Genetic Analysis of Inheritance ofmei8, sy1, and sy10Meiotic Mutations in Rye Secale cereale L.

It is shown that mutations mei8 (irregular condensation and fragmentation of meiotic chromosomes),sy1(asynapsis), and sy10 (heterologous synapsis) of rye Secale cereale L. are nonallelic. In double mutants mei8 sy1 and mei8 sy10 both mutations are expressed simultaneously and independently of each other. A study of joint inheritance of mutations sy1 and sy10 revealed their interaction by means of recessive epistasis: the double mutants sy1sy10 has the sy10 phenotype. This means that the sy10 gene controls an earlier stage of synapsis in meiotic prophase than the sy1 gene. Mutation mei8is inherited independently of sy1 but it is linked to sy10 (recombination frequency 26.8 ± 3.58%).     

The study of joint inheritance of mutations impairing the structure of meiotic chromosomes in rye Secale cereale L

Genetic analysis has demonstrated that meiotic mutations mei8 (irregular condensation and fragmentation of meiotic chromosomes) and mei10 (chromosome overcompaction) are nonallelic. Mutation mei10 exhibits digenic inheritance (with a segregation ratio of 13:3) in the combinations of crosses studied. It is assumed that the phenotypic expression of mutation mei10 is suppressed by the effect of recessive gene lch1 or lch2 (long chromosomes), both of which have been revealed in one of the parental lines (Mc10). These genes determine weak condensation of meiotic chromosomes. In double mutants mei8 mei10, the mutations are expressed independently of each other. Gene mei10 is linked with gene mei8 (r = 36.8 +/- 5.38%); genes lch1 and lch2 are not linked either with them or with each other. Taking into account the data on the linkage between genes mei10 and sy10 and between mei8 and sy10, the order of genes in the linkage group is shown to the following: mei8-sy10-mei10.     

Genetic analysis of mutation sy2 which causes nonhomologous meiotic synapsis in chromosomes of diploid rye Secale cereale L

Partially nonhomologous (heterologous) synapsis of meiotic chromosomes in a spontaneous desynaptic mutant form of rye is determined by two recessive genes, sy2a and sy2b, that have independent expression and inheritance. The third gene, dominant inhibitor suppressing the mutant phenotype, has been revealed in hybrid combinations between sy2 mutants and lines segregating other meiotic mutants: sy10 (heterologous synapsis), sy1, and sy9 (asynapsis). All three genes determining desynapsis (sy2a, sy2b, and I) were shown to be nonallelic to monogenic mutations sy10, sy1, and sy9, inherited independently of them and expressed at later stages of prophase I than the sy10 gene. The possibility of modifying monogenic segregation of mutation sy2 by gametophyte selection for a locus linked to the gene expressed as sy2 at particular frequencies of recombination between this gene and selected locus is discussed.     

Meiotic mutations in rye Secale cereale L

Spontaneous meiotic mutations of winter rye Secale cereale L. (2n = 14) were revealed in inbred F2 progenies, which were obtained by self-pollination of F1 hybrids resulting from crosses of individual plants of cultivar Vyatka or weedy rye with plants of self-fertile inbred lines. The mutations cause partial or complete sterility, and are maintained in heterozygote condition. Six types of mutations were distinguished as the result of cytological analysis of meiosis and genetic analysis. (1) Plants with nonallelic asynaptic mutations sy1 and sy9 lacked bivalents in 96.8 and 67.0% metaphase I cells, respectively, formed only axial elements but not the mature synaptonemal complex (SC), and had defects in telomere clustering in early prophase I. (2) Weak asynaptic mutant sy3 showed incomplete synapsis at the start of SC degradation at diplotene and lower chiasma number; yet only 2% meiocytes lacked bivalents in MI. (3) Mutations sy2, sy6, sy7, sy8, sy10, and sy19 caused nonhomologous synapsis; i.e., a varying number of univalents and occasional multivalents were observed in MI, which was preceded by switches of pairing partners and fold-back synapsis at mid-prophase I. (4) Mutation mei6 led to the formation of protrusions and minor branched structures of the SC lateral elements. (5) Allelic mutations mei8 and mei8-10 caused irregular chromatin condensation along the chromosome length in prophase I, which was accompanied by chromosome sticking and fragmentation in MI. (6) Allelic mutations mei5 and mei10 determined chromosome supercondensation, caused the disturbance of meiotic spindle assembly, arrested meiosis at various stages but did not affect formation of the pollen wall, thus arrested meiocytes got covered with the pollen wall. Analysis of double mutants revealed recessive epistatic interactions for some mutations; the epistatic group was sy9 > sy1 > sy3 > sy19. This reflects the sequence of meiotic events controlled by the corresponding genes. The expression of sy2 and sy19 proved to be modified by additional genes. Most meiotic mutations found in rye have analogs in other plants.     

