Publications by authors named "Eric Campos"

27 Publications

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Comprehensive interactome profiling of the human Hsp70 network highlights functional differentiation of J domains.

Mol Cell 2021 Apr 27. Epub 2021 Apr 27.

Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 1A8, Canada; Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, ON M5S 3E1, Canada. Electronic address:

Hsp70s comprise a deeply conserved chaperone family that has a central role in maintaining protein homeostasis. In humans, Hsp70 client specificity is provided by 49 different co-factors known as J domain proteins (JDPs). However, the cellular function and client specificity of JDPs have largely remained elusive. We have combined affinity purification-mass spectrometry (AP-MS) and proximity-dependent biotinylation (BioID) to characterize the interactome of all human JDPs and Hsp70s. The resulting network suggests specific functions for many uncharacterized JDPs, and we establish a role of conserved JDPs DNAJC9 and DNAJC27 in histone chaperoning and ciliogenesis, respectively. Unexpectedly, we find that the J domain of DNAJC27 but not of other JDPs can fully replace the function of endogenous DNAJC27, suggesting a previously unappreciated role for J domains themselves in JDP specificity. More broadly, our work expands the role of the Hsp70-regulated proteostasis network and provides a platform for further discovery of JDP-dependent functions.
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http://dx.doi.org/10.1016/j.molcel.2021.04.012DOI Listing
April 2021

Estimation of genetic parameters for body areas in Nile tilapia measured by digital image analysis.

J Anim Breed Genet 2021 Apr 23. Epub 2021 Apr 23.

Department of Animal Science, Universidade Federal de Viçosa, Viçosa, Brazil.

Digital image analysis is a practical, non-invasive, and relatively low-cost tool that may assist in the evaluation of body traits in Nile tilapia, being particularly useful for assessing difficult-to-measure variables, such as body areas. In this study, we aimed to estimate variance components and genetic parameters for body areas of Nile tilapia obtained by digital images. The data set comprised body weight (BW) records of 1,917 pond-reared fish at 366 days of age. Of this total, 656 animals were photographed and subjected to image analysis of trunk area (TA), head area (HA), caudal fin area (CFA) and fillet area (FA). Heritabilities and genetic correlations were estimated through multiple-trait models based on Bayesian inference. Heritability estimates for BW, TA, HA, CFA and FA were 0.25, 0.23, 0.26, 0.21 and 0.25, respectively. Genetic correlations between the traits were high and positive, ranging from 0.70 to 0.98. We highlight the genetic correlation between BW and TA (r  = 0.98) and FA (r  = 0.97). In view of the observed results, it can be concluded that trunk and fillet areas obtained by digital image analysis can lead to indirect genetic gains in weight and other body areas. In addition, the areas studied have potential as a selection criterion and may assist in studies on changes in the body shape in Nile tilapia.
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http://dx.doi.org/10.1111/jbg.12551DOI Listing
April 2021

Evaluation of a panel of microsatellite markers to study their applications in and to reveal the genetic diversity in .

Anim Biotechnol 2020 Oct 5:1-9. Epub 2020 Oct 5.

Department of Animal Sciences and Postgraduate Program in Animal Sciences, Center of Agricultural Sciences, State University of Londrina, Londrina, Brazil.

Jewel tetra () is a freshwater fish found in several rivers and basins in South America. The present study is the first study to create a panel of microsatellite markers for detecting genetic diversity in and evaluating the application of these markers in . In total, 44 individuals were genotyped from the natural (WIL,  = 20) and stock in captivity (CAP,  = 24) population. Moreover, 19 microsatellite markers were obtained, of which only 8 loci presented a high degree polymorphism. In total, 45 alleles were detected, ranging from 126 bp (Hype2G2) to 420 bp (Hype2E2). The Hardy-Weinberg equilibrium ( < 0.05) revealed significant difference in one locus in WIL (Hype1G4) and three loci in CAP (Hype1F4, Hype2C3, and Hype2G2). Null alleles ( < 0.05) were present in only one locus (Hype1G4). The WIL and CAP populations revealed high genetic diversity during F analysis. The cross-amplification test for revealed that only two loci (Hype2C3 and Hype2G2B) presented satisfactory transferability results. The developed microsatellite primers will be useful in studying the genetic diversity and population structure of in wild populations and fish farms in the Brazilian and other South American basins.
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http://dx.doi.org/10.1080/10495398.2020.1826957DOI Listing
October 2020

Interactions With Histone H3 & Tools to Study Them.

