Biomedical subjects

Werner Schempp

Publications and source records attributed to Werner Schempp.

12 recordsLinked to original sources

PubMed2006

Complex patterns of copy number variation at sites of segmental duplications: an important category of structural variation in the human genome.

The structural diversity of the human genome is much higher than previously assumed although its full extent remains unknown. To investigate the association between segmental duplications that display constitutive copy number differences (CNDs) between humans and the great apes and those which exhibit polymorphic copy number variations (CNVs) between humans, we analysed a BAC array enriched with segmental duplications displaying such CNDs. This study documents for the first time that in addition to human-specific gains common to all humans, these duplication clusters (DCs) also exhibit polymorphic CNVs > 40 kb. Segmental duplication is known to have been a frequent event during human genome evolution. Importantly, among the CNV-associated genes identified here, those involved in transcriptional regulation were found to be significantly overrepresented. Complex patterns of variation were evident at sites of DCs, manifesting as inter-individual differentially sized copy number alterations at the same genomic loci. Thus, CNVs associated with segmental duplications do not simply represent insertion/deletion polymorphisms, but rather constitute a wide variety of rearrangements involving differential amplification and partial gains and losses with high inter-individual variability. Although the number of CNVs was not found to differ between Africans and Caucasians/Asians, the average number of variant patterns per locus was significantly lower in Africans. Thus, complex variation patterns characterizing segmental duplications result from relatively recent genomic rearrangements. The high number of these rearrangements, some of which are potentially recurrent, together with differences in population size and expansion dynamics, may account for the greater diversity of CNV in Caucasians/Asians as compared with Africans.

Animals↗

PubMed2006

Identification of large-scale human-specific copy number differences by inter-species array comparative genomic hybridization.

Copy number differences (CNDs), and the concomitant differences in gene number, have contributed significantly to the genomic divergence between humans and other primates. To assess its relative importance, the genomes of human, common chimpanzee, bonobo, gorilla, orangutan and macaque were compared by comparative genomic hybridization using a high-resolution human BAC array (aCGH). In an attempt to avoid potential interference from frequent intra-species polymorphism, pooled DNA samples were used from each species. A total of 322 sites of large-scale inter-species CND were identified. Most CNDs were lineage-specific but frequencies differed considerably between the lineages; the highest CND frequency among hominoids was observed in gorilla. The conserved nature of the orangutan genome has already been noted by karyotypic studies and our findings suggest that this degree of conservation may extend to the sub-microscopic level. Of the 322 CND sites identified, 14 human lineage-specific gains were observed. Most of these human-specific copy number gains span regions previously identified as segmental duplications (SDs) and our study demonstrates that SDs are major sites of CND between the genomes of humans and other primates. Four of the human-specific CNDs detected by aCGH map close to the breakpoints of human-specific karyotypic changes [e.g., the human-specific inversion of chromosome 1 and the polymorphic inversion inv(2)(p11.2q13)], suggesting that human-specific duplications may have predisposed to chromosomal rearrangement. The association of human-specific copy number gains with chromosomal breakpoints emphasizes their potential importance in mediating karyotypic evolution as well as in promoting human genomic diversity.

Animals↗

PubMed2005

Polymorphic micro-inversions contribute to the genomic variability of humans and chimpanzees.

A combination of inter- and intra-species genome comparisons is required to identify and classify the full spectrum of genetic changes, both subtle and gross, that have accompanied the evolutionary divergence of humans and other primates. In this study, gene order comparisons of 11,518 human and chimpanzee orthologous gene pairs were performed to detect regions of inverted gene order that are potentially indicative of small-scale rearrangements such as inversions. By these means, a total of 71 potential micro-rearrangements were detected, nine of which were considered to represent micro-inversions encompassing more than three genes. These putative inversions were then investigated by FISH and/or PCR analyses and the authenticity of five of the nine inversions, ranging in size from approximately 800 kb to approximately 4.4 Mb, was confirmed. These inversions mapped to 1p13.2-13.3, 7p22.1, 7p13-14.1, 18p11.21-11.22 and 19q13.12 and encompass 50, 14, 16, 7 and 16 known genes, respectively. Intriguingly, four of the confirmed inversions turned out to be polymorphic: three were polymorphic in the chimpanzee and one in humans. It is concluded that micro-inversions make a significant contribution to genomic variability in both humans and chimpanzees and inversion polymorphisms may be more frequent than previously realized.