Genetic collection of meiotic mutants of rye Secale cereale L

Genetic collection of meiotic mutants of winter rye Secale cereale L. (2n = 14) was created. Mutations were detected in inbred F2 generations after self-fertilization of the F1 hybrids, obtained by individual crossing of rye plants (cultivar Vyatka) or weedy rye with plants from autofertile lines. The mutations cause partial or complete plant sterility and are maintained in collection in a heterozygous state. Genetic analysis accompanied by cytogenetic study of meiosis has revealed six mutation types. (1) Nonallelic asynaptic mutations sy1 and sy9 caused the formation of only axial chromosome elements in prophase and anaphase. The synaptonemal complexes (SCs) were absent, the formation of the chromosome "bouquet" was impaired, and all chromosomes were univalent in meiotic metaphase I in 96% (sy1) and 67% (sy2) of cells. (2) Weak asynaptic mutation sy3, which hindered complete termination of synapsis in prophase II. Subterminal asynaptic segments were always observed in the SC, and at least one pair of univalents was present in metaphase I, but the number of cells with univalents did not exceed 2%. (3) Mutations sy2, sy6, sy7, sy8, sy10, and sy19, which caused partially nonhomologous synapsis: change in pairing partners and fold-back chromosome synapsis in prophase I. In metaphase I, the number of univalents varied and multivalents were observed. (4) Mutation mei6, which causes the formation of ultrastructural protrusions on the lateral SC elements, gaps and branching of these elements. (5) Allelic mutations mei8 and mei10, which caused irregular chromatin condensation along chromosomes in prophase I, sticking and fragmentation of chromosomes in metaphase I. (6) Allelic mutations mei5 and mei10, which caused chromosome hypercondensation, defects of the division spindle formation, and random arrest of cells at different meiotic stages. However, these mutations did not affect the formation of microspore envelopes even around the cells, whose development was blocked at prophase I. Analysis of cytological pictures of meiosis in double rye mutants reveled epistatic interaction in the mutation series sy9 > sy1 > sy3 > sy19, which reflects the order of switching these genes in the course of meiosis. The expression of genes sy2 and sy19 was shown to be controlled by modifier genes. Most meiotic mutations found in rye have analogs in other plant species.     

Genetic Analysis of Mutation sy2 which Causes Nonhomologous Meiotic Synapsis in Chromosomes of Diploid Rye Secale cereale L.

Partially nonhomologous (heterologous) synapsis of meiotic chromosomes in a spontaneous desynaptic mutant form of rye is determined by two recessive genes, sy2a and sy2b, that have independent expression and inheritance. The third gene, dominant inhibitor suppressing the mutant phenotype, has been revealed in hybrid combinations between sy2 mutants and lines segregating other meiotic mutants: sy10 (heterologous synapsis), sy1, and sy9(asynapsis). All three genes determining desynapsis (sy2a, sy2b, and I) were shown to be nonallelic to monogenic mutations sy10, sy1, and sy9, inherited independently of them and expressed at later stages of prophase I than the sy10 gene. The possibility of modifying monogenic segregation of mutation sy2 by gametophyte selection for a locus linked to the gene expressed as sy2 at particular frequencies of recombination between this gene and selected locus is discussed.     