Front Cell Dev Biol 2020 31;8:701. Epub 2020 Jul 31.

Genetics & Genome Biology Program, The Hospital for Sick Children, Toronto, ON, Canada.

Histones are an integral part of chromatin and thereby influence its structure, dynamics, and functions. The effects of histone variants, posttranslational modifications, and binding proteins is therefore of great interest. From the moment that they are deposited on chromatin, nucleosomal histones undergo dynamic changes in function of the cell cycle, and as DNA is transcribed and replicated. In the process, histones are not only modified and bound by various proteins, but also shuffled, evicted, or replaced. Technologies and tools to study such dynamic events continue to evolve and better our understanding of chromatin and of histone proteins proper. Here, we provide an overview of H3.1 and H3.3 histone dynamics throughout the cell cycle, while highlighting some of the tools used to study their protein-protein interactions. We specifically discuss how histones are chaperoned, modified, and bound by various proteins at different stages of the cell cycle. Established and select emerging technologies that furthered (or have a high potential of furthering) our understanding of the dynamic histone-protein interactions are emphasized. This includes experimental tools to investigate spatiotemporal changes on chromatin, the role of histone chaperones, histone posttranslational modifications, and histone-binding effector proteins.
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http://dx.doi.org/10.3389/fcell.2020.00701DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7411163PMC
July 2020

Effects of genotype × environment interaction on the estimation of genetic parameters and gains in Nile tilapia.

J Appl Genet 2020 Dec 19;61(4):575-580. Epub 2020 Aug 19.

PeixeGen Research Group - Management, Breeding and Molecular Genetics of Freshwater Fish, Department of Animal Science, State University of Maringá, Avenida Colombo 5790, Maringá, Parana, Brazil.

Nile tilapia (Oreochromis niloticus) is the major fish species produced in Brazil, a country with a vast territory and great climate diversity. This study assessed the effects of the genotype × environment interaction on heritability estimates and selection responses in Nile tilapia (Tilamax strain) cultivated in earthen ponds and net cages. The weight at harvest, trunk length, and head percentage of 4400 individuals were determined. Trait heritabilities were higher in pond fish (0.27-0.52) than in caged fish (0.09-0.33). Genetic correlations between farming systems were lower than 0.5 for the three traits. The rank position of the top 10 families differed according to the environment, as did the response to direct and indirect selection. The results revealed significant genotype × environment effects on the heritability of Nile tilapia farmed under different systems.
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http://dx.doi.org/10.1007/s13353-020-00576-2DOI Listing
December 2020

Derivation of a continuous time dynamic planar system with two unstable foci from a three-dimensional chaotic piecewise linear system.

Authors:
Eric Campos

Chaos 2020 May;30(5):053114

División de Matemáticas Aplicadas, Instituto Potosino de Investigación Científica y Tecnológica A. C., Camino a la Presa San José 2055, col. Lomas 4a Sección, 78216 San Luis Potosí, SLP, Mexico.

In this paper, we introduce a class of continuous time dynamical planar systems that is capable of generating attractors in the plane by means of the use of hysteresis and at least two unstable foci. This class of systems shows stretching and folding behavior due to unstable equilibria and hysteresis. Hysteresis is used to overwhelm the constraints on the behavior of planar systems. This class of systems is derived from three-dimensional piecewise linear systems that have two manifolds, one stable and the other unstable, to generate heteroclinic chaos. Two numerical examples are given accordingly to the developed theory.
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http://dx.doi.org/10.1063/1.5144709DOI Listing
May 2020

The Pluripotency Regulator PRDM14 Requires Hematopoietic Regulator CBFA2T3 to Initiate Leukemia in Mice.

Mol Cancer Res 2019 07 23;17(7):1468-1479. Epub 2019 Apr 23.

Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada.

