Evgenia Pak

National Human Genome Research Institute

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Amalia Dutra

National Human Genome Research Institute

Svetlana Pack

National Institutes of Health

Zhengping Zhuang

National Institutes of Health

Robert J. Weil

SouthCoast Health

A. Dutra

Universidade Federal do Estado do Rio de Janeiro

Stephen Wincovitch

National Human Genome Research Institute

Constantine A. Stratakis

Eunice Kennedy Shriver National Institute of Child Health and Human Development

Alexander O. Vortmeyer

Indiana University School of Medicine

Maximilian Muenke

National Institutes of Health

Evan E. Eichler

Howard Hughes Medical Institute

Cooperative Institutions

National Institutes of Health

312

National Human Genome Research Institute

117

National Cancer Institute

National Institute of Neurological Disorders and Stroke

Cancer Genetics (United States)

Johns Hopkins University

Center for Cancer Research

University of Washington

National Institute of Mental Health

National Heart Lung and Blood Institute

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The complete sequence and comparative analysis of ape sex chromosomes

Nature (2024)

Kateryna D. Makova Brandon D. Pickett Robert S. Harris Gabrielle A. Hartley Monika Čechová

Abstract Apes possess two sex chromosomes—the male-specific Y chromosome and the X chromosome, which is present in both males and females. The Y chromosome is crucial for male reproduction, with deletions being linked to infertility 1 . The X chromosome is vital for reproduction and cognition 2 . Variation in mating patterns and brain function among apes suggests corresponding differences in their sex chromosomes. However, owing to their repetitive nature and incomplete reference assemblies, ape sex chromosomes have been challenging to study. Here, using the methodology developed for the telomere-to-telomere (T2T) human genome, we produced gapless assemblies of the X and Y chromosomes for five great apes (bonobo ( Pan paniscus ), chimpanzee ( Pan troglodytes ), western lowland gorilla ( Gorilla gorilla gorilla ), Bornean orangutan ( Pongo pygmaeus ) and Sumatran orangutan ( Pongo abelii )) and a lesser ape (the siamang gibbon ( Symphalangus syndactylus )), and untangled the intricacies of their evolution. Compared with the X chromosomes, the ape Y chromosomes vary greatly in size and have low alignability and high levels of structural rearrangements—owing to the accumulation of lineage-specific ampliconic regions, palindromes, transposable elements and satellites. Many Y chromosome genes expand in multi-copy families and some evolve under purifying selection. Thus, the Y chromosome exhibits dynamic evolution, whereas the X chromosome is more stable. Mapping short-read sequencing data to these assemblies revealed diversity and selection patterns on sex chromosomes of more than 100 individual great apes. These reference assemblies are expected to inform human evolution and conservation genetics of non-human apes, all of which are endangered species.

Gorilla

Pan paniscus

Bonobo

Hominidae

Lineage (genetic)

Pongo pygmaeus

10.1038/s41586-024-07473-2

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Citations (46)

The human inactive X chromosome modulates expression of the active X chromosome

bioRxiv (Cold Spring Harbor Laboratory) (2021)

Adrianna K. San Roman Alexander K. Godfrey Helen Skaletsky Daniel W. Bellott Abigail F. Groff

Summary Researchers have presumed that the “inactive” X chromosome (Xi) has little impact, in trans, on the “active” X (Xa). To test this, we quantified Xi and Xa gene expression in individuals with one Xa and zero to three Xi’s. Our linear modeling revealed modular Xi and Xa transcriptomes and significant Xi-driven expression changes for 38% (162/423) of expressed X-chromosome genes. By integrating allele-specific analyses, we found that modulation of Xa transcript levels by Xi contributes to many of these Xi-driven changes (≥ 121 genes). By incorporating metrics of evolutionary constraint, we identified 10 X-chromosome genes most likely to drive sex differences in common disease, and sex chromosome aneuploidy syndromes. We conclude that human X chromosomes are regulated both in cis, through Xi-wide transcriptional attenuation, and in trans, through positive or negative modulation of individual Xa genes by Xi. The sum of cis and trans effects differs widely among genes.

