Regulation of Msh4-Msh5 association with meiotic chromosomes in budding yeast

Genetics ◽  
2021 ◽  
Author(s):  
Krishnaprasad G Nandanan ◽  
Sagar Salim ◽  
Ajith V Pankajam ◽  
Miki Shinohara ◽  
Gen Lin ◽  
...  
Keyword(s):  
Binding Sites ◽  
Holliday Junction ◽  
Crossing Over ◽  
Wild Type ◽  
Yeast Saccharomyces Cerevisiae ◽  
Meiotic Chromosomes ◽  
Genome Wide ◽  
Double Holliday Junction

Abstract In the baker’s yeast Saccharomyces cerevisiae, most of the meiotic crossovers are generated through a pathway involving the highly conserved mismatch repair related Msh4-Msh5 complex. To understand the role of Msh4-Msh5 in meiotic crossing over, we determined its genome wide in vivo binding sites in meiotic cells. We show that Msh5 specifically associates with DSB hotspots, chromosome axes, and centromeres on chromosomes. A basal level of Msh5 association with these chromosomal features is observed even in the absence of DSB formation (spo11Δ mutant) at the early stages of meiosis. But efficient binding to DSB hotspots and chromosome axes requires DSB formation and resection and is enhanced by double Holliday junction structures. Msh5 binding is also correlated to DSB frequency and enhanced on small chromosomes with higher DSB and crossover density. The axis protein Red1 is required for Msh5 association with the chromosome axes and DSB hotspots but not centromeres. Although binding sites of Msh5 and other pro-crossover factors like Zip3 show extensive overlap, Msh5 associates with centromeres independent of Zip3. These results on Msh5 localization in wild type and meiotic mutants have implications for how Msh4-Msh5 works with other pro-crossover factors to ensure crossover formation.

Control of meiotic recombination in Arabidopsis: role of the MutL and MutS homologues

2006 ◽  
Vol 34 (4) ◽  
pp. 542-544 ◽  
Author(s):  
F.C.H. Franklin ◽  
J.D. Higgins ◽  
E. Sanchez-Moran ◽  
S.J. Armstrong ◽  
K.E. Osman ◽  
...  
Keyword(s):  
Mismatch Repair ◽  
Meiotic Recombination ◽  
Holliday Junction ◽  
Crossing Over ◽  
Wild Type ◽  
Pathway Model ◽  
Junction Formation ◽  
Double Holliday Junction ◽  
Dependent Pathway

Immunocytochemistry reveals that the Arabidopsis mismatch repair proteins AtMSH4, AtMLH3 and AtMLH1 are expressed during prophase I of meiosis. Expression of AtMSH4 precedes AtMLH3 and AtMLH1 which co-localize as foci during pachytene. Co-localization between AtMSH4 and AtMLH3 occurs, but appears transient. AtMLH3 foci are not detected in an Atmsh4 mutant. However, localization of AtMSH4 is unaffected in Atmlh3, suggesting that recombination may proceed to dHj (double Holliday junction) formation. Mean chiasma frequency in Atmsh4 is reduced to 1.55 compared with 9.86 in wild-type. In contrast with wild-type, the distribution of residual crossovers in Atmsh4 closely fits a Poisson distribution. This is consistent with a two-pathway model for meiotic crossing-over whereby most crossovers occur via an AtMSH4-dependent pathway that is subject to interference, with the remaining crossovers arising via an interference-independent pathway. Loss of AtMLH3 results in an approx. 60% reduction in crossovers. Results suggest that dHj resolution can occur, but in contrast with wild-type where most or all dHjs are directed to form crossovers, the outcome is biased in favour of a non-crossover outcome. The results are compatible with a model whereby the MutL complex maintains or imposes a dHj conformation that ensures crossover formation.