PR domain-containing 14 () is a pluripotency regulator central to embryonic stem cell identity and primordial germ cell specification. Genomic regions containing are often amplified leading to misexpression in human cancer. expression in mouse hematopoietic stem cells (HSC) leads to progenitor cell expansion prior to the development of T-cell acute lymphoblastic leukemia (T-ALL), consistent with PRDM14's role in cancer initiation. Here, we demonstrate mechanistic insight into PRDM14-driven leukemias . Mass spectrometry revealed novel PRDM14-protein interactions including histone H1, RNA-binding proteins, and the master hematopoietic regulator CBFA2T3. In mouse leukemic cells, CBFA2T3 and PRDM14 associate independently of the related ETO family member CBFA2T2, PRDM14's primary protein partner in pluripotent cells. CBFA2T3 plays crucial roles in HSC self-renewal and lineage commitment, and participates in oncogenic translocations in acute myeloid leukemia. These results suggest a model whereby PRDM14 recruits CBFA2T3 to DNA, leading to gene misregulation causing progenitor cell expansion and lineage perturbations preceding T-ALL development. Strikingly, -induced T-ALL does not occur in mice deficient for , demonstrating that is required for leukemogenesis. Moreover, T-ALL develops in heterozygotes with a significantly longer latency, suggesting that PRDM14-associated T-ALL is sensitive to levels. Our study highlights how an oncogenic protein uses a native protein in progenitor cells to initiate leukemia, providing insight into PRDM14-driven oncogenesis in other cell types. IMPLICATIONS: The pluripotency regulator PRDM14 requires the master hematopoietic regulator CBFA2T3 to initiate leukemia in progenitor cells, demonstrating an oncogenic role for CBFA2T3 and providing an avenue for targeting cancer-initiating cells.
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http://dx.doi.org/10.1158/1541-7786.MCR-18-1327DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6610658PMC
July 2019

Morphospace exploration reveals divergent fitness optima between plants and pollinators.

PLoS One 2019 13;14(3):e0213029. Epub 2019 Mar 13.

Department of Biology, University of Washington, Seattle, Washington, United States of America.

The obligate mutualism and exquisite specificity of many plant-pollinator interactions lead to the expectation that flower phenotypes (e.g., corolla tube length) and corresponding pollinator traits (e.g., hawkmoth proboscis length) are congruent as a result of coevolution by natural selection. However, the effect of variation in flower morphology on the fitness of plants and their pollinators has not been quantified systematically. In this study, we employed the theoretical morphospace paradigm using a combination of 3D printing, electronic sensing, and machine vision technologies to determine the influence of two flower morphological features (corolla curvature and nectary diameter) on the fitness of both parties: the artificial flower and its hawkmoth pollinator. Contrary to the expectation that the same flower morphology maximizes the fitness of both plant and pollinator, we found that the two parties have divergent optima for corolla curvature, with non-overlapping fitness peaks in flower morphospace. The divergent fitness optima between plants and pollinators could lead to evolutionary diversification in both groups.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0213029PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6415803PMC
November 2019

H3-H4 Histone Chaperone Pathways.

Annu Rev Genet 2018 11 5;52:109-130. Epub 2018 Sep 5.

Genetics & Genome Biology Program, The Hospital for Sick Children, Toronto, Ontario M5G 0A4, Canada; email:

Nucleosomes compact and organize genetic material on a structural level. However, they also alter local chromatin accessibility through changes in their position, through the incorporation of histone variants, and through a vast array of histone posttranslational modifications. The dynamic nature of chromatin requires histone chaperones to process, deposit, and evict histones in different tissues and at different times in the cell cycle. This review focuses on the molecular details of canonical and variant H3-H4 histone chaperone pathways that lead to histone deposition on DNA as they are currently understood. Emphasis is placed on the most established pathways beginning with the folding, posttranslational modification, and nuclear import of newly synthesized H3-H4 histones. Next, we review the deposition of replication-coupled H3.1-H4 in S-phase and replication-independent H3.3-H4 via alternative histone chaperone pathways. Highly specialized histone chaperones overseeing the deposition of histone variants are also briefly discussed.
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http://dx.doi.org/10.1146/annurev-genet-120417-031547DOI Listing
November 2018

A Screen for Epstein-Barr Virus Proteins That Inhibit the DNA Damage Response Reveals a Novel Histone Binding Protein.