Genetic control of chromosomal synapsis in Secale cereale L. rye meiosis: sy19 gene, causing heterologous synapsis

Analysis of manifestation and inheritance of a new mutation inducing irregular synapsis in rye showed that abnormal phenotype is determined by a recessive allele of the sy19 gene. In the homozygotes for this mutation, even at the light microscopic level, abnormal formation of bivalents is already observed at pachytene-diakinesis. At metaphase I, the univalent frequency varies from 0 to 14; in a few cells, multivalent associations of chromosomes, which are not clearly oriented in the spindle, are detected. Electron microscopy of synaptonemal complexes revealed both homologous and heterologous synapsis in homozygotes for sy19, namely partial loss of the ability to stringent homology search. Analysis of joint inheritance of sy19 and asynaptic sy1 mutations showed that they are nonallelic, inherited independently, and interact by recessive epistasis. The phenotype of double sy1sy19 mutants indicates that the sy19 gene conditioning heterologous synapsis operates at meiosis later than the synaptic gene sy1. The epistatic group of mutations, sy9 > sy1 > sy19 and sy3, was determined.     

The Study of Joint Inheritance of Mutations Impairing the Structure of Meiotic Chromosomes in Rye Secale cereale L.

Genetic analysis has demonstrated that meiotic mutations mei8 (irregular condensation and fragmentation of meiotic chromosomes) andmei10 (chromosome overcompaction) are nonallelic. Mutation mei10 exhibits digenic inheritance (with a segregation ratio of 13 : 3) in the combinations of crosses studied. It is assumed that the phenotypic expression of mutation mei10is suppressed by the effect of recessive genelch1 or lch2 (long chromosomes), both of which have been revealed in one of the parental lines (Mc10). These genes determine weak condensation of meiotic chromosomes. In double mutantsmei8 mei10, the mutations are expressed independently of each other. Gene mei10 is linked with gene mei8(r^ = 36.8 ± 5.38%); genes lch1 and lch2 are not linked either with them or with each other. Taking into account the data on the linkage between genes mei10and sy10 and between mei8andsy10, the order of genes in the linkage group is shown to be the following: mei8–sy10–mei10.     

Meiotic mutants of rye Secale cereale L. II. The nonhomologous synapsis in desynaptic mutants sy7 and sy10

We studied the expression and inheritance of two spontaneous mutations found in different populations of rye Secale cereale L. that cause high univalent frequency in meiosis and low fertility. Both mutations were inherited as monogenic recessives. For each of the mutations the corresponding gene symbols (sy7 and sy10) were suggested although their allelism has not been studied. These mutants differ in chiasma frequency and in the number of univalents per meiocyte. Electron microscopy of the wholemount surface-spread synaptonemal complexes (SCs) from microsporocytes of both mutants revealed that during meiotic prophase I random synapsis began and progressed that involved not only homologous but also nonhomologous chromosomes. SCs were formed with frequent changes of pairing partners (switches) and intrachromosomal foldbacks of unpaired axial elements. As a result, incompletely synapsed, non-homologous and multivalent SCs were formed in mutants by the stage analogous to pachytene in normal plants. In sy7 a maximum in the number of switches and foldbacks were observed at zygotene, whereas in sy10 this occurred at pachytene. We suggest that it is the process of recognition of homology that is impaired in both mutants. This leads to indiscriminate synapsis and prevents chiasma formation. Both mutants may be classified as desynaptic.     

Genetic Collection of Meiotic Mutants of Rye <Emphasis Type="Italic">Secale cereale</Emphasis> L.

Genetic collection of meiotic mutants of winter rye Secale cereale L. (2n = 14) was created. Mutations were detected in inbred F₂ generations after self-fertilization of the F₁ hybrids, obtained by individual crossing of rye plants (cultivar Vyatka) or weedy rye with plants from autofertile lines. The mutations cause partial or complete plant sterility and are maintained in collection in a heterozygous state. Genetic analysis accompanied by cytogenetic study of meiosis has revealed six mutation types. (1) Nonallelic asynaptic mutations sy1 and sy9 caused the formation of only axial chromosome elements in prophase and anaphase. The synaptonemal complexes (SCs) were absent, the formation of the chromosome “bouquet” was impaired, and all chromosomes were univalent in meiotic metaphase I in 96.8% (sy1) and 67% (sy2) of cells. (2) Weak asynaptic mutation sy3, which hindered complete termination of synapsis in prophase I. Subterminal asynaptic segments were always observed in the SC, and at least one pair of univalents was present in metaphase I, but the number of cells with 14 univalents did not exceed 2%. (3) Mutations sy2, sy6, sy7, sy8, sy10, and sy19, which caused partially nonhomologous synapsis: change in pairing partners and fold-back chromosome synapsis in prophase I. In metaphase I, the number of univalents varied and multivalents were observed. (4) Mutation mei6, which causes the formation of ultrastructural protrusions on the lateral SC elements, gaps and branching of these elements. (5) Allelic mutations mei8 and mei8-10, which caused irregular chromatin condensation along chromosomes in prophase I, sticking and fragmentation of chromosomes in metaphase I. (6) Allelic mutations mei5 and mei10, which caused chromosome hypercondensation, defects of the division spindle formation, and random arrest of cells at different meiotic stages. However, these mutations did not affect the formation of microspore envelopes even around the cells, whose development was blocked at prophase I. Analysis of cytological pictures of meiosis in double rye mutants reveled epistatic interaction in the mutation series sy9 > sy1 > sy3 > sy19, which reflects the order of switching these genes in the course of meiosis. The expression of genes sy2 and sy19 was shown to be controlled by modifier genes. Most meiotic mutations found in rye have analogs in other plant species.     