10.1101/2021.08.09.455676

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Multiomic Profiling Identifies cis-Regulatory Networks Underlying Human Pancreatic β Cell Identity and Function

Cell Reports (2019)

Nathan Lawlor Eladio J. Márquez Peter Orchard Narisu Narisu Muhammad S. Shamim

EndoC-βH1 is emerging as a critical human β cell model to study the genetic and environmental etiologies of β cell (dys)function and diabetes. Comprehensive knowledge of its molecular landscape is lacking, yet required, for effective use of this model. Here, we report chromosomal (spectral karyotyping), genetic (genotyping), epigenomic (ChIP-seq and ATAC-seq), chromatin interaction (Hi-C and Pol2 ChIA-PET), and transcriptomic (RNA-seq and miRNA-seq) maps of EndoC-βH1. Analyses of these maps define known (e.g., PDX1 and ISL1) and putative (e.g., PCSK1 and mir-375) β cell-specific transcriptional cis-regulatory networks and identify allelic effects on cis-regulatory element use. Importantly, comparison with maps generated in primary human islets and/or β cells indicates preservation of chromatin looping but also highlights chromosomal aberrations and fetal genomic signatures in EndoC-βH1. Together, these maps, and a web application we created for their exploration, provide important tools for the design of experiments to probe and manipulate the genetic programs governing β cell identity and (dys)function in diabetes.

Epigenomics

PDX1

RNA-Seq

Chromosome conformation capture

10.1016/j.celrep.2018.12.083

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Citations (78)

Juvenile myelomonocytic leukemia due to a germline CBL Y371C mutation: 35-year follow-up of a large family

Human Genetics (2015)

Anand Pathak Alexander Pemov Mary L. McMaster Ramita Dewan Sarangan Ravichandran

Juvenile myelomonocytic leukemia

Penetrance

Myeloproliferative neoplasm

10.1007/s00439-015-1550-9

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Citations (23)

Differentiation of human bone marrow-derived cells into buccal epithelial cells in vivo: a molecular analytical study

The Lancet (2003)

Simon D. Tran Stanley R. Pillemer Amalia Dutra A. John Barrett Michael Brownstein

Transdifferentiation

10.1016/s0140-6736(03)12894-2

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Citations (184)

Construction of a Human-BAC Array for a high resolution analysis of genomic changes in cancer

Cancer Research (2006)

Virginia Rodriguez Abdel Elkahloun Amalia Dutra Evgenia Pak Settara C. Chandrasekharappa

Proc Amer Assoc Cancer Res, Volume 47, 2006 169 A high density BAC array of the human genome will be very useful to assess global genomic DNA changes in cancer cells and deletion syndromes at a very high resolution using CGH (comparative genomic hybridization) techniques. We are generating an array of over 32,000 BAC clones to give comprehensive coverage of the genome. To begin this process on a small scale, a BAC array of chromosome 8 was developed and tested. This array contains 1463 BAC clones creating a tiling path along the chromosome 8 and includes 48 BAC clones from chromosome X. The BAC clones DNA were amplified using the Phi29 DNA polymerase to generate the required amounts; a modified protocol allowed for efficient amplification of large number of BAC clones. Labeling/hybridization protocols were optimized to detect single copy changes using a panel of cell lines with an increasing number of X chromosomes. DNA from a breast cancer cell line (SKBR3) with two known amplified regions in 8q21 and 8q24 was hybridized to evaluate the usefulness of this high density BAC array. We identified 6 distinct amplified and 3 deleted regions in 8q. Each of these regions was verified by q-PCR (quantitative PCR) and FISH (fluorescence in situ hybridization). These techniques were also used to define the precise boundaries of amplicons in an independent manner. In addition, expression of genes that were part of the amplified/deleted regions was evaluated by expression arrays and q-PCR. Similar high-resolution amplification/deletion maps of chromosome 8 of several pancreatic cancer cell lines are generated. The entire 32,000 BAC clone array is now being assembled, and it is anticipated that a high-resolution analysis will help identify hitherto unknown genomic changes that will lead to a better understanding of cancer.

