Publications by authors named "Vanessa L Horner"

14 Publications

  • Page 1 of 1

Chromosomal instability upregulates interferon in acute myeloid leukemia.

Genes Chromosomes Cancer 2020 11 18;59(11):627-638. Epub 2020 Jul 18.

Department of Medicine, Division of Hematology/Oncology, School of Medicine and Public Health, University of Wisconsin, Madison, Wisconsin, USA.

Chromosome instability (CIN) generates genetic and karyotypic diversity that is common in hematological malignancies. Low to moderate levels of CIN are well tolerated and can promote cancer proliferation. However, high levels of CIN are lethal. Thus, CIN may serve both as a prognostic factor to predict clinical outcome and as a predictive biomarker. A retrospective study was performed to evaluate CIN in acute myeloid leukemia (AML). Chromosome mis-segregation frequency was correlated with clinical outcome in bone marrow core biopsy specimens from 17 AML cases. Additionally, we induced chromosome segregation errors in AML cell lines with AZ3146, an inhibitor of the Mps1 mitotic checkpoint kinase, to quantify the phenotypic effects of high CIN. We observed a broad distribution of chromosome mis-segregation frequency in AML bone marrow core specimens. High CIN correlated with complex karyotype in AML, as expected, although there was no clear survival effect. In addition to CIN, experimentally inducing chromosome segregation errors by Mps1 inhibition in AML cell lines causes DNA damage, micronuclei formation, and upregulation of interferon stimulated genes. High levels of CIN appear to be immunostimulatory, suggesting an opportunity to combine mitotic checkpoint inhibitors with immunotherapy in treatment of AML.
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http://dx.doi.org/10.1002/gcc.22880DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7597364PMC
November 2020

Technical laboratory standards for interpretation and reporting of acquired copy-number abnormalities and copy-neutral loss of heterozygosity in neoplastic disorders: a joint consensus recommendation from the American College of Medical Genetics and Genomics (ACMG) and the Cancer Genomics Consortium (CGC).

Genet Med 2019 09 29;21(9):1903-1916. Epub 2019 May 29.

Department of Pathology and Laboratory Medicine, Children's Hospital Los Angeles, Keck School of Medicine, University of Southern California, Los Angeles, CA, USA.

The detection of acquired copy-number abnormalities (CNAs) and copy-neutral loss of heterozygosity (CN-LOH) in neoplastic disorders by chromosomal microarray analysis (CMA) has significantly increased over the past few years with respect to both the number of laboratories utilizing this technology and the broader number of tumor types being assayed. This highlights the importance of standardizing the interpretation and reporting of acquired variants among laboratories. To address this need, a clinical laboratory-focused workgroup was established to draft recommendations for the interpretation and reporting of acquired CNAs and CN-LOH in neoplastic disorders. This project is a collaboration between the American College of Medical Genetics and Genomics (ACMG) and the Cancer Genomics Consortium (CGC). The recommendations put forth by the workgroup are based on literature review, empirical data, and expert consensus of the workgroup members. A four-tier evidence-based categorization system for acquired CNAs and CN-LOH was developed, which is based on the level of available evidence regarding their diagnostic, prognostic, and therapeutic relevance: tier 1, variants with strong clinical significance; tier 2, variants with some clinical significance; tier 3, clonal variants with no documented neoplastic disease association; and tier 4, benign or likely benign variants. These recommendations also provide a list of standardized definitions of terms used in the reporting of CMA findings, as well as a framework for the clinical reporting of acquired CNAs and CN-LOH, and recommendations for how to deal with suspected clinically significant germline variants.
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http://dx.doi.org/10.1038/s41436-019-0545-7DOI Listing
September 2019

Calcium waves occur as Drosophila oocytes activate.

Proc Natl Acad Sci U S A 2015 Jan 6;112(3):791-6. Epub 2015 Jan 6.

Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY 14853; and

Egg activation is the process by which a mature oocyte becomes capable of supporting embryo development. In vertebrates and echinoderms, activation is induced by fertilization. Molecules introduced into the egg by the sperm trigger progressive release of intracellular calcium stores in the oocyte. Calcium wave(s) spread through the oocyte and induce completion of meiosis, new macromolecular synthesis, and modification of the vitelline envelope to prevent polyspermy. However, arthropod eggs activate without fertilization: in the insects examined, eggs activate as they move through the female's reproductive tract. Here, we show that a calcium wave is, nevertheless, characteristic of egg activation in Drosophila. This calcium rise requires influx of calcium from the external environment and is induced as the egg is ovulated. Pressure on the oocyte (or swelling by the oocyte) can induce a calcium rise through the action of mechanosensitive ion channels. Visualization of calcium fluxes in activating eggs in oviducts shows a wave of increased calcium initiating at one or both oocyte poles and spreading across the oocyte. In vitro, waves also spread inward from oocyte pole(s). Wave propagation requires the IP3 system. Thus, although a fertilizing sperm is not necessary for egg activation in Drosophila, the characteristic of increased cytosolic calcium levels spreading through the egg is conserved. Because many downstream signaling effectors are conserved in Drosophila, this system offers the unique perspective of egg activation events due solely to maternal components.
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http://dx.doi.org/10.1073/pnas.1420589112DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4311822PMC
January 2015

Density matters: comparison of array platforms for detection of copy-number variation and copy-neutral abnormalities.

Genet Med 2013 Sep 4;15(9):706-12. Epub 2013 Apr 4.

Department of Human Genetics, Emory University School of Medicine, Atlanta, Georgia, USA.

Purpose: A combination of oligonucleotide and single-nucleotide polymorphism probes on the same array platform can detect copy-number abnormalities and copy-neutral aberrations such as uniparental disomy and long stretches of homozygosity. The single-nucleotide polymorphism probe density in commercially available platforms varies widely, which may affect the detection of copy-neutral abnormalities.

Methods: We evaluated the ability of array platforms with low (Oxford Gene Technology CytoSure ISCA uniparental disomy), mid-range (Agilent custom array), and high (Affymetrix CytoScan HD) single-nucleotide polymorphism probe density to detect copy-number variation, mosaicism, uniparental isodisomy, and absence of heterozygosity in 50 clinical samples.

Results: All platforms reliably detected copy-number variation, mosaicism, and uniparental isodisomy; however, absence-of-heterozygosity detection varied significantly. The low-density array called absence-of-heterozygosity regions not confirmed by the other platforms and also overestimated the length of true absence-of-heterozygosity regions. Furthermore, the low- and mid-density platforms failed to detect some small absence-of-heterozygosity regions that were identified by the high-density platform.

Conclusion: Variation in single-nucleotide polymorphism density can lead to major discrepancies in the detection of and confidence in copy-neutral abnormalities. Although suitable for uniparental disomy detection, copy-number plus single-nucleotide polymorphism arrays with 30,000 or fewer unique single-nucleotide polymorphism probes miscall absence-of-heterozygosity regions due to identity by descent.
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http://dx.doi.org/10.1038/gim.2013.36DOI Listing
September 2013

Protein phosphorylation changes reveal new candidates in the regulation of egg activation and early embryogenesis in D. melanogaster.

Dev Biol 2012 Oct 31;370(1):125-34. Epub 2012 Jul 31.

Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY 14853, United States.

Egg activation is the series of events that must occur for a mature oocyte to become capable of supporting embryogenesis. These events include changes to the egg's outer coverings, the resumption and completion of meiosis, the translation of new proteins, and the degradation of specific maternal mRNAs. While we know some of the molecules that direct the initial events of egg activation, it remains unclear how multiple pathways are coordinated to change the cellular state from mature oocyte to activated egg. Using a proteomic approach we have identified new candidates for the regulation and progression of egg activation. Reasoning that phosphorylation can simultaneously and rapidly modulate the activity of many proteins, we identified proteins that are post-translationally modified during the transition from oocyte to activated egg in Drosophila melanogaster. We find that at least 311 proteins change in phosphorylation state between mature oocytes and activated eggs. These proteins fall into various functional classes related to the events of egg activation including calcium binding, proteolysis, and protein translation. Our set of candidates includes genes already associated with egg activation, as well as many genes not previously studied during this developmental period. RNAi knockdown of a subset of these genes revealed a new gene, mrityu, necessary for embryonic development past the first mitosis. Thus, by identifying phospho-modulated proteins we have produced a focused candidate set for future genetic studies to test their roles in egg activation and the initiation of embryogenesis.
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http://dx.doi.org/10.1016/j.ydbio.2012.07.024DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3441184PMC
October 2012

Multiplex Chromosomal Exome Sequencing Accelerates Identification of ENU-Induced Mutations in the Mouse.

G3 (Bethesda) 2012 Jan 1;2(1):143-50. Epub 2012 Jan 1.

