Functional epialleles at an endogenous human centromere

Human centromeres are defined by megabases of homogenous alpha-satellite DNA arrays that are packaged into specialized chromatin marked by the centromeric histone variant, centromeric protein A (CENP-A). Although most human chromosomes have a single higher-order repeat (HOR) array of alpha satellites, several chromosomes have more than one HOR array. Homo sapiens chromosome 17 (HSA17) has two juxtaposed HOR arrays, D17Z1 and D17Z1-B. Only D17Z1 has been linked to CENP-A chromatin assembly. Here, we use human artificial chromosome assembly assays to show that both D17Z1 and D17Z1-B can support de novo centromere assembly independently. We extend these in vitro studies and demonstrate, using immunostaining and chromatin analyses, that in human cells the centromere can be assembled at D17Z1 or D17Z1-B. Intriguingly, some humans are functional heterozygotes, meaning that CENP-A is located at a different HOR array on the two HSA17 homologs. The site of CENP-A assembly on HSA17 is stable and is transmitted through meiosis, as evidenced by inheritance of CENP-A location through multigenerational families. Differences in histone modifications are not linked clearly with active and inactive D17Z1 and D17Z1-B arrays; however, we detect a correlation between the presence of variant repeat units of D17Z1 and CENP-A assembly at the opposite array, D17Z1-B. Our studies reveal the presence of centromeric epialleles on an endogenous human chromosome and suggest genomic complexities underlying the mechanisms that determine centromere identity in humans.

[1]  G. Vance,et al.  Cancer-associated alteration of pericentromeric heterochromatin may contribute to chromosome instability , 2012, Oncogene.

[2]  T. Fukagawa,et al.  The ABCs of CENPs , 2011, Chromosoma.

[3]  B. Sullivan,et al.  Genomic size of CENP-A domain is proportional to total alpha satellite array size at human centromeres and expands in cancer cells , 2011, Chromosome Research.

[4]  V. Rakyan,et al.  The hunt for the epiallele , 2011, Environmental and molecular mutagenesis.

[5]  Hiroshi Kimura,et al.  Epigenetic engineering shows H3K4me2 is required for HJURP targeting and CENP-A assembly on a synthetic human kinetochore , 2010, The EMBO journal.

[6]  A. Jauch,et al.  Telomere Disruption Results in Non-Random Formation of De Novo Dicentric Chromosomes Involving Acrocentric Human Chromosomes , 2010, PLoS genetics.

[7]  B. Sullivan,et al.  Histone Modifications within the Human X Centromere Region , 2009, PloS one.

[8]  F. Hubé,et al.  Non-coding murine centromeric transcripts associate with and potentiate Aurora B kinase , 2009, Nucleic acids research.

[9]  Yusuke Nakamura,et al.  HJURP Is a Cell-Cycle-Dependent Maintenance and Deposition Factor of CENP-A at Centromeres , 2009, Cell.

[10]  J. Yates,et al.  Centromere-Specific Assembly of CENP-A Nucleosomes Is Mediated by HJURP , 2009, Cell.

[11]  V. Noskov,et al.  Inactivation of a Human Kinetochore by Specific Targeting of Chromatin Modifiers , 2008, Developmental cell.

[12]  Takeshi Urano,et al.  Heterochromatin and RNAi Are Required to Establish CENP-A Chromatin at Centromeres , 2008, Science.

[13]  H. Masumoto,et al.  CENP-B Controls Centromere Formation Depending on the Chromatin Context , 2007, Cell.

[14]  R. Martienssen,et al.  Transposable elements and the epigenetic regulation of the genome , 2007, Nature Reviews Genetics.

[15]  P. Ridgway,et al.  H2A.Z contributes to the unique 3D structure of the centromere , 2007, Proceedings of the National Academy of Sciences.

[16]  S. Henikoff,et al.  Human centromeric chromatin is a dynamic chromosomal domain that can spread over noncentromeric DNA. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[17]  D. Murphy,et al.  Simultaneous genotyping of four functional loci of human SLC6A4, with a reappraisal of 5-HTTLPR and rs25531 , 2006, Molecular Psychiatry.

