meiotic segregation and recombination

Crossovers by themselves are insufficient to direct disjunction, however. Each haploid was subsequently transformed with pSH18-34Spe (provided by S. Hollenberg, Oregon Health Sciences University) digested with StuI to target integration of the plasmid to ura3. The SC fragments in red1-K348E may represent a leaky phenotype for the red1-K348E mutant in which local SC formation occurs because some Red1p-Hop1p hetero-oligomers are formed. Mutations in either HOP1 or RED1 create unique as well as overlapping phenotypes, indicating that the two proteins act alone as well as in concert with each other. In fact, there are mammalian AE components (COR1 and SCP3) that may serve similar roles as Red1p (12, 24). Simultaneous detection of structural and numerical chromosome abnormalities in sperm of healthy men by multicolor fluorescence in situ hybridization. The wild-type diploid produced 85.0% mature asci, compared to just 1.2% for the sae2 strain. The https:// ensures that you are connecting to the Would you like email updates of new search results? Chromosome spreads were prepared, Meiotic progression in various red1 sae2 SK1 diploids. This value is statistically significantly lower than the 50% expected if spore viability is directly correlated with crossing over (2; P < 0.001). (Fig.7).7). If the original parts of the same chromosome subsequently rejoin, no recombination occurs; however, if a strand of one chromosome joins together with a strand from the second chromosome, the pair of recombined chromosomes will differ in the combination of their alleles from the two original chromosomes. Please enable it to take advantage of the complete set of features! The improvement in the spore viability of red1-K348E diploids when HOP1 is overexpressed is presumably due to the ability to now form SCs. Some of these threads seem to be thickened AEs, but the majority are segments of tripartite SCs. This library was generated using primers complementary to sequences 200 bp upstream of the 5 BglII site and 200 bp downstream of the 3 BglII site in pNH223 with pNH223 as the template. [email protected]. Genetics. The truncation plasmids pDW35 (lexA-RED1326), pDW61 (lexA-RED11359), and pDW62 (lexA-RED11400) were all constructed by PCR using pNH207 as the template and Vent polymerase (New England Biolabs, Beverly, Mass.). Would you like email updates of new search results? Chromosome spreads were prepared for electron microscopy as described in Materials and Methods. Plasmids pRS402, pBB14, and pBB16 were digested with StuI and integrated at ade2 in the two red1::LEU2 sae2 haploids. Fig.4).4). Mitotic Recombination - an overview | ScienceDirect Topics Dephosphorylation of Red1p by the Glc7p phosphatase is then necessary for cells to exit pachytene in the BR strain background (3). Our findings are helpful in understanding the specific features of meiotic chromatid recombination and segregation in human oocytes. spo13 mutants undergo a single meiotic division, thereby obviating the need for crossovers to produce viable spores (27). Red1p and Mek1p have been proposed to provide a chromosomal context in which recombination intermediates must be generated if they are to be sensed by the checkpoint (49). RED1, but not HOP1, is required for wild-type levels of sister chromatid cohesion (4). This plasmid was constructed identically to pDW28 except that the 1.2-kb EcoRI/BglII fragment was obtained from pNH223-K348E. DNA was isolated from vegetative cells (0 h) or cells 5 h after transfer to sporulation medium (5 h) as described in Materials and Methods. In the course of this work we discovered that GAD-HOP1 produces a much stronger signal when combined with the lexA-RED11426 C-terminal truncation (Fig. In yeast, HOP1 and RED1 are required during meiosis for proper chromosome segregation and the consequent formation of viable spores. Bethesda, MD 20894, Web Policies National Library of Medicine Repression of harmful meiotic recombination in centromeric regions. No signal was observed with any of these combinations, indicating that homo-oligomerization is specific to the C-terminal fragment of RED1 (Fig. Three independent cultures were examined for each strain. Despite just a twofold reduction in recombination, however, the red1-K348E SPO13 diploid exhibited low levels of spore viability (13.8%). Quantitation of the most prominent DSB fragment indicated that the hop1::LEU2 and red1::LEU2 diploids exhibited 11.9 and 46.8%, respectively, of the level of wild-type DSBs. We thank Neta Dean, JoAnne Engebrecht, Bernadette Holdener, Michael Lichten, and Aaron Neiman for helpful discussions. Consistent with published work from other labs, deletion of RED1 reduced, but did not abolish, crossing over (Table (Table5).5). 2022 Apr 27;13(5):777. doi: 10.3390/genes13050777. The process of meiotic recombination involves the following: (a) the initiation of meiosis (a developmental switch), (b) enzymes required to perform the steps in recombination, (c) the association of homologous chromosomes, (d) the initiation of recombination, and (e) physical exchange of genetic material. This result provides strong evidence that a lack of Red1p-Hop1p hetero-oligomers is responsible for the SC formation defect. While none of the nuclei from the red1-K348E diploid NH305::pBB14 formed wild-type SCs (Table (Table6),6), 10.5% of 200 nuclei from this strain exhibited nearly wild-type SCs when HOP1 was overexpressed (Fig. Probing of the Hop1p IPs with -HA antibodies reproduced the Red1-3HAp co-IP observed previously (Fig. Genetics. 