J Virol 2018 07 29;92(14). Epub 2018 Jun 29.

Department of Molecular Genetics, University of Toronto, Toronto, Canada

To replicate and persist in human cells, linear double-stranded DNA (dsDNA) viruses, such as Epstein-Barr virus (EBV), must overcome the host DNA damage response (DDR) that is triggered by the viral genomes. Since this response is necessary to maintain cellular genome integrity, its inhibition by EBV is likely an important factor in the development of cancers associated with EBV infection, including gastric carcinoma. Here we present the first extensive screen of EBV proteins that inhibit dsDNA break signaling. We identify the BKRF4 tegument protein as a DDR inhibitor that interferes with histone ubiquitylation at dsDNA breaks and recruitment of the RNF168 histone ubiquitin ligase. We further show that BKRF4 binds directly to histones through an acidic domain that targets BKRF4 to cellular chromatin and is sufficient to inhibit dsDNA break signaling. BKRF4 transcripts were detected in EBV-positive gastric carcinoma cells (AGS-EBV), and these increased in lytic infection. Silencing of BKRF4 in both latent and lytic AGS-EBV cells (but not in EBV-negative AGS cells) resulted in increased dsDNA break signaling, confirming a role for BKRF4 in DDR inhibition in the context of EBV infection and suggesting that BKRF4 is expressed in latent cells. BKRF4 was also found to be consistently expressed in EBV-positive gastric tumors in the absence of a full lytic infection. The results suggest that BKRF4 plays a role in inhibiting the cellular DDR in latent and lytic EBV infection and that the resulting accumulation of DNA damage might contribute to development of gastric carcinoma. Epstein-Barr virus (EBV) infects most people worldwide and is causatively associated with several types of cancer, including ∼10% of gastric carcinomas. EBV encodes ∼80 proteins, many of which are believed to manipulate cellular regulatory pathways but are poorly characterized. The DNA damage response (DDR) is one such pathway that is critical for maintaining genome integrity and preventing cancer-associated mutations. In this study, a screen for EBV proteins that inhibit the DDR identified BKRF4 as a DDR inhibitor that binds histones and blocks their ubiquitylation at the DNA damage sites. We also present evidence that BKRF4 is expressed in both latent and lytic forms of EBV infection, where it downregulates the DDR, as well as in EBV-positive gastric tumors. The results suggest that BKRF4 could contribute to the development of gastric carcinoma through its ability to inhibit the DDR.
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http://dx.doi.org/10.1128/JVI.00262-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6026726PMC
July 2018

Analysis of the Histone H3.1 Interactome: A Suitable Chaperone for the Right Event.

Mol Cell 2015 Nov 29;60(4):697-709. Epub 2015 Oct 29.

Howard Hughes Medical Institute, New York University School of Medicine, NY 10016, USA; Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, NY 10016, USA. Electronic address:

Despite minimal disparity at the sequence level, mammalian H3 variants bind to distinct sets of polypeptides. Although histone H3.1 predominates in cycling cells, our knowledge of the soluble complexes that it forms en route to deposition or following eviction from chromatin remains limited. Here, we provide a comprehensive analysis of the H3.1-binding proteome, with emphasis on its interactions with histone chaperones and components of the replication fork. Quantitative mass spectrometry revealed 170 protein interactions, whereas a large-scale biochemical fractionation of H3.1 and associated enzymatic activities uncovered over twenty stable protein complexes in dividing human cells. The sNASP and ASF1 chaperones play pivotal roles in the processing of soluble histones but do not associate with the active CDC45/MCM2-7/GINS (CMG) replicative helicase. We also find TONSL-MMS22L to function as a H3-H4 histone chaperone. It associates with the regulatory MCM5 subunit of the replicative helicase.
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http://dx.doi.org/10.1016/j.molcel.2015.08.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4656108PMC
November 2015

BRD4 assists elongation of both coding and enhancer RNAs by interacting with acetylated histones.

Nat Struct Mol Biol 2014 Dec 10;21(12):1047-57. Epub 2014 Nov 10.

Program in Genomics of Differentiation, National Institutes of Child Health and Human Development, Bethesda, Maryland, USA.

