Fanconi anemia ortholog FANCM regulates meiotic crossover distribution in plants

Abstract Meiotic recombination increases genetic diversity and manipulation of its frequency and distribution holds great promise in crop breeding. In Arabidopsis thaliana, FANCM (a homolog of mammalian Fanconi anemia complementation group M) suppresses recombination and its function seems conserved in other species including the rosids Brassica spp. and pea (Pisum sativum), and the monocot rice (Oryza sativa). To examine the role of FANCM during meiotic recombination in lettuce (Lactuca sativa, an asterid), we characterized the function of lettuce LsFANCM and found that it can functionally substitute for AtFANCM in transgenic Arabidopsis plants. Moreover, three independent CRISPR/Cas9-edited lettuce Lsfancm mutants showed reduced pollen viability and seed setting. Unexpectedly, analyses of chromosome behavior revealed that 77.8% of Lsfancm meiocytes exhibited univalents. The normal formation of double-strand breaks in DNA and the discontinuous assembly of synaptonemal complex in Lsfancm mutants supports the hypothesis that LsFANCM might be dispensable for the initiation of meiotic recombination but required for normal synapsis. Furthermore, the frequency of lettuce HEI10 (Human Enhancer of Invasion 10) foci, a marker for Class-I crossovers (COs), was similar between wild-type (WT) and Lsfancm. Strikingly, the distribution of LsHEI10 foci and chiasmata in Lsfancm meiotic chromosomes was markedly different from the WT. A similar alteration in the distribution of Class-I COs was also observed in the Arabidopsis Atfancm mutant. Taken together, these results demonstrate that FANCM is important for shaping the distribution of meiotic Class-I COs in plants, and reveal an evolutionarily divergent role for FANCM in meiotic bivalent formation between Arabidopsis and lettuce.

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Multiple mechanisms limit meiotic crossovers: TOP3α and two BLM homologs antagonize crossovers in parallel to FANCM

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1423107112 ◽

2015 ◽

Vol 112 (15) ◽

pp. 4713-4718 ◽

Cited By ~ 56

Author(s):

Mathilde Séguéla-Arnaud ◽

Wayne Crismani ◽

Cécile Larchevêque ◽

Julien Mazel ◽

Nicole Froger ◽

...

Keyword(s):

Chromosome Segregation ◽

Complementation Group ◽

Great Promise ◽

Dna Double Strand Breaks ◽

Strand Breaks ◽

Evolutionary Forces ◽

Mechanical Constraints ◽

Syndrome Protein

Meiotic crossovers (COs) have two important roles, shuffling genetic information and ensuring proper chromosome segregation. Despite their importance and a large excess of precursors (i.e., DNA double-strand breaks, DSBs), the number of COs is tightly regulated, typically one to three per chromosome pair. The mechanisms ensuring that most DSBs are repaired as non-COs and the evolutionary forces imposing this constraint are poorly understood. Here we identified Topoisomerase3α (TOP3α) and the RECQ4 helicases—the Arabidopsis slow growth suppressor 1 (Sgs1)/Bloom syndrome protein (BLM) homologs—as major barriers to meiotic CO formation. First, the characterization of a specific TOP3α mutant allele revealed that, in addition to its role in DNA repair, this topoisomerase antagonizes CO formation. Further, we found that RECQ4A and RECQ4B constitute the strongest meiotic anti-CO activity identified to date, their concomitant depletion leading to a sixfold increase in CO frequency. In both top3α and recq4ab mutants, DSB number is unaffected, and extra COs arise from a normally minor pathway. Finally, both TOP3α and RECQ4A/B act independently of the previously identified anti-CO Fanconi anemia of complementation group M (FANCM) helicase. This finding shows that several parallel pathways actively limit CO formation and suggests that the RECQA/B and FANCM helicases prevent COs by processing different substrates. Despite a ninefold increase in CO frequency, chromosome segregation was unaffected. This finding supports the idea that CO number is restricted not because of mechanical constraints but likely because of the long-term costs of recombination. Furthermore, this work demonstrates how manipulating a few genes holds great promise for increasing recombination frequency in plant-breeding programs.

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Mouse TEX15 is essential for DNA double-strand break repair and chromosomal synapsis during male meiosis

The Journal of Cell Biology ◽

10.1083/jcb.200709057 ◽

2008 ◽

Vol 180 (4) ◽

pp. 673-679 ◽

Cited By ~ 75

Author(s):

Fang Yang ◽

Sigrid Eckardt ◽

N. Adrian Leu ◽

K. John McLaughlin ◽

Peijing Jeremy Wang

Keyword(s):

Meiotic Recombination ◽

Strand Break ◽

Male Meiosis ◽

Double Strand Breaks ◽

Dna Double Strand Breaks ◽

Strand Breaks ◽

Homologous Chromosomes ◽

Recombination Proteins ◽

Meiotic Chromosomes ◽

Dna Double Strand Break

During meiosis, homologous chromosomes undergo synapsis and recombination. We identify TEX15 as a novel protein that is required for chromosomal synapsis and meiotic recombination. Loss of TEX15 function in mice causes early meiotic arrest in males but not in females. Specifically, TEX15-deficient spermatocytes exhibit a failure in chromosomal synapsis. In mutant spermatocytes, DNA double-strand breaks (DSBs) are formed, but localization of the recombination proteins RAD51 and DMC1 to meiotic chromosomes is severely impaired. Based on these data, we propose that TEX15 regulates the loading of DNA repair proteins onto sites of DSBs and, thus, its absence causes a failure in meiotic recombination.