Animals↗

PubMed2005

Interchromosomal segmental duplications of the pericentromeric region on the human Y chromosome.

Basic medical research critically depends on the finished human genome sequence. Two types of gaps are known to exist in the human genome: those associated with heterochromatic sequences and those embedded within euchromatin. We identified and analyzed a euchromatic island within the pericentromeric repeats of the human Y chromosome. This 450-kb island, although not recalcitrant to subcloning and present in 100 tested males from different ethnic origins, was not detected and is not contained within the published Y chromosomal sequence. The entire 450-kb interval is almost completely duplicated and consists predominantly of interchromosomal rather than intrachromosomal duplication events that are usually prevalent on the Y chromosome. We defined the modular structure of this interval and detected a total of 128 underlying pairwise alignments (>/=90% and >/=1 kb in length) to various autosomal pericentromeric and ancestral pericentromeric regions. We also analyzed the putative gene content of this region by a combination of in silico gene prediction and paralogy analysis. We can show that even in this exceptionally duplicated region of the Y chromosome, eight putative genes with open reading frames reside, including fusion transcripts formed by the splicing of exons from two different duplication modules as well as members of the homeobox gene family DUX.

Amino Acid Sequence↗

PubMed2005

Polymorphisms associated with the DAZ genes on the human Y chromosome.

The human Y chromosome is unique in that it does not engage in pairing and crossing over during meiosis for most of its length. Y chromosome microdeletions, a frequent finding in infertile men, thus occur through intrachromosomal recombination, either within a single chromatid or between sister chromatids. A recently identified polymorphism associated with increased risk for spermatogenic failure, the gr/gr deletion, removes two of the four Deleted in Azoospermia (DAZ) genes in the AZFc region on the Y-chromosome long arm. We found the likely reciprocal duplication product of gr/gr deletion in 5 (6%) of 82 males using a novel DNA-blot hybridization strategy and confirmed the presence of six DAZ genes in three cases by FISH analysis. Additional polymorphisms identified within the DAZ repeat regions of the DAZ genes indicate that sister chromatid exchange plays a significant role in the genesis of deletions, duplications, and polymorphisms of the Y chromosome.

Alleles↗

PubMed2005

Human endogenous retrovirus HERV-K14 families: status, variants, evolution, and mobilization of other cellular sequences.

The human genome harbors many distinct families of human endogenous retroviruses (HERVs) that stem from exogenous retroviruses that infected the germ line millions of years ago. Many HERV families remain to be investigated. We report in the present study the detailed characterization of the HERV-K14I and HERV-K14CI families as they are represented in the human genome. Most of the 68 HERV-K14I and 23 HERV-K14CI proviruses are severely mutated, frequently displaying uniform deletions of retroviral genes and long terminal repeats (LTRs). Both HERV families entered the germ line approximately 39 million years ago, as evidenced by homologous sequences in hominoids and Old World primates and calculation of evolutionary ages based on a molecular clock. Proviruses of both families were formed during a brief period. A majority of HERV-K14CI proviruses on the Y chromosome mimic a higher evolutionary age, showing that LTR-LTR divergence data can indicate false ages. Fully translatable consensus sequences encoding major retroviral proteins were generated. Most HERV-K14I loci lack an env gene and are structurally reminiscent of LTR retrotransposons. A minority of HERV-K14I variants display an env gene. HERV-K14I proviruses are associated with three distinct LTR families, while HERV-K14CI is associated with a single LTR family. Hybrid proviruses consisting of HERV-K14I and HERV-W sequences that appear to have produced provirus progeny in the genome were detected. Several HERV-K14I proviruses harbor TRPC6 mRNA portions, exemplifying mobilization of cellular transcripts by HERVs. Our analysis contributes essential information on two more HERV families and on the biology of HERV sequences in general.