Department of Human Genetics, Emory University School of Medicine, Atlanta, Georgia 30322.

Forward genetic screens in Mus musculus have proved powerfully informative by revealing unsuspected mechanisms governing basic biological processes. This approach uses potent chemical mutagens, such as N-ethyl-N-nitrosourea (ENU), to randomly induce mutations in mice, which are then bred and phenotypically screened to identify lines that disrupt a specific biological process of interest. Although identifying a mutation using the rich resources of mouse genetics is straightforward, it is unfortunately neither fast nor cheap. Here we show that detecting newly induced causal variants in a forward genetic screen can be accelerated dramatically using a methodology that combines multiplex chromosome-specific exome capture, next-generation sequencing, rapid mapping, sequence annotation, and variation filtering. The key innovation of our method is multiplex capture and sequence that allows the simultaneous survey of both mutant, parental, and background strains in a single experiment. By comparing variants identified in mutant offspring with those found in dbSNP, the unmutagenized background strains, and parental lines, induced causative mutations can be distinguished immediately from preexisting variation or experimental artifact. Here we demonstrate this approach to find the causative mutations induced in four novel ENU lines identified from a recent ENU screen. In all four cases, after applying our method, we found six or fewer putative mutations (and sometimes only a single one). Determining the causative variant was then easily achieved through standard segregation approaches. We have developed this process into a community resource that will speed up individual labs' ability to identify the genetic lesion in mutant mouse lines; all of our reagents and software tools are open source and available to the broader scientific community.
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http://dx.doi.org/10.1534/g3.111.001669DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3276189PMC
January 2012

Creating a "hopeful monster": mouse forward genetic screens.

Methods Mol Biol 2011 ;770:313-36

Department of Human Genetics, Emory University School of Medicine, Atlanta, GA, USA.

One of the most straightforward approaches to making novel biological discoveries is the forward genetic screen. The time is ripe for forward genetic screens in the mouse since the mouse genome is sequenced, but the function of many of the genes remains unknown. Today, with careful planning, such screens are within the reach of even small individual labs. In this chapter we first discuss the types of screens in existence, as well as how to design a screen to recover mutations that are relevant to the interests of a lab. We then describe how to create mutations using the chemical N-ethyl-N-nitrosourea (ENU), including a detailed injection protocol. Next, we outline breeding schemes to establish mutant lines for each type of screen. Finally, we explain how to map mutations using recombination and how to ensure that a particular mutation causes a phenotype. Our goal is to make forward genetics in the mouse accessible to any lab with the desire to do it.
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http://dx.doi.org/10.1007/978-1-61779-210-6_12DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3827720PMC
November 2011

Disrupted dorsal neural tube BMP signaling in the cilia mutant Arl13b hnn stems from abnormal Shh signaling.

Dev Biol 2011 Jul 22;355(1):43-54. Epub 2011 Apr 22.

Department of Human Genetics, Emory University School of Medicine, Atlanta, GA 30322, USA.

In the embryonic neural tube, multiple signaling pathways work in concert to create functional neuronal circuits in the adult spinal cord. In the ventral neural tube, Sonic hedgehog (Shh) acts as a graded morphogen to specify neurons necessary for movement. In the dorsal neural tube, bone morphogenetic protein (BMP) and Wnt signals cooperate to specify neurons involved in sensation. Several signaling pathways, including Shh, rely on primary cilia in vertebrates. In this study, we used a mouse mutant with abnormal cilia, Arl13b(hnn), to study the relationship between cilia, cell signaling, and neural tube patterning. Arl13b(hnn) mutants have abnormal ventral neural tube patterning due to disrupted Shh signaling; in addition, dorsal patterning defects occur, but the cause of these is unknown. Here we show that the Arl13b(hnn) dorsal patterning defects result from abnormal BMP signaling. In addition, we find that Wnt ligands are abnormally expressed in Arl13b(hnn) mutants; surprisingly, however, downstream Wnt signaling is normal. We demonstrate that Arl13b is required non-autonomously for BMP signaling and Wnt ligand expression, indicating that the abnormal Shh signaling environment in Arl13b(hnn) embryos indirectly causes dorsal defects.
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http://dx.doi.org/10.1016/j.ydbio.2011.04.019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3119544PMC
July 2011

Microarray oligonucleotide probe designer (MOPeD): A web service.

Open Access Bioinformatics 2010 Nov;2(2010):145-155

Department of Human Genetics, Emory University School of Medicine, Atlanta, GA, USA.