scholarly journals Corepression of the P1 Addiction Operon by Phd and Doc

1998 ◽  
Vol 180 (23) ◽  
pp. 6342-6351 ◽  
Author(s):  
Roy Magnuson ◽  
Michael B. Yarmolinsky
Keyword(s):  
Fusion Protein ◽  
Binding Sites ◽  
Wild Type ◽  
Dna Complexes ◽  
Present Evidence ◽  
Cooperative Interactions ◽  
Regulatory Properties

ABSTRACT The P1 plasmid addiction operon encodes Doc, a toxin that kills plasmid-free segregants, and Phd, an unstable antidote that neutralizes the toxin. Additionally, these products repress transcription of the operon. The antidote binds to two adjacent sites in the promoter. Here we present evidence concerning the regulatory role of the toxin, which we studied with the aid of a mutation,docH66Y. The DocH66Y protein retained the regulatory properties of the wild-type protein, but not its toxicity. In vivo, DocH66Y enhanced repression by Phd but failed to affect repression in the absence of Phd, suggesting that DocH66Y contacts Phd. In vitro, a MalE-DocH66Y fusion protein was found to bind Phd. Binding of toxin to antidote may be the physical basis for the neutralization of toxin. DocH66Y failed to bind DNA in vitro yet enhanced the affinity, cooperativity, and specificity with which Phd bound the operator. Although DocH66Y enhanced the binding of Phd to two adjacent Phd-binding sites, DocH66Y had relatively little effect on the binding of Phd to a single Phd-binding site, indicating that DocH66Y mediates cooperative interactions between adjacent Phd-binding sites. Several electrophoretically distinct protein-DNA complexes were observed with different amounts of DocH66Y relative to Phd. Maximal repression and specificity of DNA binding were observed with subsaturating amounts of DocH66Y relative to Phd. Analogous antidote-toxin pairs appear to have similar autoregulatory circuits. Autoregulation, by dampening fluctuations in the levels of toxin and antidote, may prevent the inappropriate activation of the toxin.

scholarly journals The Role of Gcr1p in the Transcriptional Activation of Glycolytic Genes in Yeast Saccharomyces cerevisiae

Genetics ◽  
1997 ◽  
Vol 147 (2) ◽  
pp. 521-532 ◽  
Author(s):  
Hiroshi Uemura ◽  
Miho Koshio ◽  
Yoko Inoue ◽  
M Cecilia Lopez ◽  
Henry V Baker
Keyword(s):  
Transcriptional Activation ◽  
Binding Sites ◽  
Regulatory Sequences ◽  
Lacz Reporter ◽  
Lacz Reporter Gene ◽  
Yeast Saccharomyces Cerevisiae ◽  
Glycolytic Gene ◽  
Synthetic Oligonucleotides

To study the interdependence of Gcr1p and Rap1p, we prepared a series of synthetic regulatory sequences that contained various numbers and combinations of CT-boxes (Gcr1p-binding sites) and RPG-boxes (Rap1p-binding sites). The ability of the synthetic oligonucleotides to function as regulatory sequences was tested using an ENO1-lacZ reporter gene. As observed previously, synthetic oligonucleotides containing both CT- and RPG-boxes conferred strong UAS activity. Likewise, a lone CT-box did not show any UAS activity. By contrast, oligonucleotides containing tandem CT-boxes but no RPG-box conferred strong promoter activity. This UAS activity was not dependent on position or orientation of the oligonucleotides in the 5′ noncoding region. However, it was dependent on both GCR1 and GCR2. These results suggest that the ability of Gcr1p to bind Gcr1p-binding sites in vivo is not absolutely dependent on Rap1p. Eleven independent mutants of GCR1 were isolated that conferred weak UAS activity to a single CT-box. Five mutants had single mutations in Gcr1p's DNA-binding domain and displayed slightly higher affinity for the CT-box. These results support the hypothesis that Gcr1p and Gcr2p play the central role in glycolytic gene expression and that the function of Rap1p is to facilitate the binding of Gcr1p to its target.

