DOCSLIB.ORG

Interrogating the Functions of PRDM9 Domains in Meiosis

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Interrogating the Functions of PRDM9 Domains in Meiosis Genetics: Early Online, published on April 19, 2018 as 10.1534/genetics.118.300565 1 Interrogating the functions of PRDM9 domains in meiosis 2 Sarah Thibault-Sennett*†, Qi Yu*‡, Fatima Smagulova†,§, Jeff Cloutier‡, Kevin Brick‡, R. 3 Daniel Camerini-Otero‡^ and Galina V. Petukhova†^. 4 5 * These authors contributed equally to the manuscript. 6 † Department of Biochemistry and Molecular Biology, Uniformed Services University of 7 Health Sciences, Bethesda, Maryland 20814, USA 8 ‡ Genetics and Biochemistry Branch, National Institute of Diabetes and Digestive and 9 Kidney Diseases (NIDDK), National Institutes of Health (NIH), Bethesda, Maryland 10 20892, USA 11 § Present address: 9 Avenue du Professeur Léon Bernard, 35000 Rennes, France 12 13 ^ Correspondence: [email protected] or [email protected]. 14 15 Data available at GEO (https://www.ncbi.nlm.nih.gov/geo/) with the accession number: 16 GSE104850. 1 Copyright 2018. 17 RUNNING TITLE: PRDM9 domains during meiosis 18 19 KEYWORDS: PRDM9, meiosis, homologous recombination and KRAB domain 20 21 CORRESPONDING AUTHORS: 22 Galina Petukhova, (301) 295-3564, [email protected], 23 Department of Biochemistry and Molecular Biology, USUHS, 4301 Jones Bridge Rd., 24 Bethesda, MD 20814, USA 25 R. Daniel Camerini-Otero, (301) 496-2710, [email protected] 26 National Institutes of Health, Building 5, Room 205A, 5 Memorial Dr., Bethesda, MD 27 20814 28 29 30 31 2 32 ABSTRACT 33 Homologous recombination is required for proper segregation of homologous 34 chromosomes during meiosis. It predominantly occurs at recombination hotspots that 35 are defined by the DNA binding specificity of the PRDM9 protein. PRDM9 contains 36 three conserved domains typically involved in regulation of transcription, yet, the role of 37 PRDM9 in gene expression control is not clear. Here we analyze the germline 38 transcriptome of Prdm9-/- male mice in comparison to Prdm9+/+ males and find no 39 apparent differences in the mRNA and miRNA profiles. We further explore the role of 40 PRDM9 in meiosis by analyzing the effect of the KRAB, SSXRD and post-SET zinc 41 finger deletions in a cell culture expression system and the KRAB domain deletion in 42 mice. We found that although the post-SET zinc finger and the KRAB domains are not 43 essential for the methyltransferase activity of PRDM9 in cell culture, the KRAB domain 44 mutant mice show only residual PRDM9 methyltransferase activity and undergo meiotic 45 arrest. In aggregate, our data indicate that domains typically involved in regulation of 46 gene expression do not serve that role in PRDM9, but are likely involved in setting the 47 proper chromatin environment for initiation and completion of homologous 48 recombination. 49 50 INTRODUCTION 51 Homologous recombination is required for the faithful segregation of homologous 52 chromosomes during meiosis. It assures that homologous chromosomes find each 53 other and stay connected until the first meiotic division, when they segregate to different 54 daughter cells. Failure to segregate properly results in aneuploidy in gametes and 3 55 offspring which accounts for 35% of miscarriages and developmental disabilities in 0.3% 56 of live births (Hassold et al. 2007). The most severe recombination defects lead to 57 meiotic arrest and consequent infertility (reviewed in (Handel and Schimenti 2010)). 58 59 During meiotic recombination the chromosomes undergo programmed DNA double- 60 stranded breaks (DSBs) (reviewed in (Keeney and Neale 2006)). This triggers a 61 genome-wide search for the intact homologous chromosome to be used for repair. 