Small-molecule BET inhibitors interfere with the epigenetic interactions between acetylated histones and the bromodomains of the BET family proteins, including BRD4, and they potently inhibit growth of malignant cells by targeting cancer-promoting genes. BRD4 interacts with the pause-release factor P-TEFb and has been proposed to release RNA polymerase II (Pol II) from promoter-proximal pausing. We show that BRD4 occupies widespread genomic regions in mouse cells and directly stimulates elongation of both protein-coding transcripts and noncoding enhancer RNAs (eRNAs), in a manner dependent on bromodomain function. BRD4 interacts with elongating Pol II complexes and assists Pol II in progression through hyperacetylated nucleosomes by interacting with acetylated histones via bromodomains. On active enhancers, the BET inhibitor JQ1 antagonizes BRD4-associated eRNA synthesis. Thus, BRD4 is involved in multiple steps of the transcription hierarchy, primarily by facilitating transcript elongation both at enhancers and on gene bodies independently of P-TEFb.
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http://dx.doi.org/10.1038/nsmb.2912DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4720983PMC
December 2014

Epigenetic inheritance: histone bookmarks across generations.

Trends Cell Biol 2014 Nov 18;24(11):664-74. Epub 2014 Sep 18.

Howard Hughes Medical Institute, Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, NY, USA. Electronic address:

Multiple circuitries ensure that cells respond correctly to the environmental cues within defined cellular programs. There is increasing evidence suggesting that cellular memory for these adaptive processes can be passed on through cell divisions and generations. However, the mechanisms by which this epigenetic information is transferred remain elusive, largely because it requires that such memory survive through gross chromatin remodeling events during DNA replication, mitosis, meiosis, and developmental reprogramming. Elucidating the processes by which epigenetic information survives and is transmitted is a central challenge in biology. In this review, we consider recent advances in understanding mechanisms of epigenetic inheritance with a focus on histone segregation at the replication fork, and how an epigenetic memory may get passed through the paternal lineage.
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http://dx.doi.org/10.1016/j.tcb.2014.08.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4254315PMC
November 2014

USP33 regulates centrosome biogenesis via deubiquitination of the centriolar protein CP110.

Nature 2013 Mar;495(7440):255-9

Department of Pathology and Cancer Institute, Smilow Research Center, New York University School of Medicine, 522 1st Avenue, New York, New York 10016, USA.

Centrosome duplication is critical for cell division, and genome instability can result if duplication is not restricted to a single round per cell cycle. Centrosome duplication is controlled in part by CP110, a centriolar protein that positively regulates centriole duplication while restricting centriole elongation and ciliogenesis. Maintenance of normal CP110 levels is essential, as excessive CP110 drives centrosome over-duplication and suppresses ciliogenesis, whereas its depletion inhibits centriole amplification and leads to highly elongated centrioles and aberrant assembly of cilia in growing cells. CP110 levels are tightly controlled, partly through ubiquitination by the ubiquitin ligase complex SCF(cyclin F) during G2 and M phases of the cell cycle. Here, using human cells, we report a new mechanism for the regulation of centrosome duplication that requires USP33, a deubiquitinating enzyme that is able to regulate CP110 levels. USP33 interacts with CP110 and localizes to centrioles primarily in S and G2/M phases, the periods during which centrioles duplicate and elongate. USP33 potently and specifically deubiquitinates CP110, but not other cyclin-F substrates. USP33 activity antagonizes SCF(cyclin F)-mediated ubiquitination and promotes the generation of supernumerary centriolar foci, whereas ablation of USP33 destabilizes CP110 and thereby inhibits centrosome amplification and mitotic defects. To our knowledge, we have identified the first centriolar deubiquitinating enzyme whose expression regulates centrosome homeostasis by countering cyclin-F-mediated destruction of a key substrate. Our results point towards potential therapeutic strategies for inhibiting tumorigenesis associated with centrosome amplification.
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http://dx.doi.org/10.1038/nature11941DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3815529PMC
March 2013

The program for processing newly synthesized histones H3.1 and H4.

Nat Struct Mol Biol 2010 Nov 17;17(11):1343-51. Epub 2010 Oct 17.

Howard Hughes Medical Institute, Department of Biochemistry, New York University School of Medicine, New York, New York, USA.