scholarly journals Genome-wide analysis of STAT3 binding in vivo predicts effectors of the anti-inflammatory response in macrophages

Blood ◽  
2012 ◽  
Vol 119 (13) ◽  
pp. e110-e119 ◽  
Author(s):  
Andrew Paul Hutchins ◽  
Stéphane Poulain ◽  
Diego Miranda-Saavedra
Keyword(s):  
Inflammatory Response ◽  
Binding Sites ◽  
Peritoneal Macrophages ◽  
Rna Seq ◽  
Genome Wide ◽  
Depth Study ◽  
Anti Inflammatory ◽  
Generation Sequencing

Abstract Inflammation is a powerful response of the immune system against invading pathogens, and must be cancelled when unneeded or otherwise death inevitably follows. In macrophages, the anti-inflammatory response (AIR) is driven by STAT3 upon IL-10 signaling. The role of STAT3 is to stimulate the expression of specific genes that in-turn suppress the transcription of proinflammatory genes. Here we describe a systematic approach to identify the elusive STAT3-controlled effectors of the AIR. In vivo STAT3-binding sites were identified by ChIP-seq, coupled to expression analysis by RNA-seq, both in resting and IL-10–treated peritoneal macrophages. We report the genomic targets of STAT3 and show that STAT3's transcriptional program during the AIR is highly specific to IL-10–stimulated macrophages, that STAT3 is a positive transcriptional regulator, and we predict severalputative AIR factors that merit further investigation. This is the first in-depth study of the AIR by next-generation sequencing and provides an unprecedented degree of detail into this fundamental physiologic response.

scholarly journals SUT1-promoted sterol uptake involves the ABC transporter Aus1 and the mannoprotein Dan1 whose synergistic action is sufficient for this process

2004 ◽  
Vol 381 (1) ◽  
pp. 195-202 ◽  
Author(s):  
Parissa ALIMARDANI ◽  
Matthieu RÉGNACQ ◽  
Carole MOREAU-VAUZELLE ◽  
Thierry FERREIRA ◽  
Tristan ROSSIGNOL ◽  
...  
Keyword(s):  
Abc Transporter ◽  
Constitutive Expression ◽  
Open Reading Frames ◽  
Wild Type ◽  
Gene Encoding ◽  
Yeast Saccharomyces Cerevisiae ◽  
Sterol Uptake ◽  
Genome Wide ◽  
A Genome

Efficient sterol influx in the yeast Saccharomyces cerevisiae is restricted to anaerobiosis or to haem deficiency resulting from mutations. Constitutive expression of SUT1, an hypoxic gene encoding a transcriptional regulator, induces sterol uptake in aerobiosis. A genome-wide approach using DNA microarray was used to identify the mediators of SUT1 effects on aerobic sterol uptake. A total of 121 ORFs (open reading frames) were significantly and differentially expressed after SUT1 overexpression, 61 down-regulated and 60 up-regulated. Among these genes, the role of the putative ABC transporter (ATP-binding-cassette transporter) Aus1, and of the cell-wall mannoprotein Dan1, was characterized better. These two genes play an essential role in aerobic sterol uptake, since their deletion compromised the SUT1 effects, but individual overexpression of either of these genes in a wild-type background was not sufficient for this process. However, constitutive co-expression of AUS1 and DAN1 in a wild-type background resulted in sterol influx in aerobiosis. These results suggest that the corresponding proteins may act synergistically in vivo to promote sterol uptake.

scholarly journals Roles of Two ATPase-Motif-containing Domains in Cyanobacterial Circadian Clock Protein KaiC

2004 ◽  
Vol 279 (50) ◽  
pp. 52331-52337 ◽  
Author(s):  
Fumio Hayashi ◽  
Noriyo Itoh ◽  
Tatsuya Uzumaki ◽  
Ryo Iwase ◽  
Yuka Tsuchiya ◽  
...  
Keyword(s):  
Reaction Mechanism ◽  
Circadian Clock ◽  
Binding Sites ◽  
Wild Type ◽  
High Affinity ◽  
High Affinity Site ◽  
Clock Protein