62 RAD51 and DMC1 recombinases bind to single-stranded tails of the resected DSB ends 63 and initiate recombinational repair of DSBs that results in either crossover or non- 64 crossover products (Keeney and Neale 2006). Meiotic DSBs preferentially occur in 65 discrete sites of the genome called recombination hotspots (reviewed in (Arnheim et al. 66 2007; Buard and de Massy 2007; Lichten 2008; Paigen and Petkov 2010; Clark et al. 67 2010)). In mice and humans, hotspot locations are defined by the DNA binding 68 specificity of the PR/SET domain containing 9 (PRDM9) protein that trimethylates 69 Lysine 4 of the histone H3 (H3K4me3) at the sites where recombination may 70 subsequently occur (Baudat et al. 2010; Parvanov et al. 2010; Myers et al. 2010; Grey 71 et al. 2011; Brick et al. 2012). The DNA binding domain of PRDM9 consists of an array 72 of zinc finger domains (ZFs) that vary primarily at three amino acids that define the 73 sequence specificity of DNA binding (reviewed in Wolfe et al. 2000; Persikov et al. 74 2009). Remarkably, the ZF array is extremely polymorphic and may consist of different 75 ZFs in different individuals, leading to a completely different distribution of 76 recombination hotspots. When PRDM9 is absent, DSBs are formed at so-called default 77 hotspots, the other available H3K4me3 sites within the genome, mostly at gene 4 78 promoters and enhancer elements (Brick et al. 2012). These default sites are almost 79 never used in wild type mice, suggesting a dominant direct interaction between PRDM9 80 and the DSB formation machinery. Indeed, recent studies implicated the KRAB domain 81 of PRDM9 in tethering the hotspot DNA to the chromosome cores, where the DSB 82 machinery resides, through an interaction with CXXC1 (Imai et al. 2017; Parvanov et al. 83 2017). Prdm9-/- mice undergo meiotic arrest due to recombination failure, leading to 84 sterility in both sexes (Hayashi et al. 2005). Interestingly, the loss of Prdm9 does not 85 universally result in sterility as Prdm9 knockouts in some species remain fertile 86 (Narasimhan et al. 2016), and the functional Prdm9 gene has been lost in several 87 lineages (Ponting 2011; Baker et al. 2017). 88 89 PRDM9 belongs to the PRDM protein family, a group that is defined by the presence of 90 a ZF array and the PR/SET domain (reviewed in (Hohenauer and Moore 2012)). The 91 PR/SET domain of PRDM9 is responsible for histone methyltransferase activity, 92 including mono-, di- and trimethylation of the H3K4 (Hayashi et al. 2005; Wu et al. 93 2013), as well as H3K36 trimethylation (Eram et al. 2014, Powers et al. 2016). This 94 domain is distantly related to the classic SET domain found in the histone lysine 95 methyltransferases, a protein family involved in regulation of gene expression. The 96 crystal structure of the PRDM9 PR/SET domain suggests that the methyltransferase 97 activity of PRDM9 is autoregulated and the lone ZF located in the post-SET region is 98 involved in this regulation (Wu et al. 2013). 99 100 PRDM family members either directly methylate their histone targets or recruit other 5 101 histone modifiers to chromatin (Hohenauer and Moore 2012). Owing to these activities 102 the PRDM family is involved in a diverse array of developmental processes through the 103 regulation of gene expression (Hohenauer and Moore 2012). PRDM9, however, is 104 unique in that it also has the Kruppel Associated Box (KRAB) domain (Liu et al. 2014). 105 This leads to inclusion of PRDM9 in a second family of transcription factors, the KRAB- 106 ZFPs (KRAB Zinc Finger Proteins). Members of this family also contain a ZF DNA- 107 binding array, but have a KRAB domain instead of PR/SET (Lupo et al. 2013). 108 109 The KRAB domain is typically involved in gene silencing through interaction with the 110 TRIM28 protein, also known as KAP-1 (reviewed in (Iyengar and Farnham 2011)). 