Base Sequence↗

PubMed2004

An excess of chromosome 1 breakpoints in male infertility.

In a search for potential infertility loci, which might be revealed by clustering of chromosomal breakpoints, we compiled 464 infertile males with a balanced rearrangement from Mendelian Cytogenetics Network database (MCNdb) and compared their karyotypes with those of a Danish nation-wide cohort. We excluded Robertsonian translocations, rearrangements involving sex chromosomes and common variants. We identified 10 autosomal bands, five of which were on chromosome 1, with a large excess of breakpoints in the infertility group. Some of these could potentially harbour a male-specific infertility locus. However, a general excess of breakpoints almost everywhere on chromosome 1 was observed among the infertile males: 26.5 versus 14.5% in the cohort. This excess was observed both for translocation and inversion carriers, especially pericentric inversions, both for published and unpublished cases, and was significantly associated with azoospermia. The largest number of breakpoints was reported in 1q21; FISH mapping of four of these breakpoints revealed that they did not involve the same region at the molecular level. We suggest that chromosome 1 harbours a critical domain whose integrity is essential for male fertility.

Chromosome Aberrations↗

PubMed2004

Human endogenous retrovirus family HERV-K(HML-5): status, evolution, and reconstruction of an ancient betaretrovirus in the human genome.

The human genome harbors numerous distinct families of so-called human endogenous retroviruses (HERV) which are remnants of exogenous retroviruses that entered the germ line millions of years ago. We describe here the hitherto little-characterized betaretrovirus HERV-K(HML-5) family (named HERVK22 in Repbase) in greater detail. Out of 139 proviruses, only a few loci represent full-length proviruses, and many lack gag protease and/or env gene regions. We generated a consensus sequence from multiple alignment of 62 HML-5 loci that displays open reading frames for the four major retroviral proteins. Four HML-5 long terminal repeat (LTR) subfamilies were identified that are associated with monophyletic proviral bodies, implying different evolution of HML-5 LTRs and genes. Sequence analysis indicated that the proviruses formed approximately 55 million years ago. Accordingly, HML-5 proviral sequences were detected in Old World and New World primates but not in prosimians. No recent activity is associated with this HERV family. We also conclude that the HML-5 consensus sequence primer binding site is identical to methionine tRNA. Therefore, the family should be designated HERV-M. Our study provides important insights into the structure and evolution of the oldest betaretrovirus in the primate genome known to date.

Animals↗

PubMed2002

The 48 bp centromeric repeat is a functionally conserved motif in great apes and man showing protein-binding properties.

The centromere-kinetochore complex is a chromosomal assembly site including repeat motifs and protein binding properties thus mediating chromosome motility and mitotic regulation. Next to the alpha-satellite DNA family as well as human satellite III DNA, contribution of other repetitive sequences has to be strongly considered in centromere function. Here, we report the identification of centromeric 48 bp motifs, isolated from chimpanzee and orang-utan using an orthologous human DNA probe. Applying Southern hybridization we show that these sequences are restricted to hominoid species. Diminishing hybrid formation in gibbons suggested that the 48 bp repeat originated approximately 25-20 million years ago. Consistently, both chimpanzee as well as human repeat probes failed to generate any hybridization signal with the monkey species Cercopithecus aethiops and Aotes trivirgatus. Sequence deviations from the consensus of human repeat monomers of 6% and 10.4% in chimpanzee and orang-utan, respectively, were found within a 16 bp region of the 48 bp repeat units. Gel mobility shift assays using chimpanzee repeat dimers as probes revealed peptide-binding properties with human and chimpanzee nuclear extracts. Species-specific DNA-protein complexes remained unaffected by competition studies and indicated the presence of at least one novel interacting protein consisting of two subunits with 90 and 95 kDa. Our data suggest that the 48 bp repeat, next to alpha-satellite DNA, provides essential sequence information for specific DNA-protein interaction and they imply phylogenetic conservation of these binding properties in primates. The complex is likely involved in the proper formation and/or function of mammalian centromeres.