Methods of genomic selection that combine high-density oligonucleotide microarrays with next-generation DNA sequencing allow investigators to characterize genomic variation in selected portions of complex eukaryotic genomes. Yet choosing which specific oligonucleotides to be use can pose a major technical challenge. To address this issue, we have developed a software package called MOPeD (Microarray Oligonucleotide Probe Designer), which automates the process of designing genomic selection microarrays. This web-based software allows individual investigators to design custom genomic selection microarrays optimized for synthesis with Roche NimbleGen's maskless photolithography. Design parameters include uniqueness of the probe sequences, melting temperature, hairpin formation, and the presence of single nucleotide polymorphisms. We generated probe databases for the human, mouse, and rhesus macaque genomes and conducted experimental validation of MOPeD-designed microarrays in human samples by sequencing the human X chromosome exome, where relevant sequence metrics indicated superior performance relative to a microarray designed by the Roche NimbleGen proprietary algorithm. We also performed validation in the mouse to identify known mutations contained within a 487-kb region from mouse chromosome 16, the mouse chromosome 16 exome (1.7 Mb), and the mouse chromosome 12 exome (3.3 Mb). Our results suggest that the open source MOPeD software package and website (http://moped.genetics.emory.edu/) will make a valuable resource for investigators in their sequence-based studies of complex eukaryotic genomes.
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http://dx.doi.org/10.2147/OAB.S13741DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3048354PMC
November 2010

SeqAnt: a web service to rapidly identify and annotate DNA sequence variations.

BMC Bioinformatics 2010 Sep 20;11:471. Epub 2010 Sep 20.

Department of Human Genetics, Emory University School of Medicine, Atlanta, GA, USA.

Background: The enormous throughput and low cost of second-generation sequencing platforms now allow research and clinical geneticists to routinely perform single experiments that identify tens of thousands to millions of variant sites. Existing methods to annotate variant sites using information from publicly available databases via web browsers are too slow to be useful for the large sequencing datasets being routinely generated by geneticists. Because sequence annotation of variant sites is required before functional characterization can proceed, the lack of a high-throughput pipeline to efficiently annotate variant sites can act as a significant bottleneck in genetics research.

Results: SeqAnt (Sequence Annotator) is an open source web service and software package that rapidly annotates DNA sequence variants and identifies recessive or compound heterozygous loci in human, mouse, fly, and worm genome sequencing experiments. Variants are characterized with respect to their functional type, frequency, and evolutionary conservation. Annotated variants can be viewed on a web browser, downloaded in a tab-delimited text file, or directly uploaded in a BED format to the UCSC genome browser. To demonstrate the speed of SeqAnt, we annotated a series of publicly available datasets that ranged in size from 37 to 3,439,107 variant sites. The total time to completely annotate these data completely ranged from 0.17 seconds to 28 minutes 49.8 seconds.

Conclusion: SeqAnt is an open source web service and software package that overcomes a critical bottleneck facing research and clinical geneticists using second-generation sequencing platforms. SeqAnt will prove especially useful for those investigators who lack dedicated bioinformatics personnel or infrastructure in their laboratories.
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http://dx.doi.org/10.1186/1471-2105-11-471DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2955049PMC
September 2010

Wispy, the Drosophila homolog of GLD-2, is required during oogenesis and egg activation.

Genetics 2008 Apr;178(4):2017-29

Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853, USA.

Egg activation is the process that modifies mature, arrested oocytes so that embryo development can proceed. One key aspect of egg activation is the cytoplasmic polyadenylation of certain maternal mRNAs to permit or enhance their translation. wispy (wisp) maternal-effect mutations in Drosophila block development during the egg-to-embryo transition. We show here that the wisp gene encodes a member of the GLD-2 family of cytoplasmic poly(A) polymerases (PAPs). The WISP protein is required for poly(A) tail elongation of bicoid, Toll, and torso mRNAs upon egg activation. In Drosophila, WISP and Smaug (SMG) have previously been reported to be required to trigger the destabilization of maternal mRNAs during egg activation. SMG is the major regulator of this activity. We report here that SMG is still translated in activated eggs from wisp mutant mothers, indicating that WISP does not regulate mRNA stability by controlling the translation of smg mRNA. We have also analyzed in detail the very early developmental arrest associated with wisp mutations. Pronuclear migration does not occur in activated eggs laid by wisp mutant females. Finally, we find that WISP function is also needed during oogenesis to regulate the poly(A) tail length of dmos during oocyte maturation and to maintain a high level of active (phospho-) mitogen-activated protein kinases (MAPKs).
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http://dx.doi.org/10.1534/genetics.107.084558DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2323793PMC
April 2008

Mechanical stimulation by osmotic and hydrostatic pressure activates Drosophila oocytes in vitro in a calcium-dependent manner.