Positional cloning and characterization of mouse mei8, a disrupted allelle of the meiotic cohesin Rec8

A novel mutation, mei8, was isolated in a forward genetic screen for infertility mutations induced by chemical mutagenesis of ES cells. Homozygous mutant mice are sterile. Mutant females exhibit ovarian dysgenesis and lack ovarian follicles at reproductive maturity. Affected males have small testes due to arrest of spermatogenesis during meiotic prophase I. Genetic mapping and positional cloning of mei8 led to the identification of a mutation in Rec8, a homolog of the yeast meiosis-specific cohesin gene REC8. Analysis of meiosis in Rec8(mei8)/Rec8(mei8) spermatocytes showed that, while initiation of recombination and synapsis occurs, REC8 is required for the completion and/or maintenance of synapsis, cohesion of sister chromatids, and the formation of chiasmata, as it is in other organisms. However, unlike yeast and Caenorhabditis elegans, localization of REC8 on meiotic chromosomes is not required for the assembly of axial elements.     

Genetic Control of Chromosome Synapsis at Meiosis in Rye Secale cereale L.: The sy19 Gene Controlling Heterologous Synapsis

Analysis of manifestation and inheritance of a new mutation inducing irregular synapsis in rye showed that abnormal phenotype is determined by a recessive allele of the sy19 gene. In the homozygotes for this mutation, even at the light microscopic level, abnormal formation of bivalents is already observed at pachytene–diakinesis. At metaphase I, the univalent frequency varies from 0 to 14; in a few cells, multivalent associations of chromosomes, which are not clearly oriented in the spindle, are detected. Electron microscopy of synaptonemal complexes revealed both homologous and heterologous synapsis in homozygotes for sy19, namely partial loss of the ability to stringent homology search. Analysis of joint inheritance of sy19 and asynaptic sy1 mutations showed that they are nonallelic, inherited independently, and interact by recessive epistasis. The phenotype of doublesy1sy19 mutants indicates that thesy19 gene conditioning heterologous synapsis operates at meiosis later than the synaptic gene sy1. The epistatic group of mutations, sy9 > sy1 > sy19 and sy3, was determined.     

MRG-1 is required for genomic integrity in Caenorhabditis elegans germ cells

During meiotic cell division, proper chromosome synapsis and accurate repair of DNA double strand breaks (DSBs) are required to maintain genomic integrity, loss of which leads to apoptosis or meiotic defects. The mechanisms underlying meiotic chromosome synapsis, DSB repair and apoptosis are not fully understood. Here, we report that the chromodomain-containing protein MRG-1 is an important factor for genomic integrity in meiosis in Caenorhabditis elegans. Loss of mrg-1 function resulted in a significant increase in germ cell apoptosis that was partially inhibited by mutations affecting DNA damage checkpoint genes. Consistently, mrg-1 mutant germ lines exhibited SPO-11-generated DSBs and elevated exogenous DNA damage-induced chromosome fragmentation at diakinesis. In addition, the excessive apoptosis in mrg-1 mutants was partially suppressed by loss of the synapsis checkpoint gene pch-2, and a significant number of meiotic nuclei accumulated at the leptotene/zygotene stages with an elevated level of H3K9me2 on the chromatin, which was similarly observed in mutants deficient in the synaptonemal complex, suggesting that the proper progression of chromosome synapsis is likely impaired in the absence of mrg-1. Altogether, these findings suggest that MRG-1 is critical for genomic integrity by promoting meiotic DSB repair and synapsis progression in meiosis.     