111 TRIM28 interacts with Heterochromatin Protein 1 (HP-1) and serves as a scaffold for 112 recruiting chromatin modifying enzymes and remodeling complexes to promoters of 113 target genes bound by the KRAB-ZFPs (Iyengar and Farnham 2011). Interestingly, the 114 KRAB domain of PRDM9 is missing the amino acids required for interaction with 115 TRIM28 (Liu et al. 2014), and recent studies found no direct interaction between these 116 two proteins (Patel et al. 2016; Imai et al. 2017). In fact, the KRAB domain of PRDM9 117 has higher similarity to the atypical KRAB domain found in the Synovial Sarcoma X 118 chromosome breakpoint (SSX) protein family (Lim et al. 1998). PRDM9 and the SSX 119 family members have another similarity: they both contain the SSX Repressive Domain 120 (SSXRD) following the atypical KRAB domain. Although the SSXRD was shown to be a 121 potent transcription repressor (Lim et al. 1998) the molecular mechanism behind this 122 function has not been described. 123 6 124 Considering that PRDM9 has three domains implicated in regulation of transcription it 125 was originally thought of as a transcription factor (Hayashi et al. 2005; Matsui and 126 Hayashi 2007; Mihola et al. 2009; reviewed in Nowick et al. 2013; Capilla et al. 2016). 127 This role was attractive, because Prdm9 is the only known speciation gene in mammals, 128 and the conflict between regulatory mechanisms is perceived as the most likely cause 129 of hybrid incompatibilities (Nowick et al. 2013; Mack and Nachman 2017). Although later 130 studies suggested a different role of Prdm9 in speciation (Smagulova et al. 2016; 131 Davies et al. 2016) they did not rule out the possibility of additional mechanisms that 132 may involve misregulation of transcription. Furthermore, it was recently reported that 133 during meiosis, PRDM9 binds to some active promoters (Grey et al. 2017), bringing 134 back the question of the possible regulatory function of PRDM9 in gene expression. In 135 this study we evaluate the potential role of PRDM9 in transcription and the role of 136 transcription-related domains within PRDM9 in PRDM9 function. 137 138 We first used mRNA sequencing to demonstrate that inactivation of Prdm9 does not 139 lead to changes in gene expression. We then showed that PRDM9 does not appear to 140 regulate miRNA expression as well.
Load more

Recommended publications
  • Hotspots of Homologous Recombination in the Human Genome: Not Comment All Homologous Sequences Are Equal James R Lupski
    Minireview Hotspots of homologous recombination in the human genome: not comment all homologous sequences are equal James R Lupski Address: Departments of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX 77030, USA. Current address (sabbatical until July 2005): Wellcome Trust Sanger Institute, Hinxton, Cambridge CB10 1SA, UK. E-mail: [email protected] Published: 28 September 2004 reviews Genome Biology 2004, 5:242 The electronic version of this article is the complete one and can be found online at http://genomebiology.com/2004/5/10/242 © 2004 BioMed Central Ltd reports Abstract Homologous recombination between alleles or non-allelic paralogous sequences does not occur uniformly but is concentrated in ‘hotspots’ with high recombination rates. Recent studies of these hotspots show that they do not share common sequence motifs, but they do have other features in common. deposited research Homologous recombination is the process whereby two DNA this is not the case and have provided evidence for local sequence substrates that share a significant stretch of iden- ‘hotspots’ - short regions of the genome where strand tity are brought together, in an enzyme-catalyzed reaction, exchanges are more common than elsewhere. These obser- and undergo strand exchange to give a product that is a vations come from pedigree studies that examined the novel amalgamation of the two substrates. It occurs during parent-to-offspring transmission of alleles, linkage disequi- meiosis, leading to crossovers between alleles (allelic homol- librium (LD) studies and, more recently, direct DNA refereed research ogous recombination, AHR), and during repair of double- sequencing of the products of recombination using either strand breaks in DNA and other processes, leading to sperm (which represent a large number of recombination recombination between paralogous sequences (non-allelic products from a single meiosis) or junction fragments from homologous recombination, NAHR, also known as ectopic ectopic recombination (NAHR) [4,5].