Dev Biol 2008 Apr 26;316(1):100-9. Epub 2008 Jan 26.

Department of Molecular Biology and Genetics, 423 Biotechnology Building, Cornell University, Ithaca, NY 14853-2703, USA.

Embryogenesis in vertebrates and marine invertebrates begins when a mature oocyte is fertilized, resulting in a rise in intracellular calcium (Ca(2+)) that activates development. Insect eggs activate without fertilization via an unknown signal imparted to the egg during ovulation or egg laying. One hypothesis for the activating signal is that deformation of eggs as they pass through a tight orifice provides a mechanical stimulus to trigger activation. Ovulation could produce two forms of mechanical stimulus: external pressure resulting from the passage of oocytes from the ovary into the narrow oviducts, and osmotic pressure caused by hydration-induced swelling of the oocyte within the oviducts. Ovulation could also trigger activation by placing the oocyte in a new environment that contains an activating substance, such as a particular ion. Here, we provide the first evidence that Drosophila oocytes require Ca(2+) for activation, and that activation can be triggered in vitro by mechanical stimuli, specifically osmotic and hydrostatic pressure. Our results suggest that activation in Drosophila is triggered by a mechanosensitive process that allows external Ca(2+) to enter the oocyte and drive the events of activation. This will allow exploitation of Drosophila genetics to dissect molecular pathways involving Ca(2+) and the activation of development.
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http://dx.doi.org/10.1016/j.ydbio.2008.01.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2372165PMC
April 2008

Transitioning from egg to embryo: triggers and mechanisms of egg activation.

Dev Dyn 2008 Mar;237(3):527-44

Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853-2703, USA.

The transition from mature oocyte to developing embryo requires a coordinated series of events, collectively known as egg activation. Egg activation includes changes to egg coverings to prevent polyspermy, release of oocyte meiotic arrest, generation of haploid female and male pronuclei, changes in maternal mRNAs and protein populations, and cytoskeletal rearrangements. In many animals, egg activation is triggered by fertilization, which increases intracellular calcium within the oocyte and thereby regulates molecular events of egg activation. In other animals, fertilization-independent external signals, including mechanical stimulation of eggs and/or changes in ionic milieu, trigger activation. Recent studies have clarified the upstream portion of pathways leading to eggshell changes and cell cycle resumption and have identified activation-induced changes in maternal mRNA and protein profiles that can identify molecular players in the downstream events of egg activation. We review signals that trigger activation and how they link to subsequent molecular events of egg activation.
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http://dx.doi.org/10.1002/dvdy.21454DOI Listing
March 2008

The Drosophila calcipressin sarah is required for several aspects of egg activation.

Curr Biol 2006 Jul;16(14):1441-6

Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853, USA.

Activation of mature oocytes initiates development by releasing the prior arrest of female meiosis, degrading certain maternal mRNAs while initiating the translation of others, and modifying egg coverings. In vertebrates and marine invertebrates, the fertilizing sperm triggers activation events through a rise in free calcium within the egg. In insects, egg activation occurs independently of sperm and is instead triggered by passage of the egg through the female reproductive tract ; it is unknown whether calcium signaling is involved. We report here that mutations in sarah, which encodes an inhibitor of the calcium-dependent phosphatase calcineurin, disrupt several aspects of egg activation in Drosophila. Eggs laid by sarah mutant females arrest in anaphase of meiosis I and fail to fully polyadenylate and translate bicoid mRNA. Furthermore, sarah mutant eggs show elevated cyclin B levels, indicating a failure to inactivate M-phase promoting factor (MPF). Taken together, these results demonstrate that calcium signaling is involved in Drosophila egg activation and suggest a molecular mechanism for the sarah phenotype. We also find the conversion of the sperm nucleus into a functional male pronucleus is compromised in sarah mutant eggs, indicating that the Drosophila egg's competence to support male pronuclear maturation is acquired during activation.
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http://dx.doi.org/10.1016/j.cub.2006.06.024DOI Listing
July 2006