Amplicon

Comparative genomic hybridization

Bacterial artificial chromosome

genomic DNA

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Advantages of q-PCR as a method of screening for gene targeting in mammalian cells using conventional and whole BAC-based constructs

Nucleic Acids Research (2008)

Julio Gómez‐Rodríguez Valance Washington Jun Cheng Amalia Dutra Evgenia Pak

We evaluate here the use of real-time quantitative PCR (q-PCR) as a method for screening for homologous recombinants generated in mammalian cells from either conventional gene-targeting constructs or whole BAC-based constructs. Using gene-targeted events at different loci, we show that q-PCR is a highly sensitive and accurate method for screening for conventional gene targeting that can reduce the number of clones requiring follow-up screening by Southern blotting. We further compared q-PCR to fluorescent in situ hybridization (FISH) for the detection of gene-targeting events using full-length BAC-based constructs designed to introduce mutations either into one gene or simultaneously into two adjacent genes. We find that although BAC-based constructs appeared to have high rates of homologous recombination when evaluated by FISH, screening by FISH was prone to false positives that were detected by q-PCR. Our results demonstrate the utility of q-PCR as a screening tool for gene targeting and further highlight potential problems with the use of whole BAC-based constructs for homologous recombination.

10.1093/nar/gkn523

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Citations (12)

The human Y and inactive X chromosomes similarly modulate autosomal gene expression

bioRxiv (Cold Spring Harbor Laboratory) (2023)

Adrianna K. San Roman Helen Skaletsky Alexander K. Godfrey Neha V. Bokil Levi S. Teitz

Somatic cells of human males and females have 45 chromosomes in common, including the "active" X chromosome. In males the 46

10.1101/2023.06.05.543763

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Citations (8)

Regulation of Telomere Length and Suppression of Genomic Instability in Human Somatic Cells by Ku86

Molecular and Cellular Biology (2004)

Kyungjae Myung Goutam Ghosh Farjana Fattah Gang Li Haeyoung Kim

Ku86 plays a key role in nonhomologous end joining in organisms as evolutionarily disparate as bacteria and humans. In eukaryotic cells, Ku86 has also been implicated in the regulation of telomere length although the effect of Ku86 mutations varies considerably between species. Indeed, telomeres either shorten significantly, shorten slightly, remain unchanged, or lengthen significantly in budding yeast, fission yeast, chicken cells, or plants, respectively, that are null for Ku86 expression. Thus, it has been unclear which model system is most relevant for humans. We demonstrate here that the functional inactivation of even a single allele of Ku86 in human somatic cells results in profound telomere loss, which is accompanied by an increase in chromosomal fusions, translocations, and genomic instability. Together, these experiments demonstrate that Ku86, separate from its role in nonhomologous end joining, performs the additional function in human somatic cells of suppressing genomic instability through the regulation of telomere length.

Non-homologous end joining

10.1128/mcb.24.11.5050-5059.2004

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Citations (107)

The Chapel Hill hemophilia A dog colony exhibits a factor VIII gene inversion

Proceedings of the National Academy of Sciences (2002)

Jay N. Lozier Amalia Dutra Evgenia Pak Nan Zhou Zhili Zheng

In the Chapel Hill colony of factor VIII-deficient dogs, abnormal sequence (ch8, for canine hemophilia 8, GenBank no. ) follows exons 1-22 in the factor VIII transcript in place of exons 23-26. The canine hemophilia 8 locus (ch8) sequence was found in a 140-kb normal dog genomic DNA bacterial artificial chromosome (BAC) clone that was completely outside the factor VIII gene, but not in BAC clones containing the factor VIII gene. The BAC clone that contained ch8 also contained a homologue of F8A (factor 8 associated) sequence, which participates in a common inversion that causes severe hemophilia A in humans. Fluorescence in situ hybridization analysis indicated that exons 1-26 normally proceed sequentially from telomere to centromere at Xq28, and ch8 is telomeric to the factor VIII gene. The appearance of an "upstream" genomic sequence element (ch8) at the end of the aberrant factor VIII transcript suggested that an inversion of genomic DNA replaced factor VIII exons 22-26 with ch8. The F8A sequence appeared also in overlapping normal BAC clones containing factor VIII sequence. We hypothesized that homologous recombination between copies of canine F8A inside and outside the factor VIII gene had occurred, as in human hemophilia A. High-resolution fluorescent in situ hybridization on hemophilia A dog DNA revealed a pattern consistent with this inversion mechanism. We also identified a HindIII restriction fragment length polymorphism of F8A fragments that distinguished hemophilia A, carrier, and normal dogs' DNA. The Chapel Hill hemophilia A dog colony therefore replicates the factor VIII gene inversion commonly seen in humans with severe hemophilia A.

Bacterial artificial chromosome

genomic DNA

10.1073/pnas.192219599

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Citations (99)