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The yeast MER2 gene is required for chromosome synapsis and the initiation of meiotic recombination.

Genetics ◽

10.1093/genetics/141.1.49 ◽

1995 ◽

Vol 141 (1) ◽

pp. 49-59

Author(s):

B Rockmill ◽

J A Engebrecht ◽

H Scherthan ◽

J Loidl ◽

G S Roeder

Keyword(s):

Meiotic Recombination ◽

Microscopic Analysis ◽

Primary Defect ◽

Electron Microscopic ◽

Strand Breaks ◽

Meiotic Chromosomes ◽

Chromosome Synapsis ◽

Damage Repair

Abstract Mutation of the MER2 gene of Saccharomyces cerevisiae confers meiotic lethality. To gain insight into the function of the Mer2 protein, we have carried out a detailed characterization of the mer2 null mutant. Genetic analysis indicates that mer2 completely eliminates meiotic interchromosomal gene conversion and crossing over. In addition, mer2 abolishes intrachromosomal meiotic recombination, both in the ribosomal DNA array and in an artificial duplication. The results of a physical assay demonstrate that the mer2 mutation prevents the formation of meiosis-specific, double-strand breaks, indicating that the Mer2 protein acts at or before the initiation of meiotic recombination. Electron microscopic analysis reveals that the mer2 mutant makes axial elements, which are precursors to the synaptonemal complex, but homologous chromosomes fail to synapse. Fluorescence in situ hybridization of chromosome-specific DNA probes to spread meiotic chromosomes demonstrates that homolog alignment is also significantly reduced in the mer2 mutant. Although the MER2 gene is transcribed during vegetative growth, deletion or overexpression of the MER2 gene has no apparent effect on mitotic recombination or DNA damage repair. We suggest that the primary defect in the mer2 mutant is in the initiation of meiotic genetic exchange.

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mei-P22Encodes a Chromosome-Associated Protein Required for the Initiation of Meiotic Recombination inDrosophila melanogaster

Genetics ◽

10.1093/genetics/162.1.245 ◽

2002 ◽

Vol 162 (1) ◽

pp. 245-258 ◽

Cited By ~ 10

Author(s):

Hao Liu ◽

Janet K Jang ◽

Naohiro Kato ◽

Kim S McKim

Keyword(s):

Meiotic Recombination ◽

Meiotic Prophase ◽

Direct Evidence ◽

Double Strand Breaks ◽

Crossing Over ◽

Wild Type ◽

Direct Role ◽

Strand Breaks ◽

Meiotic Chromosomes ◽

X Irradiation

AbstractDouble-strand breaks (DSB) initiate meiotic recombination in a variety of organisms. Here we present genetic evidence that the mei-P22 gene is required for the induction of DSBs during meiotic prophase in Drosophila females. Strong mei-P22 mutations eliminate meiotic crossing over and suppress the sterility of DSB repair-defective mutants. Interestingly, crossing over in mei-P22 mutants can be restored to almost 50% of wild-type by X irradiation. In addition, an antibody-based assay was used to demonstrate that DSBs are not formed in mei-P22 mutants. This array of phenotypes is identical to that of mei-W68 mutants; mei-W68 encodes the Drosophila Spo11 homolog that is proposed to be an enzyme required for DSB formation. Consistent with a direct role in DSB formation, mei-P22 encodes a basic 35.7-kD protein, which, when examined by immunofluorescence, localizes to foci on meiotic chromosomes. MEI-P22 foci appear transiently in early meiotic prophase, which is when meiotic recombination is believed to initiate. By using an antibody to C(3)G as a marker for synaptonemal complex (SC) formation, we observed that SC is present before MEI-P22 associates with the chromosomes, thus providing direct evidence that the development of SC precedes the initiation of meiotic recombination. Similarly, we found that MEI-P22 foci did not appear in a c(3)G mutant in which SC does not form, suggesting that DSB formation is dependent on SC formation in Drosophila. We propose that MEI-P22 interacts with meiosis-specific chromosome proteins to facilitate DSB creation by MEI-W68.