The mouse Spo11 gene is required for meiotic chromosome synapsis

The Spo11 protein initiates meiotic recombination by generating DNA double-strand breaks (DSBs) and is required for meiotic synapsis in S. cerevisiae. Surprisingly, Spo11 homologs are dispensable for synapsis in C. elegans and Drosophila yet required for meiotic recombination. Disruption of mouse Spo11 results in infertility. Spermatocytes arrest prior to pachytene with little or no synapsis and undergo apoptosis. We did not detect Rad51/Dmc1 foci in meiotic chromosome spreads, indicating DSBs are not formed. Cisplatin-induced DSBs restored Rad51/Dmc1 foci and promoted synapsis. Spo11 localizes to discrete foci during leptotene and to homologously synapsed chromosomes. Other mouse mutants that arrest during meiotic prophase (Atm -/-, Dmc1 -/-, mei1, and Morc(-/-)) showed altered Spo11 protein localization and expression. We speculate that there is an additional role for Spo11, after it generates DSBs, in synapsis.     

Impairment of homologous chromosome synapsis in meiosis in rye <Emphasis Type="Italic">Secale cereale</Emphasis> L. Caused by a recessive mutation of the <Emphasis Type="Italic">sy18</Emphasis> gene

Expression and inheritance of the sy18 mutation causing impairment of synapsis homology were studied. It was established that the abnormal phenotype is determined by a recessive allele of the sy18 gene. Univalents and multivalents are observed in homozygotes for this mutant allele. According to the electron microscopic analysis of synaptonemal complexes in mutants, homologous synapsis occurs together with nonhomologous synapsis. The sy18 gene was found to have no allelism with asynaptic genes sy1 and sy9 and with genes sy10 and sy19 causing, like sy18, disturbances in synapsis homology.     

The Arabidopsis MEI1 gene encodes a protein with five BRCT domains that is involved in meiosis-specific DNA repair events independent of SPO11-induced DSBs

Arabidopsis thaliana MEI1 was first described as a gene involved in male meiosis, encoding a short protein showing homology with a human acrosin-trypsin inhibitor. We have isolated a new allele of mei1, and shown that in both mutants male and female meiosis are affected. In both reproductive pathways, meiosis proceeds while chromosomes become fragmented, resulting in aberrant meiotic products and in a strongly reduced fertility. We have shown that the gene mutated in mei1 mutants actually encodes a protein of 972 amino acids that contains five BRCA1 C-terminus (BRCT) domains and is similar to proteins involved in the response to DNA damage and replication blocks in eukaryotes. During meiosis, recombination is initiated by the formation of DNA double strand breaks (DSBs) induced by the protein SPO11. We analysed meiotic chromosome behaviour of the mei1 mutant in a spo11 mutant background and proved that the meiotic fragmentation observed in mei1 mutants was not the consequence of defects in the repair of meiotic DSBs induced by SPO11. We also analysed the effect of mei1 on the mitotic cell cycle but could not detect any sensitivity of mei1 seedlings to DNA-damaging agents like gamma-rays or UV. Therefore, MEI1 is a BRCT-domain-containing protein that could be specific to the meiotic cell cycle and that plays a crucial role in some DNA repair events independent of SPO11 DSB recombination repair.     

Molecular assembly of meiotic proteins Asy1 and Zyp1 and pairing promiscuity in rye (Secale cereale L.) and its synaptic mutant sy10

Assembly of two orthologous proteins associated with meiotic chromosome axes in Arabidopsis thaliana (Asy1 and Zyp1) was studied immunologically at meiotic prophase of meiosis of wild-type rye (Secale cereale) and its synaptic mutant sy10, using antibodies derived from A. thaliana. The temporal and spatial expression of the two proteins were similar in wild-type rye, but with one notable difference. Unlike A. thaliana, in which foci of the transverse filament protein Zyp1 appear to linearize commensurately with synapsis, linear tracts of Asy1 and Zyp1 protein form independently at leptotene and early zygotene of rye and coalign into triple structures resembling synaptonemal complexes (SCs) only at later stages of synapsis. The sy10 mutant used in this study also forms spatially separate linear tracts of Asy1 and Zyp1 proteins at leptotene and early zygotene, and these coalign but do not form regular triple structures at midprophase. Electron microscopy of spread axial elements reveals extensive asynapsis with some exchanges of pairing partners. Indiscriminate SCs support nonhomologous chiasma formation at metaphase I, as revealed by multi-color fluorescence in situ hybridization enabling reliable identification of all the chromosomes of the complement. Scrutiny of chiasmate associations of chromosomes at this stage revealed some specificity in the associations of homologous and nonhomologous chromosomes. Inferences about the nature of synapsis in this mutant were drawn from such observations.     