    [Show full text]
  • Th2bs11ph Histone Mark Is Enriched in the Unsynapsed Axes of the XY Body and Predominantly Associates with H3k4me3-Containing Ge
    Mahadevan et al. Epigenetics & Chromatin (2019) 12:53 https://doi.org/10.1186/s13072-019-0300-y Epigenetics & Chromatin RESEARCH Open Access TH2BS11ph histone mark is enriched in the unsynapsed axes of the XY body and predominantly associates with H3K4me3-containing genomic regions in mammalian spermatocytes Iyer Aditya Mahadevan1†, Satyakrishna Pentakota2†, Raktim Roy3, Utsa Bhaduri1 and Manchanahalli R. Satyanarayana Rao1* Abstract Background: TH2B is a major histone variant that replaces about 80–85% of somatic H2B in mammalian spermato- cytes and spermatids. The post-translational modifcations (PTMs) on TH2B have been well characterised in spermato- cytes and spermatids. However, the biological function(s) of these PTMs on TH2B have not been deciphered in great detail. In our attempt to decipher the unique function(s) of histone variant TH2B, we detected the modifcation in the N-terminal tail, Serine 11 phosphorylation on TH2B (TH2BS11ph) in spermatocytes. Results: The current study is aimed at understanding the function of the TH2BS11ph modifcation in the context of processes that occur during meiotic prophase I. Immunofuorescence studies with the highly specifc antibodies revealed that TH2BS11ph histone mark is enriched in the unsynapsed axes of the sex body and is associated with XY body-associated proteins like Scp3, γH2AX, pATM, ATR, etc. Genome-wide occupancy studies as determined by ChIP sequencing experiments in P20 C57BL6 mouse testicular cells revealed that TH2BS11ph is enriched in X and Y chromosomes confrming the immunofuorescence staining pattern in the pachytene spermatocytes. Apart from the localisation of this modifcation in the XY body, TH2BS11ph is majorly associated with H3K4me3-containing genomic regions like gene promoters, etc.
    [Show full text]
  • Activates the M26 Meiotic Recombination Hotspot in Schizosaccharomyces Pombe
    Proc. Natl. Acad. Sci. USA Vol. 94, pp. 13765–13770, December 1997 Genetics Transcription factor Mts1yMts2 (Atf1yPcr1, Gad7yPcr1) activates the M26 meiotic recombination hotspot in Schizosaccharomyces pombe NING KON*, MICHELLE D. KRAWCHUK*, B. GREG WARREN*, GERALD R. SMITH†, AND WAYNE P. WAHLS*‡ *Department of Biochemistry, Vanderbilt University School of Medicine, 621 Light Hall, Nashville, TN 37232-0146; and †Fred Hutchinson Cancer Research Center, 1100 Fairview Avenue N., Seattle, WA 98109 Communicated by Sydney Kustu, University of California, Berkeley, CA, October 13, 1997 (received for review July 11, 1997) ABSTRACT Homologous recombination hotspots in- regulatory elements, there are proteins that interact with crease the frequency of recombination in nearby DNA. The hotspots to mediate their biological activity. In Escherichia coli M26 hotspot in the ade6 gene of Schizosaccharomyces pombe is the RecBCD enzyme interacts with Chi sites to enhance a meiotic hotspot with a discrete, cis-acting nucleotide se- recombination (12). The M26 recombination hotspot of the quence (5*-ATGACGT-3*) defined by extensive mutagenesis. A fission yeast Schizosaccharomyces pombe (5) is a well- heterodimeric M26 DNA binding protein, composed of sub- characterized eukaryotic hotspot. The M26 mutation is a single units Mts1 and Mts2, has been identified and purified 40,000- base pair substitution in ade6 that increases meiotic recombi- fold. Cloning, disruption, and genetic analyses of the mts genes nation up to 20-fold relative to other ade6 alleles, such as M375 demonstrate that the Mts1yMts2 heterodimer is essential for (Fig. 1) (5, 7, 13). Mutational analysis revealed that a specific hotspot activity. This provides direct evidence that a specific 7-bp nucleotide sequence at M26 is required for hotspot trans-acting factor, binding to a cis-acting site with a unique activity (14) (Fig.