Animals↗

PubMed2002

Direct evidence for the Homo-Pan clade.

For a long time, the evolutionary relationship between human and African apes, the 'trichotomy problem', has been debated with strong differences in opinion and interpretation. Statistical analyses of different molecular DNA data sets have been carried out and have primarily supported a Homo-Pan clade. An alternative way to address this question is by the comparison of evolutionarily relevant chromosomal breakpoints. Here, we made use of a P1-derived artificial chromosome (PAC)/bacterial artificial chromosome (BAC) contig spanning approximately 2.8 Mb on the long arm of the human Y chromosome, to comparatively map individual PAC clones to chromosomes from great apes, gibbons, and two species of Old World monkeys by fluorescence in-situ hybridization. During our search for evolutionary breakpoints on the Y chromosome, it transpired that a transposition of an approximately 100-kb DNA fragment from chromosome 1 onto the Y chromosome must have occurred in a common ancestor of human, chimpanzee and bonobo. Only the Y chromosomes of these three species contain the chromosome-1-derived fragment; it could not be detected on the Y chromosomes of gorillas or the other primates examined. Thus, this shared derived (synapomorphic) trait provides clear evidence for a Homo-Pan clade independent of DNA sequence analysis.

Animals↗

PubMed2002

A fiber-FISH contig spanning the non-recombining region of the human Y chromosome.

Using fluorescence in-situ hybridization on interphase chromatin fibers (fiber-FISH), we have constructed an overlapping fiber-FISH contig spanning the non-recombining region of the human Y chromosome (NRY). We first established a standard FISH-signal pattern for a distinct panel of DNA clones on prometaphase Y chromosomes in six healthy fertile men. Clones in the panel were selected from all R-bands as well as deletion intervals 1 through 7 plus PAR1 and PAR2 of the human Y chromosome. We next used signals of these marker clones to build a fiber-FISH contig for the multicopy gene families, CDY, DAZ, RBMY, TSPY and XKRY, along the NRY. Our fiber-FISH contig of human NRY may help to close the four gaps that still exist in the current physical map of the human Y chromosome. Furthermore, it provides a more complete picture with respect to the positions and arrangements of the multicopy gene families along the human NRY.

Chromatin↗

PubMed2002

Identification and characterization of the cynomolgus monkey chromodomain gene cynCDY, an orthologue of the human CDY gene family.

Microdeletions within the AZF (azoospermia factor) a, b and c regions of the Y chromosome can be detected worldwide in 1-10% of infertile men. AZFc, containing genes such as DAZ, CDY, RBMY and others, is most frequently deleted and associated with oligo- or azoospermia. The function of the different genes within AZFc is not yet understood. Here we report the identification and first characterization of the cynomolgus monkey (Macaca fascicularis) homologue of the human CDY gene. cynCDY encodes a 541 aa protein, which like human CDY possesses two putative functional domains: an N-terminal chromodomain, possibly involved in heterochromatin interactions, and a C-terminal domain showing similarity to enoyl-CoA-isomerase, which is involved in fatty acid oxidation. Northern analysis and in-situ hybridization experiments revealed testis- and stage-specific expression of cynCDY mRNA, mainly confined to round and elongating spermatids. Fluorescence in-situ hybridization (FISH) performed on monkey metaphase chromosomes displayed exclusively Y-specific signals in Yq12.1. Using fibre FISH, short signal stretches that indicate the presence of three CDY copies could be visualized, although their integrity or function remains unknown. cynCDY is similar to human CDY with features of a retrotransposon, but different in the 3'UTR. It seems to represent a more ancestral form of CDY and its characterization yields insights into the evolution of candidate genes for AZF.

Amino Acid Sequence↗