Use of homeotic mutation tasselseed2 for investigation of the action of maize meiotic genes during micro- and megasporogenesis

The manifestation of the ms43 maize meiotic mutation in the megasporogenesis of ts2 ms43 double mutants has been studied. Combined genetical and cytological analysis of the progeny of diheterozygote selfing showed that the ms43 mei gene was not microsporogenesis-specific. The manifestation of ms43 in megasporogenesis of the double mutants proved to be affected by the ts2 mutation. It prevented formation of the phenotype characteristic of ms43 and distorted early developmental stages of the entire ovule. It is the first study of megasporogenesis in tasselseed2 mutant tassels. Cytological data on the afd1 mutation in single and double mutants are presented. Possible mechanisms of the interaction between the ms43 and ts2 mutations are discussed.     

The Role of the Ameiotic1 Gene in the Initiation of Meiosis and in Subsequent Meiotic Events in Maize

Understanding the initiation of meiosis and the relationship of this event with other key cytogenetic processes are major goals in studying the genetic control of meiosis in higher plants. Our genetic and structural analysis of two mutant alleles of the ameiotic1 gene (am1 and am1-praI) suggest that this locus plays an essential role in the initiation of meiosis in maize. The product of the ameiotic1 gene affects an earlier stage in the meiotic sequence than any other known gene in maize and is important for the irreversible commitment of cells to meiosis and for crucial events marking the passage from premeiotic interphase into prophase I including chromosome synapsis. It appears that the period of ameiotic1 gene function in meiosis at a minimum covers the interval from some point during premeiotic interphase until the early zygotene stage of meiosis. To study the interaction of genes in the progression of meiosis, several double meiotic mutants were constructed. In these double mutants (i) the ameiotic1 mutant allele was brought together with the meiotic mutation (afd1) responsible for the fixation of centromeres in meiosis; and with the mutant alleles of the three meiotic genes that control homologous chromosome segregation (dv1, ms43 and ms28), which impair microtubule organizing center organization, the orientation of the spindle fiber apparatus, and the depolymerization of spindle filaments after the first meiotic division, respectively; (ii) the afd1 mutation was combined with two mutations (dsy1 and as1) affecting homologous pairing; (iii) the ms43 mutation was combined with the as1, the ms28 and the dv1 mutations; and (iv) the ms28 mutation was combined with the dv1 mutation and the ms4 (polymitotic1) mutations. An analysis of gene interaction in the double mutants led us to conclude that the ameiotic1 gene is epistatic over the afd1, the dv1, the ms43 and the ms28 genes but the significance of this relationship requires further analysis. The afd gene appears to function from premeiotic interphase throughout the first meiotic division, but it is likely that its function begins after the start of the ameiotic1 gene expression. The afd1 gene is epistatic over the two synaptic mutations dsy1 and as1 and also over the dv1 mutation. The new ameiotic*-485 and leptotene arrest*-487 mutations isolated from an active ROBERTSON's Mutator stocks take part in the control of the initiation of meiosis.     

Mapping of meiotic genes in rye (Secale cereale L.): Localization of sy19 mutation, impairing homologous synapsis, by means of isozyme and microsatellite markers

The sy19 mutation, which impairs the homology of meiotic chromosome synapsis in rye, were mapped using a specially created F₂ population by means of isozyme Acph1 locus and microsatellite (SSR) markers. The sy19 gene was localized in the chromosome 7R in the pericentromeric region of long arm based on the linked inheritance with the Acph1 locus. The locus was linked with five rye SSR markers, with the Xrems1234 locus being located closest to the sy19 gene (6.4 cM). The genetic map of the analyzed chromosome 7R region includes ten markers and the sy19 locus. A possible function of the Sy1 and Sy19 genes based on the data on comparative genomics is discussed.