    [Show full text]
  • A Second Generation Human Haplotype Map of Over 3.1 Million Snps
    Vol 449 | 18 October 2007 | doi:10.1038/nature06258 ARTICLES A second generation human haplotype map of over 3.1 million SNPs The International HapMap Consortium* We describe the Phase II HapMap, which characterizes over 3.1 million human single nucleotide polymorphisms (SNPs) genotyped in 270 individuals from four geographically diverse populations and includes 25–35% of common SNP variation in the populations surveyed. The map is estimated to capture untyped common variation with an average maximum r2 of between 0.9 and 0.96 depending on population. We demonstrate that the current generation of commercial genome-wide genotyping products captures common Phase II SNPs with an average maximum r2 of up to 0.8 in African and up to 0.95 in non-African populations, and that potential gains in power in association studies can be obtained through imputation. These data also reveal novel aspects of the structure of linkage disequilibrium. We show that 10–30% of pairs of individuals within a population share at least one region of extended genetic identity arising from recent ancestry and that up to 1% of all common variants are untaggable, primarily because they lie within recombination hotspots. We show that recombination rates vary systematically around genes and between genes of different function. Finally, we demonstrate increased differentiation at non-synonymous, compared to synonymous, SNPs, resulting from systematic differences in the strength or efficacy of natural selection between populations. Advances made possible by the Phase I haplotype map In Phase II of the HapMap Project, a further 2.1 million SNPs The International HapMap Project was launched in 2002 with the were successfully genotyped on the same individuals.
    [Show full text]
  • Characterization of a Meiotic Recombination Hotspot in Arabidopsis Thaliana Hossein Khademian
    Characterization of a meiotic recombination hotspot in Arabidopsis thaliana Hossein Khademian To cite this version: Hossein Khademian. Characterization of a meiotic recombination hotspot in Arabidopsis thaliana. Agricultural sciences. Université Paris Sud - Paris XI, 2012. English. NNT : 2012PA112051. tel- 00800551 HAL Id: tel-00800551 https://tel.archives-ouvertes.fr/tel-00800551 Submitted on 14 Mar 2013 HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. UNIVERSITE PARIS-SUD 11 U.F.R. Scientifique d’Orsay Thèse Présentée pour l’obtention du grade de Docteur en Sciences de l’Université Paris-Sud XI Spécialité : Sciences du Végétal par Hossein KHADEMIAN Caractérisation d’un point chaud de recombinaison méiotique chez Arabidopsis thaliana Composition du jury : Valérie BORDE Rapporteur Michel DRON Président du Jury Corinne GREY Examinateur Christine MEZARD Directeur de Thèse Minoo RASSOULZADEGAN Rapporteur Abstract Meiotic recombination initiated in prophase I of meiosis generates either crossovers (COs), which are reciprocal exchanges between chromosome segments, or gene conversion not associated to crossovers (NCOs). Both kinds of events occur in narrow regions (less than 10 kilobases) called hotspots, which are distributed non-homogenously along chromosomes. The aim of my PhD was the characterization of a hotspot of meiotic recombination (named 14a) in Arabidopsis thaliana (i) across different accessions (ii) in msh4 mutant, a gene involved in CO formation.