The mechanism by which newly synthesized histones are imported into the nucleus and deposited onto replicating chromatin alongside segregating nucleosomal counterparts is poorly understood, yet this program is expected to bear on the putative epigenetic nature of histone post-translational modifications. To define the events by which naive pre-deposition histones are imported into the nucleus, we biochemically purified and characterized the full gamut of histone H3.1-containing complexes from human cytoplasmic fractions and identified their associated histone post-translational modifications. Through reconstitution assays, biophysical analyses and live cell manipulations, we describe in detail this series of events, namely the assembly of H3-H4 dimers, the acetylation of histones by the HAT1 holoenzyme and the transfer of histones between chaperones that culminates with their karyopherin-mediated nuclear import. We further demonstrate the high degree of conservation for this pathway between higher and lower eukaryotes.
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http://dx.doi.org/10.1038/nsmb.1911DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2988979PMC
November 2010

Traf7, a MyoD1 transcriptional target, regulates nuclear factor-κB activity during myogenesis.

EMBO Rep 2010 Dec 15;11(12):969-76. Epub 2010 Oct 15.

Department of Pathology, New York University School of Medicine, New York 10016, USA.

We have identified the E3 ligase Traf7 as a direct MyoD1 target and show that cell cycle exit-an early event in muscle differentiation-is linked to decreased Traf7 expression. Depletion of Traf7 accelerates myogenesis, in part through downregulation of nuclear factor-κB (NF-κB) activity. We used a proteomic screen to identify NEMO, the NF-κB essential modulator, as a Traf7-interacting protein. Finally, we show that ubiquitylation of NF-κB essential modulator is regulated exclusively by Traf7 activity in myoblasts. Our results suggest a new mechanism by which MyoD1 function is coupled to NF-κB activity through Traf7, regulating the balance between cell cycle progression and differentiation during myogenesis.
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http://dx.doi.org/10.1038/embor.2010.154DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2999857PMC
December 2010

New chaps in the histone chaperone arena.

Genes Dev 2010 Jul;24(13):1334-8

Department of Biochemistry, Howard Hughes Medical Institute, New York University School of Medicine, New York, NY 10016, USA.

Understanding exactly how chromatin is assembled is paramount to addressing how select histone modifications may be transmitted, a putative epigenetic process. In the June 15, 2010, issue of Genes & Development, Drané and colleagues (pp. 1253-1265) identified DAXX as a novel H3.3-specific chaperone. This finding, in the context of others published by Goldberg and colleagues in Cell and Sawatsubashi and colleagues (pp. 159-170) in the January 15, 2010, issue of Genes & Development, provides the impetus for uncovering the mechanistic and functional properties of alternative histone deposition pathways.
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http://dx.doi.org/10.1101/gad.1946810DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2895192PMC
July 2010

Histones: annotating chromatin.

Annu Rev Genet 2009 ;43:559-99

Department of Biochemistry, Howard Hughes Medical Institute, NYU School of Medicine, New York, New York 10016, USA.

Chromatin is a highly regulated nucleoprotein complex through which genetic material is structured and maneuvered to elicit cellular processes, including transcription, cell division, differentiation, and DNA repair. In eukaryotes, the core of this structure is composed of nucleosomes, or repetitive histone octamer units typically enfolded by 147 base pairs of DNA. DNA is arranged and indexed through these nucleosomal structures to adjust local chromatin compaction and accessibility. Histones are subject to multiple covalent posttranslational modifications, some of which alter intrinsic chromatin properties, others of which present or hinder binding modules for non-histone, chromatin-modifying complexes. Although certain histone marks correlate with different biological outputs, we have yet to fully appreciate their effects on transcription and other cellular processes. Tremendous advancements over the past years have uncovered intriguing histone-related matters and raised important related questions. This review revisits past breakthroughs and discusses novel developments that pertain to histone posttranslational modifications and the affects they have on transcription and DNA packaging.
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http://dx.doi.org/10.1146/annurev.genet.032608.103928DOI Listing
December 2009

NAD(P)H quinone oxidoreductase 1 inhibits the proteasomal degradation of the tumour suppressor p33(ING1b).

EMBO Rep 2008 Jun 4;9(6):576-81. Epub 2008 Apr 4.