8600 Rockville Pike Only 13.8% of the spores from the red1-K348E SPO13 diploid (NH246::pSB3-K348E Spo13+; 76 asci) were viable. Deconstructing meiosis one kinase at a time: polo pushes past pachytene Bypass of a meiotic checkpoint by overproduction of meiotic chromosomal proteins. One plus sign indicates that the cells turned blue in 90 min at 30C; three plus signs indicates the cells turned blue by 15 min; a minus sign indicates a failure to turn blue by 4 h. Secondary structure programs predict two distinct regions of helix in Red1p. 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. Cells were removed at different time points and fixed with 3.7% formaldehyde for 24 h. The cells were then washed twice with phosphate-buffered saline and stained with 1 M DAPI (4,6-diamidino-2-phenylindole) for 10 min. The SK1 diploids were generated using transformed derivatives of two different combinations of parental strains (S2683 RKY1145 and NH212-1-1 NH212-35-1 [Table 2]). Single cell genome sequencing data of PB1, PB2 and FPN that originated from the same oocyte were used to analyse the human oocyte homologous chromosome interaction and segregation. Furthermore, the red1-K348E sae2 mutant produced increased levels of tetranucleate cells relative to the sae2 strain (82.7% versus 1.2%), as well more mature asci (20.2% versus 1.2%). Sperm chromosome complements in a man heterozygous for a reciprocal translocation 46,XY,t(9;13)(q21.1;q21.2) and a review of the literature. The haploidization that occurs during meiosis results in one-fourth of the spores being resistant to both drugs. Our assay allowed us to evaluate recombination events in the interstitial segments at adjacent II segregation. Thus, four sex cells, haploid cells, can be formed from one diploid cell. Meiotic analyses show adaptations to maintenance of fertility in The red1-K348E mutant exhibited only a 2-fold reduction in crossing over, compared to the 11-fold decrease in the red1 mutant. These experiments suggested that a Hop1p interaction domain is contained within the 100 amino acids located between positions 326 and 426. Estop AM, Van Kirk V, Cieply K. Segregation analysis of four translocations, t(2;18), t(3;15), t(5;7), and t(10;12), by sperm chromosome studies and a review of the literature. Bailis J M, Roeder G S. Pachytene exit controlled by reversal of Mek1-dependent phosphorylation. An official website of the United States government. Before Meiotic crossovers facilitate the segregation of homologous chromosomes and increase genetic diversity. All of the SK1 diploids should, therefore, be isogenic with the exception of the markers listed in the genotypes. Methods: Single cell genome sequencing data of PB1, PB2 and FPN that originated from the same oocyte were used to analyse the human oocyte homologous chromosome interaction and segregation. To test whether the C-terminal 291 amino acids are sufficient for Red1p-Red1p interaction, a lexA fusion to codons 536 to 827 of RED1 was assayed for interaction with GAD-RED1537827. doi: 10.1093/genetics/iyab054. indicating a previously unknown role for HOP1 in the meiotic recombination checkpoint. The lexA-RED11400 protein ends just after the predicted -helical domain, while the lexA-RED11359 protein deletes the predicted helix. (Fig.1).1). Ma JY, Feng X, Xie FY, Li S, Chen LN, Luo SM, Yin S, Ou XH. It is possible that Red1p may interact with itself through these -helical regions, one of which is contained within the C-terminal fragment known to undergo homo-oligomerization. The FPN genome carries an assembly of maternal and paternal genome that resulted from homologous recombination during the prophase of the first meiosis. Muniyappa K, Anuradha S, Byers B. Yeast meiosis-specific protein Hop1 binds to G4 DNA and promotes its formation. Overexpression of HOP1 significantly increased spore viability in the red1-K348E strain (2; P < 0.001), while the YEp24 vector had no effect (Table (Table4,4, experiment B). A likely function for Red1p-Hop1p complexes is in the formation of crossovers that are effective for MI segregation. Sequencing of the entire gene confirmed the absence of any additional mutations. The first method utilized three DNA probes (a telomeric and a centromeric probe on chromosome 1 plus a centromeric probe on c These DSBs occur in GC-rich domains (40). The https:// ensures that you are connecting to the The lexA-RED1536827 allele was also created by PCR. SAE2/COM1 is a gene which, when deleted, exhibits the same phenotypes as rad50S, including unprocessed DSBs, a delay in the onset of MI and MII, and a decrease in the number of mature asci (30, 35). In contrast, mutants that disrupt AE formation exhibit defects in sister chromatid cohesion, recombination, and chromosome segregation (4, 23, 36). Red1p, a MEK1-dependent phosphoprotein that physically interacts with Hop1p during meiosis in yeast. 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. 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