[18]  H. Willard,et al.  Analysis of the centromeric regions of the human genome assembly. , 2004, Trends in genetics : TIG.

[19]  H. Willard,et al.  Assembly and characterization of heterochromatin and euchromatin on human artificial chromosomes , 2004, Genome Biology.

[20]  G. Karpen,et al.  Centromeric chromatin exhibits a histone modification pattern that is distinct from both euchromatin and heterochromatin , 2004, Nature Structural &Molecular Biology.

[21]  Huntington F. Willard,et al.  Interhomologue sequence variation of alpha satellite DNA from human chromosome 17: Evidence for concerted evolution along haplotypic lineages , 1995, Journal of Molecular Evolution.

[22]  S. Schwartz,et al.  Human Artificial Chromosomes with Alpha Satellite-Based De Novo Centromeres Show Increased Frequency of Nondisjunction and Anaphase Lag , 2003, Molecular and Cellular Biology.

[23]  H. Willard,et al.  Alpha-satellite DNA and vector composition influence rates of human artificial chromosome formation. , 2002, Molecular therapy : the journal of the American Society of Gene Therapy.

[24]  Gary H Karpen,et al.  Conserved organization of centromeric chromatin in flies and humans. , 2002, Developmental cell.

[25]  U. Storb,et al.  An efficient method for high-fidelity BAC/PAC retrofitting with a selectable marker for mammalian cell transfection. , 2001, Genome research.

[26]  K. Yoda,et al.  Human centromere protein A (CENP-A) can replace histone H3 in nucleosome reconstitution in vitro. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[27]  K. Yoda,et al.  In vitro assembly of the CENP‐B/α‐satellite DNA/core histone complex: CENP‐B causes nucleosome positioning , 1998, Genes to cells : devoted to molecular & cellular mechanisms.

[28]  H. Masumoto,et al.  Construction of YAC–based mammalian artificial chromosomes , 1998, Nature Biotechnology.

[29]  J. Dudar,et al.  Technical note: improved DNA extraction from ancient bones using silica-based spin columns. , 1998, American journal of physical anthropology.

[30]  C. Tyler-Smith,et al.  Immunolocalization of CENP-A suggests a distinct nucleosome structure at the inner kinetochore plate of active centromeres , 1997, Current Biology.

[31]  H. Willard,et al.  Formation of de novo centromeres and construction of first-generation human artificial microchromosomes , 1997, Nature Genetics.

[32]  Nihon Hassei Seibutsu Gakkai,et al.  Genes to cells , 1996 .

[33]  K. Sullivan,et al.  Human CENP-A contains a histone H3 related histone fold domain that is required for targeting to the centromere , 1994, The Journal of cell biology.

[34]  H. Willard,et al.  Nonrandom localization of recombination events in human alpha satellite repeat unit variants: implications for higher-order structural characteristics within centromeric heterochromatin , 1993, Molecular and cellular biology.

[35]  H. Willard,et al.  PCR amplification of chromosome-specific alpha satellite DNA: definition of centromeric STS markers and polymorphic analysis. , 1991, Genomics.

[36]  H. Willard,et al.  Physical map of the centromeric region of human chromosome 7: relationship between two distinct alpha satellite arrays. , 1991, Nucleic acids research.

[37]  H. Willard,et al.  Genomic analysis of sequence variation in tandemly repeated DNA. Evidence for localized homogeneous sequence domains within arrays of alpha-satellite DNA. , 1990, Journal of molecular biology.

[38]  B. Vissel,et al.  Identification of two distinct subfamilies of alpha satellite DNA that are highly specific for human chromosome 15. , 1990, Genomics.

[39]  R. Margolis,et al.  Biochemical analysis of CENP-A, a centromeric protein with histone-like properties. , 1989, Progress in clinical and biological research.

[40]  H. Willard,et al.  Molecular organization and haplotype analysis of centromeric DNA from human chromosome 17: implications for linkage in neurofibromatosis. , 1987, Genomics.

[41]  H. Willard,et al.  Structure, organization, and sequence of alpha satellite DNA from human chromosome 17: evidence for evolution by unequal crossing-over and an ancestral pentamer repeat shared with the human X chromosome , 1986, Molecular and cellular biology.