Department of Dermatology and Skin Science, Jack Bell Research Centre, Vancouver Coastal Health Research Institute, University of British Columbia, 2660 Oak Street, Vancouver, British Columbia V6G 3Z6, Canada.

The tumour suppressor p33(ING1b) ((ING1b) for inhibitor of growth family, member 1b) is important in cellular stress responses, including cell-cycle arrest, apoptosis, chromatin remodelling and DNA repair; however, its degradation pathway is still unknown. Recently, we showed that genotoxic stress induces p33(ING1b) phosphorylation at Ser 126, and abolishment of Ser 126 phosphorylation markedly shortened its half-life. Therefore, we suggest that Ser 126 phosphorylation modulates the interaction of p33(ING1b) with its degradation machinery, stabilizing this protein. Combining the use of inhibitors of the main degradation pathways in the nucleus (proteasome and calpains), partial isolation of the proteasome complex, and in vitro interaction and degradation assays, we set out to determine the degradation mechanism of p33(ING1b). We found that p33(ING1b) is degraded in the 20S proteasome and that NAD(P)H quinone oxidoreductase 1 (NQO1), an oxidoreductase previously shown to modulate the degradation of p53 in the 20S proteasome, inhibits the degradation of p33(ING1b). Furthermore, ultraviolet irradiation induces p33(ING1b) phosphorylation at Ser 126, which, in turn, facilitates its interaction with NQO1.
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http://dx.doi.org/10.1038/embor.2008.48DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2427386PMC
June 2008

Phosphorylation of the tumor suppressor p33(ING1b) at Ser-126 influences its protein stability and proliferation of melanoma cells.

FASEB J 2007 Nov 21;21(13):3705-16. Epub 2007 Jun 21.

Department of Dermatology and Skin Science, Vancouver Coastal Health Research Institute, Jack Bell Research Centre, 2660 Oak St., Vancouver, BC, Canada V6H 3Z6.

ING (inhibitor of growth) tumor suppressors regulate cell-cycle checkpoints, apoptosis, and ultimately tumor suppression. Among the ING family members, p33(ING1b) is the most intensively studied and plays an important role in the cellular stress response to DNA damage. Here we demonstrate that there is basal phosphorylation of p33(ING1b) at Ser-126 in normal physiological conditions and that this phosphorylation is increased on DNA damage. The mutation of Ser-126 to alanine dramatically shortened the half-life of p33(ING1b). Furthermore, we found that both Chk1 and Cdk1 can phosphorylate this residue. Interestingly, while Cdk1 can phosphorylate p33(ING1b) at Ser-126 in nonstress conditions, Chk1 predominantly phosphorylates this residue on DNA damage, which suggests that p33(ING1b) is a downstream target of the ATM/ATR response cascade to genotoxic stress. More importantly, our data indicate that the Ser-126 residue plays a key role in regulating the expression of cyclin B1 and proliferation of melanoma cells.
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http://dx.doi.org/10.1096/fj.07-8069comDOI Listing
November 2007

The ING1b tumor suppressor facilitates nucleotide excision repair by promoting chromatin accessibility to XPA.

Exp Cell Res 2007 May 22;313(8):1628-38. Epub 2007 Feb 22.

Department of Dermatology and Skin Science, Jack Bell Research Centre, Vancouver Coastal Health Research Institute, The University of British Columbia, Vancouver, Canada BC V6H 3Z6.

ING1b is the most studied ING family protein and perhaps the most ubiquitously and abundantly expressed. This protein is involved in the regulation of various biological functions ranging from senescence, cell cycle arrest, apoptosis, to DNA repair. ING1b is upregulated by UV irradiation and enhances the removal of bulky nucleic acid photoproducts. In this study, we provide evidence that ING1b mediates nucleotide excision repair by facilitating the access to damaged nucleosomal DNA. We demonstrate that ING1b is not recruited to UV-induced DNA lesions but enhances nucleotide excision repair only in XPC-proficient cells, implying an essential role in early steps of the 'access, repair, restore' model. We also find that ING1b alters histone acetylation dynamics upon exposure to UV radiation and induces chromatin relaxation in microccocal nuclease digestion assay, revealing that ING1b may allow better access to nucleotide excision repair machinery. More importantly, ING1b associates with chromatin in a UV-inducible manner and facilitates DNA access to nucleotide excision repair factor XPA. Furthermore, depletion of the endogenous ING1b results to the sensitization of cells at S-phase to UV irradiation. Taken together, these observations establish a role of ING1b acting as a chromatin accessibility factor for DNA damage recognition proteins upon genotoxic injury.
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http://dx.doi.org/10.1016/j.yexcr.2007.02.010DOI Listing
May 2007

Osteopontin expression correlates with melanoma invasion.