    [Show full text]
  • Fine-Scale Recombination Landscapes Between a Freshwater and Marine Population of Threespine 4 Stickleback Fish 5 6 Alice F
    bioRxiv preprint doi: https://doi.org/10.1101/430249; this version posted September 29, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-ND 4.0 International license. 1 Research Article 2 3 Fine-scale recombination landscapes between a freshwater and marine population of threespine 4 stickleback fish 5 6 Alice F. Shanfelter1, Sophie L. Archambeault2,3, Michael A. White1* 7 8 1Department of Genetics, University of Georgia, Athens, GA, 30602, USA 9 2Institute of Ecology and Evolution, University of Bern, 3012 Bern, Switzerland 10 3Graduate Program in Molecular and Cellular Biology, University of Washington, Seattle, WA, 11 98195, USA 12 13 *Author for Correspondence: Michael White, Department of Genetics, University of Georgia, 14 Athens, USA, Voice: 706-542-2464, Fax: 706-542-3910, [email protected] 15 16 Data deposition: Raw sequences are deposited in NCBI’s Short Read Archive, reference number 17 SRP137809 (https://submit.ncbi.nlm.nih.gov/subs/sra/SUB3748706/overview). 18 19 Short title: Recombination hotspots in threespine stickleback fish 20 21 22 23 24 25 26 27 28 29 30 31 1 bioRxiv preprint doi: https://doi.org/10.1101/430249; this version posted September 29, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-ND 4.0 International license.
    [Show full text]
  • Meiotic Recombination and Chromosome Segregation in Schizosaccharomyces Pombe
    Colloquium Meiotic recombination and chromosome segregation in Schizosaccharomyces pombe Luther Davis and Gerald R. Smith* Fred Hutchinson Cancer Research Center, 1100 Fairview Avenue North, A1–162, Seattle, WA 98109-1024 In most organisms homologous recombination is vital for the ing studies of meiotic recombination-deficient (RecϪ) mutants. proper segregation of chromosomes during meiosis, the formation Biochemical analyses are aided by a mutant, described below, of haploid sex cells from diploid precursors. This review compares that undergoes rapid, synchronous meiosis when the tempera- meiotic recombination and chromosome segregation in the fission ture is raised. Finally, the nucleotide sequence of the S. pombe yeast Schizosaccharomyces pombe and the distantly related bud- genome is essentially complete (www.sanger.ac.uk͞projects͞ ding yeast Saccharomyces cerevisiae, two especially tractable mi- S_pombe), and the near isogenicity of the commonly used S. croorganisms. Certain features, such as the occurrence of DNA pombe strains aids comparisons between different studies. breaks associated with recombination, appear similar, suggesting S. pombe is only distantly related to the budding yeast Sac- that these features may be common in eukaryotes. Other features, charomyces cerevisiae, in which meiosis also has been extensively such as the role of these breaks and the ability of chromosomes to studied (1–3). Some features of meiosis are the same in the two segregate faithfully in the absence of recombination, appear yeasts, suggesting that they may be common among eukaryotes. different, suggesting multiple solutions to the problems faced in Other features, however, appear to be distinctly different and meiosis. illustrate the diversity of meiosis. In this review we shall em- phasize some of these similarities and differences.
    [Show full text]
  • Molecular Features of Meiotic Recombination Hot Spots K.T
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Publications of the IAS Fellows Review articles Molecular features of meiotic recombination hot spots K.T. Nishant1 and M.R.S. Rao1,2* Summary double-strand breaks (DSBs) by the Spo11 endonuclease that Meiotic recombination occurs preferentially at certain initiates recombination,(1) homolog pairing through DSB- regions called hot spots and is important for generating (2) genetic diversity and proper segregation of chromo- dependent and -independent mechanisms, formation of somes during meiosis. Hot spots have been character- the synaptonemal complex (SC) and chiasma between the ized most extensively in yeast, mice and humans. The homologs (Fig. 1). The temporal sequence of these events is development of methods based on sperm typing and likely to be different in D. melanogaster and C. elegans based population genetics has facilitated rapid and high- on the study of mutants of spo11 homologs in these organisms resolution mapping of hot spots in mice and humans in recent years. With increasing information becoming that eliminate DSB formation but allow normal SC formation available on meiotic recombination in different species, and synapsis. The repair of the DSBs that give rise to both it is now possible to compare several molecular features conversion and crossover events through distinct pathways associated with hot-spot loci. Further, there have been have been reviewed recently.(3) Meiotic homologous recombi- advances in our knowledge of the factors influencing hot- nation serves two basic functions. It resorts the linkages spot activity and the role that they play in structuring the genome into haplotype blocks.
