 localization requires the recombination genes spo-11 and mre-11 as well as chk-2, which is necessary for homology recognition and presynaptic alignment. Mutational analysis with synapsis- and recombination-defective strains, as well as the analysis of strains bearing heterozygous translocation chromosomes, suggests that presynaptic alignment may be required for RAD-51 focus formation, whereas homologous synaptonemal complex formation is required to remove RAD-51 foci.

Meiotic chromosome synapsis in a haploid yeast.

An extensive synaptonemal complex (SC) is found at pachytene in whole mount spread preparations of a haploid yeast, Saccharomyces cerevisiae, strain. Whereas unsynapsed axial elements are present only in a few nuclei, in others non-homologous synapsis involves virtually the whole chromosome set. This suggests that homology is not an indispensable precondition for SC formation in yeast but that chromosomes engage in non-homologous synapsis if no homologous partner is available. Recent evidence that in the sporulation deficient yeast mutants rad50 and mer1 axial elements do form but remain unsynapsed in the majority of nuclei is discussed in the light of the above findings.

Meiotic telomere clustering requires actin for its formation and cohesin for its resolution

In diploid organisms, meiosis reduces the chromosome number by half during the formation of haploid gametes. During meiotic prophase, telomeres transiently cluster at a limited sector of the nuclear envelope (bouquet stage) near the spindle pole body (SPB). Cohesin is a multisubunit complex that contributes to chromosome segregation in meiosis I and II divisions. In yeast meiosis, deficiency for Rec8 cohesin subunit induces telomere clustering to persist, whereas telomere cluster–SPB colocalization is defective. These defects are rescued by expressing the mitotic cohesin Scc1 in rec8Δ meiosis, whereas bouquet-stage exit is independent of Cdc5 pololike kinase. An analysis of living Saccharomyces cerevisiae meiocytes revealed highly mobile telomeres from leptotene up to pachytene, with telomeres experiencing an actin- but not microtubule-dependent constraint of mobility during the bouquet stage. Our results suggest that cohesin is required for exit from actin polymerization–dependent telomere clustering and for linking the SPB to the telomere cluster in synaptic meiosis.

Study of an RNA helicase implicates small RNA–noncoding RNA interactions in programmed DNA elimination in Tetrahymena

Tetrahymena eliminates micronuclear-limited sequences from the developing macronucleus during sexual reproduction. Homology between the sequences to be eliminated and approximately 28-nucleotide small RNAs (scnRNAs) associated with an Argonaute family protein Twi1p likely underlies this elimination process. However, the mechanism by which Twi1p-scnRNA complexes identify micronuclear-limited sequences is not well understood. We show that a Twi1p-associated putative RNA helicase Ema1p is required for the interaction between Twi1p and chromatin. This requirement explains the phenotypes of EMA1 KO strains, including loss of selective down-regulation of scnRNAs homologous to macronuclear-destined sequences, loss of H3K9 and K27 methylation in the developing new macronucleus, and failure to eliminate DNA. We further demonstrate that Twi1p interacts with noncoding transcripts derived from parental and developing macronuclei and this interaction is greatly reduced in the absence of Ema1p. We propose that Ema1p functions in DNA elimination by stimulating base-pairing interactions between scnRNAs and noncoding transcripts in both parental and developing new macronuclei.

Meiotic segregation, synapsis, and recombination checkpoint functions require physical interaction between the chromosomal proteins Red1p and Hop1p.

ABSTRACT In yeast, HOP1 and RED1 are required during meiosis for proper chromosome segregation and the consequent formation of viable spores. Mutations in either HOP1 orRED1 create unique as well as overlapping phenotypes, indicating that the two proteins act alone as well as in concert with each other. To understand which meiotic processes specifically require Red1p-Hop1p hetero-oligomers, a novel genetic screen was used to identify a single-point mutation of RED1,red1-K348E, that separates Hop1p binding from Red1p homo-oligomerization. The Red1-K348E protein is stable, phosphorylated in a manner equivalent to Red1p, and undergoes efficient homo-oligomerization; however, its ability to interact with Hop1p both by two-hybrid and coimmunoprecipitation assays is greatly reduced. Overexpression of HOP1 specifically suppressesred1-K348E, supporting the idea that the only defect in the protein is a reduced affinity for Hop1p. red1-K348E mutants exhibit reduced levels of crossing over and spore viability and fail to undergo chromosome synapsis, thereby implicating a role for Red1p-Hop1p hetero-oligomers in these processes. Furthermore,red1-K348E suppresses the sae2/com1 defects in meiotic progression and sporulation, indicating a previously unknown role for HOP1 in the meiotic recombination checkpoint.

Meiotic chromosome condensation and pairing in Saccharomyces cerevisiae studied by chromosome painting.

Non-isotopic high resolution in sity hybridization was applied to cytological preparations of sporulating yeast cells. Ribosomal DNA (rDNA) and chromosome V-specific recombinant lambda clones were used to tag individual chromosomes and chromosome subregions. This allowed the study of chromosome behaviour during early meiotic prophase. It was found that chromatin becomes condensed and homologous DNA sequences then appear to become aligned prior to synaptonemal complex formation.