Cyanobacterial clock protein KaiC has a hexagonal, pot-shaped structure composed of six identical dumbbell-shaped subunits. Each subunit has duplicated domains, and each domain has a set of ATPase motifs. The two spherical regions of the dumbbell are likely to correspond to two domains. We examined the role of the two sets of ATPase motifs by analyzing thein vitroactivity of ATPγS binding, AMPPNP-induced hexamerization, thermostability, and phosphorylation of KaiC and byin vivorhythm assays both in wild type KaiC (KaiCWT) and KaiCs carrying mutations in either Walker motif A or deduced catalytic Glu residues. We demonstrated that 1) the KaiC subunit had two types of ATP-binding sites, a high affinity site in N-terminal ATPase motifs and a low affinity site in C-terminal ATPase motifs, 2) the N-terminal motifs were responsible for hexamerization, and 3) the C-terminal motifs were responsible for both stabilization and phosphorylation of the KaiC hexamer. We proposed the following reaction mechanism. ATP preferentially binds to the N-terminal high affinity site, inducing the hexamerization of KaiC. Additional ATP then binds to the C-terminal low affinity site, stabilizing and phosphorylating the hexamer. We discussed the effect of these KaiC mutations on circadian bioluminescence rhythm in cells of cyanobacteria.

The Budding Yeast Msh4 Protein Functions in Chromosome Synapsis and the Regulation of Crossover Distribution

Genetics ◽  
2001 ◽  
Vol 158 (3) ◽  
pp. 1013-1025 ◽  
Author(s):  
Janet E Novak ◽  
Petra B Ross-Macdonald ◽  
G Shirleen Roeder
Keyword(s):  
Mismatch Repair ◽  
Budding Yeast ◽  
Null Mutation ◽  
Crossing Over ◽  
Wild Type ◽  
Crossover Interference ◽  
Meiotic Chromosomes ◽  
Chromosome Synapsis ◽  
Protein Functions ◽  
Or Gene

AbstractThe budding yeast MSH4 gene encodes a MutS homolog produced specifically in meiotic cells. Msh4 is not required for meiotic mismatch repair or gene conversion, but it is required for wild-type levels of crossing over. Here, we show that a msh4 null mutation substantially decreases crossover interference. With respect to the defect in interference and the level of crossing over, msh4 is similar to the zip1 mutant, which lacks a structural component of the synaptonemal complex (SC). Furthermore, epistasis tests indicate that msh4 and zip1 affect the same subset of meiotic crossovers. In the msh4 mutant, SC formation is delayed compared to wild type, and full synapsis is achieved in only about half of all nuclei. The simultaneous defects in synapsis and interference observed in msh4 (and also zip1 and ndj1/tam1) suggest a role for the SC in mediating interference. The Msh4 protein localizes to discrete foci on meiotic chromosomes and colocalizes with Zip2, a protein involved in the initiation of chromosome synapsis. Both Zip2 and Zip1 are required for the normal localization of Msh4 to chromosomes, raising the possibility that the zip1 and zip2 defects in crossing over are indirect, resulting from the failure to localize Msh4 properly.

scholarly journals Arabidopsis Disulfide Reductase, Trx-h2, Functions as an RNA Chaperone under Cold Stress

2021 ◽  
Vol 11 (15) ◽  
pp. 6865
Author(s):  
Eun Seon Lee ◽  
Joung Hun Park ◽  
Seong Dong Wi ◽  
Ho Byoung Chae ◽  
Seol Ki Paeng ◽  
...  
Keyword(s):  
Cold Stress ◽  
Wild Type ◽  
Rna Chaperone ◽  
E Coli ◽  
Cold Sensitive ◽  
Thioredoxin H ◽  
Functional Rnas