    [Show full text]
  • Recombination Occurs Uniformly Within the Bronze Gene, a Meiotic Recombination Hotspot in the Maize Genome
    The Plant Cell, Vol. 9, 1633-1 646, September 1997 O 1997 American Society of Plant Physi&g@m Recombination Occurs Uniformly within the bronze Gene, a Meiotic Recombination Hotspot in the Maize Genome Hugo K. Doonerl and Isabel M. Martínez-Férez The Waksman Institute, Rutgers University, Piscataway, New Jersey 08855 The bronze (62)gene is a recombinational hotspot in the maize genome: its level of meiotic recombination per unit of physical length is >lOO-fold higher than the genome’s average and is the highest of any plant gene analyzed to date. Here, we examine whether recombination is also unevenly distributed within the bz gene. In yeast genes, recombina- tion (conversion) is polarized, being higher at the end of the gene where recombination is presumably initiated. We have analyzed products of meiotic recombination between heteroallelic pairs of bz mutations in both the presence and ab- sence of heterologies and have sequenced the recombination junction in 130 such Bz intragenic recombinants. We have found that in the absence of heterologies, recombination is proportional to physical distance across the bz gene. The simplest interpretation for this lack of polarity is that recombination is initiated randomly within the gene. lnsertion mutations affect the frequency and distribution of intragenic recombination events at bz, creating hotspots and cold- spots. Single base pair heterologies also affect recombination, with fewer recombination events than expected by chance occurring in regions of the bz gene with a high density of heterologies. We also provide evidence that meiotic recombination in maize is conservative, that is, it does not introduce changes, and that meiotic conversion tracts are continuous and similar in size to those in yeast.
    [Show full text]
  • A Family of Camp-Response-Element-Related DNA Sequences with Meiotic Recombination Hotspot Activity in Schizosaccharomyces Pombe
    Copyright 2000 by the Genetics Society of America A Family of cAMP-Response-Element-Related DNA Sequences With Meiotic Recombination Hotspot Activity in Schizosaccharomyces pombe Mary E. Fox,* Takatomi Yamada,² Kunihiro Ohta² and Gerald R. Smith* *Fred Hutchinson Cancer Research Center, Seattle, Washington 98109 and ²Genetic Dynamics Research Unit Laboratory, Institute of Physical and Chemical Research, Wako, Saitamo 351-01, Japan Manuscript received March 28, 2000 Accepted for publication May 15, 2000 ABSTRACT The heptamer sequence ATGACGT is essential for activity of the M26 meiotic recombination hotspot in the ade6 gene of Schizosaccharomyces pombe. Hotspot activity is associated with binding of the heterodimeric transcription factor Atf1´Pcr1 to M26. We have found that the sequences (C/T/G) TGACGT also bound Atf1´Pcr1 and acted as meiotic hotspots, but unlike M26 they must be followed by A or C for Atf1´Pcr1 binding and hotspot activity. The basis of the hotspot activity of CTGACGTA (ade6-3013) appears to be identical to that of M26: hotspot activity of both sequences was abolished in cells mutant for atf1, pcr1, spc1,orwis1 and was undetectable in mitotic recombination and in meiotic recombination when located on a plasmid. Both hotspot sequences were sites of micrococcal nuclease hypersensitivity in meiotic chromatin, suggesting that they create an open chromatin structure during meiosis at the site of the hotspots. The newly identi®ed hotspot sequences (C/T/G)TGACGT(A/C) and M26 are closely related to the cAMP response element (CRE) consensus sequence for binding of cAMP-responsive transcription factors such as Atf1´Pcr1, suggesting a link between transcription and meiotic recombination.