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Functional Interactions Between SPO11 and REC102 During Initiation of Meiotic Recombination in Saccharomyces cerevisiae

Genetics ◽

10.1093/genetics/160.1.111 ◽

2002 ◽

Vol 160 (1) ◽

pp. 111-122

Author(s):

Kehkooi Kee ◽

Scott Keeney

Keyword(s):

Saccharomyces Cerevisiae ◽

Meiotic Recombination ◽

Dna Double Strand Breaks ◽

Strand Breaks ◽

Whole Cells ◽

Meiotic Gene ◽

Meiotic Chromosomes ◽

Cell Extracts ◽

Cold Sensitive

Abstract In Saccharomyces cerevisiae, formation of the DNA double-strand breaks (DSBs) that initiate meiotic recombination requires the products of at least 10 genes. Spo11p is thought to be the catalytic subunit of the DNA cleaving activity, but the roles of the other proteins, and the interactions among them, are not well understood. This study demonstrates genetic and physical interactions between the products of SPO11 and another early meiotic gene required for DSB formation, REC102. We found that epitope-tagged versions of SPO11 and REC102 that by themselves were capable of supporting normal or nearly normal levels of meiotic recombination conferred a severe synthetic cold-sensitive phenotype when combined in the same cells. DSB formation, meiotic gene conversion, and spore viability were drastically reduced in the doubly tagged strain at a nonpermissive temperature. This conditional defect could be partially rescued by expression of untagged SPO11, but not by expression of untagged REC102, indicating that tagged REC102 is fully dominant for this synthetic phenotype. Both tagged and wild-type Spo11p co-immunoprecipitated with tagged Rec102p from meiotic cell extracts, indicating that these proteins are present in a common complex in vivo. Tagged Rec102p localized to the nucleus in whole cells and to chromatin on spread meiotic chromosomes. Our results are consistent with the idea that a multiprotein complex that includes Spo11p and Rec102p promotes meiotic DSB formation.

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Faculty Opinions recommendation of Fanconi anemia complementation group A (FANCA) localizes to centrosomes and functions in the maintenance of centrosome integrity.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.718038032.793492303 ◽

2014 ◽

Author(s):

Wade Clapp ◽

Grzegorz Nalepa

Keyword(s):

Fanconi Anemia ◽

Complementation Group ◽

Group A

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A human ortholog of archaeal DNA repair protein Hef is defective in Fanconi anemia complementation group M

Nature Genetics ◽

10.1038/ng1626 ◽

2005 ◽

Vol 37 (9) ◽

pp. 958-963 ◽

Cited By ~ 298

Author(s):

Amom Ruhikanta Meetei ◽

Annette L Medhurst ◽

Chen Ling ◽

Yutong Xue ◽

Thiyam Ramsing Singh ◽

...

Keyword(s):

Dna Repair ◽

Fanconi Anemia ◽

Complementation Group ◽

Repair Protein ◽

Dna Repair Protein ◽

Human Ortholog

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Correction of the spontaneous and DEB-induced chromosomal aberrations in Fanconi anemia cells of the FA(C) complementation group by the FACC gene

Cytogenetic and Genome Research ◽

10.1159/000134187 ◽

1996 ◽

Vol 72 (2-3) ◽

pp. 194-196 ◽

Cited By ~ 2

Author(s):

D.J. Stavropoulos ◽

S. Sood ◽

D.J. Tomkins ◽

M. Buchwald

Keyword(s):

Fanconi Anemia ◽

Chromosomal Aberrations ◽

Complementation Group

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Cytosine and adenine deaminase base-editors induce broad and nonspecific changes in gene expression and splicing

Communications Biology ◽

10.1038/s42003-021-02406-5 ◽

2021 ◽

Vol 4 (1) ◽

Author(s):

Jiao Fan ◽

Yige Ding ◽

Chao Ren ◽

Ziguo Song ◽

Jie Yuan ◽

...

Keyword(s):

Gene Expression ◽

Double Strand Breaks ◽

Great Promise ◽

Single Nucleotide ◽

Strand Breaks ◽

Dna Base ◽

Target Dna ◽

Single Nucleotide Variations ◽

Adenine Deaminase ◽

Target Rna

AbstractCytosine or adenine base editors (CBEs or ABEs) hold great promise in therapeutic applications because they enable the precise conversion of targeted base changes without generating of double-strand breaks. However, both CBEs and ABEs induce substantial off-target DNA editing, and extensive off-target RNA single nucleotide variations in transfected cells. Therefore, the potential effects of deaminases induced by DNA base editors are of great importance for their clinical applicability. Here, the transcriptome-wide deaminase effects on gene expression and splicing is examined. Differentially expressed genes (DEGs) and differential alternative splicing (DAS) events, induced by base editors, are identified. Both CBEs and ABEs generated thousands of DEGs and hundreds of DAS events. For engineered CBEs or ABEs, base editor-induced variants had little effect on the elimination of DEGs and DAS events. Interestingly, more DEGs and DAS events are observed as a result of over expressions of cytosine and adenine deaminases. This study reveals a previously overlooked aspect of deaminase effects in transcriptome-wide gene expression and splicing, and underscores the need to fully characterize such effects of deaminase enzymes in base editor platforms.

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