The thioredoxin-h (Trx-h) family of Arabidopsis thaliana comprises cytosolic disulfide reductases. However, the physiological function of Trx-h2, which contains an additional 19 amino acids at its N-terminus, remains unclear. In this study, we investigated the molecular function of Trx-h2 both in vitro and in vivo and found that Arabidopsis Trx-h2 overexpression (Trx-h2OE) lines showed significantly longer roots than wild-type plants under cold stress. Therefore, we further investigated the role of Trx-h2 under cold stress. Our results revealed that Trx-h2 functions as an RNA chaperone by melting misfolded and non-functional RNAs, and by facilitating their correct folding into active forms with native conformation. We showed that Trx-h2 binds to and efficiently melts nucleic acids (ssDNA, dsDNA, and RNA), and facilitates the export of mRNAs from the nucleus to the cytoplasm under cold stress. Moreover, overexpression of Trx-h2 increased the survival rate of the cold-sensitive E. coli BX04 cells under low temperature. Thus, our data show that Trx-h2 performs function as an RNA chaperone under cold stress, thus increasing plant cold tolerance.

scholarly journals Interplay between Pds5 and Rec8 in regulating chromosome axis length and crossover frequency

2021 ◽  
Vol 7 (11) ◽  
pp. eabe7920
Author(s):  
Meihui Song ◽  
Binyuan Zhai ◽  
Xiao Yang ◽  
Taicong Tan ◽  
Ying Wang ◽  
...  
Keyword(s):  
Recombination Frequency ◽  
Crossover Frequency ◽  
Wild Type ◽  
Protein Levels ◽  
Homolog Pairing ◽  
Meiotic Chromosomes ◽  
Homologous Chromosome Pairing ◽  
Axis Length ◽  
Chromosome Axis

Meiotic chromosomes have a loop/axis architecture, with axis length determining crossover frequency. Meiosis-specific Pds5 depletion mutants have shorter chromosome axes and lower homologous chromosome pairing and recombination frequency. However, it is poorly understood how Pds5 coordinately regulates these processes. In this study, we show that only ~20% of wild-type level of Pds5 is required for homolog pairing and that higher levels of Pds5 dosage-dependently regulate axis length and crossover frequency. Moderate changes in Pds5 protein levels do not explicitly impair the basic recombination process. Further investigations show that Pds5 does not regulate chromosome axes by altering Rec8 abundance. Conversely, Rec8 regulates chromosome axis length by modulating Pds5. These findings highlight the important role of Pds5 in regulating meiosis and its relationship with Rec8 to regulate chromosome axis length and crossover frequency with implications for evolutionary adaptation.

scholarly journals Essential role of glucose transporter GLUT3 for post-implantation embryonic development

2008 ◽  
Vol 200 (1) ◽  
pp. 23-33 ◽  
Author(s):  
S Schmidt ◽  
A Hommel ◽  
V Gawlik ◽  
R Augustin ◽  
N Junicke ◽  
...  
Keyword(s):  
Essential Role ◽  
Glucose Transporter ◽  
Growth Arrest ◽  
Complete Loss ◽  
Transporter Gene ◽  
Wild Type ◽  
Post Implantation ◽  
Time Point

Deletion of glucose transporter geneSlc2a3(GLUT3) has previously been reported to result in embryonic lethality. Here, we define the exact time point of growth arrest and subsequent death of the embryo.Slc2a3−/−morulae and blastocysts developed normally, implantedin vivo, and formed egg-cylinder-stage embryos that appeared normal until day 6.0. At day 6.5, apoptosis was detected in the ectodermal cells ofSlc2a3−/−embryos resulting in severe disorganization and growth retardation at day 7.5 and complete loss of embryos at day 12.5. GLUT3 was detected in placental cone, in the visceral ectoderm and in the mesoderm of 7.5-day-old wild-type embryos. Our data indicate that GLUT3 is essential for the development of early post-implanted embryos.

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