    [Show full text]
  • Recombination Hot Spots and Human Disease Smita M
    Downloaded from genome.cshlp.org on September 28, 2021 - Published by Cold Spring Harbor Laboratory Press REVIEW Recombination Hot Spots and Human Disease Smita M. Purandare1 and Pragna I. Patel1–4 Departments of 1Neurology, 2Molecular and Human Genetics and 3Division of Neuroscience, Baylor College of Medicine, Houston, Texas 77030 Recombination between homologous DNA se- phic VNTRs (variable number tandem repeats) or quences occurs in all organisms, and the resultant hypervariable minisatellite DNA, where the size of exchange of information is critical for the survival the repeat unit ranges from 9 to 24 bp. VNTRs have of species. Recombination is an essential cellular been shown to be hot spots for homologous recom- process catalyzed by proteins explicitly expressed bination in human cells (Wahls et al. 1990). Micro- for this purpose. It provides an effective means of satellite DNA consists of small arrays of tandem re- generating genetic diversity that is important for peats (usually 1–4 bp units) that are interspersed evolution. The proteins involved in recombination throughout the genome, in blocks consisting of allow cells to retrieve sequences lost when DNA is <150 bp. damaged by radiation or chemicals, by replacing the In contrast to tandemly repeated DNA, inter- damaged section with an undamaged strand from a spersed repetitive DNA consists of repeat units dis- homologous chromosome. The process of homolo- persed throughout the genome. Based on the repeat gous recombination has also been used to study unit length, two major classes are recognized: short gene function by way of gene knockouts. However, interspersed nuclear elements [(SINES) e.g., the Alu recombination and factors involved in recombina- repeat family] and long interspersed nuclear ele- tion may also be a source of harmful mutations and ments [(LINES) e.g., the LINE-1 or L1 element disease.
    [Show full text]
  • 24 Recombination Hotspots in Nonallelic Homologous Recombination
    Chapter 24 / NAHR Hotspots 341 24 Recombination Hotspots in Nonallelic Homologous Recombination Matthew E. Hurles, PhD and James R. Lupski, MD, PhD CONTENTS INTRODUCTION IDENTIFICATION OF RECOMBINATION HOTSPOTS FEATURES OF AHR AND NAHR HOTSPOTS MECHANISTIC BASIS OF RECOMBINATION HOTSPOTS EVOLUTIONARY ORIGINS OF HOTSPOTS CONCLUSIONS AND FUTURE WORK SUMMARY REFERENCES INTRODUCTION Rearrangement breakpoints resulting from nonallelic homologous recombination (NAHR) are typically clustered within small, well-defined portions of the segmental duplications that promote the rearrangement. These NAHR “hotspots” have been identified in every NAHR- promoted rearrangement in which breakpoint junctions have been sequenced in sufficient numbers. Enhancement of recombinatorial activity in NAHR hotspots varies from 3 to 237 times more than in the surrounding “cold” duplicated sequence. NAHR hotspots share many features in common with allelic homologous recombination (AHR) hotspots. Both AHR and NAHR hotspots appear to be relatively small (<2 kb) and are initiated by double-strand breaks. Gene conversion events as well as crossovers are enhanced at NAHR hotspots. Recent work has improved our understanding of the origins of NAHR and AHR hotspots, with both appear- ing to be relatively short-lived phenomena. Our present understanding of NAHR hotspots comes from a limited number of locus-specific studies. In the future, we can expect genome- wide analyses to provide many further insights. During meiosis, AHR occurs between homologous chromosomes, generating haplotypic diversity in the succeeding generation. The distribution of these recombination events through- out the genome has been known to be nonrandom for decades, but in the past 5 years ever-finer spatial resolutions have revealed dramatic heterogeneity in AHR rates at the DNA sequence level.
    [Show full text]