J Invest Dermatol 2005 May;124(5):1044-52

Division of Dermatology, Department of Medicine, University British Columbia, Vancouver, Canada.

Melanoma is one of the most aggressive cancers affecting humans. Although early melanomas are curable with surgical excision, metastatic melanomas are associated with high mortality. The mechanism of melanoma development, progression, and metastasis is largely unknown. In order to uncover genes unique to melanoma cells, we used high-density DNA microarrays to examine the gene expression profiles of metastatic melanoma nodules using benign nevi as controls. Over 190 genes were significantly overexpressed in metastatic melanomas compared with normal nevi by at least 2-fold. One of the most abundantly expressed genes in metastatic melanoma nodules is osteopontin (OPN). Immunohistochemistry staining on tissue microarrays and individual skin biopsies representing different stages of melanoma progression revealed that OPN expression is first acquired at the step of melanoma tissue invasion. In addition, blocking of OPN expression by RNA interference reduced melanoma cell numbers in vitro. Our observations suggest that OPN may be acquired early in melanoma development and progression, and may enhance tumor cell growth in invasive melanoma.
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http://dx.doi.org/10.1111/j.0022-202X.2005.23680.xDOI Listing
May 2005

Development of a PAN-specific, affinity-purified anti-acetylated lysine antibody for detection, identification, isolation, and intracellular localization of acetylated protein.

J Immunoassay Immunochem 2005 ;26(1):13-23

Department of Medicine, ImmuneChem Pharmaceuticals Inc, Burnaby, British Columbia, Canada.

Acetylation on the lysine residue is an important event of posttranslational modification of proteins. In this study, we developed a simple method to produce and to affinity purify the specific anti-acetylated lysine polyclonal antibody, which is useful for the detection, identification, isolation, and intracellular localization of acetylated proteins on the lysine residues. We utilized the chemically acetylated hemocyanin of keyhole limpets (KLH) as an immunogen to raise the immune serum and to isolate the population of the acetylated lysine specific antibody using the immobilized acetylated lysine as immunoaffinity-ligand. The isolated antibody was tested to be useful for ELISA, immunoblotting detection, immunofluorescent localization, and affinity isolation of the acetylated proteins.
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http://dx.doi.org/10.1081/ias-200041153DOI Listing
September 2005

XAF1 expression is significantly reduced in human melanoma.

J Invest Dermatol 2004 Dec;123(6):1127-34

Department of Medicine, Division of Dermatology, University of British Columbia, Vancouver, British Columbia, Canada.

Deregulation of apoptotic processes is likely one of the key factors contributing to the malignant nature of melanoma marked by strong chemoresistance. X-linked inhibitor of apoptosis protein (XIAP) suppresses apoptosis through the inhibition of various caspases. Recently, XIAP-associated factor 1 (XAF1) has been identified as a XIAP-binding protein that antagonizes the anti-apoptotic activity of XIAP. In this study, we sought to determine the role of XAF1 in melanoma progression. Analysis of XAF1 mRNA expression in melanoma cell lines revealed that XAF1 mRNA was downregulated in 15 of 16 cell lines examined. We next evaluated XAF1 protein expression on a tissue microarray representing 40 benign nevi and 70 primary melanomas. Our results showed that XAF1 expression in melanoma tissues was significantly reduced compared with benign melanocytic nevi (p<0.05). Our data also demonstrated that the substantial reduction of XAF1 expression occurred in both nucleus and cytoplasm in the tumor cells (p<0.0001 for both). Reduced XAF1 expression, however, was not significantly correlated with tumor thickness and 5-y patient survival. Further studies are required to understand the molecular mechanisms governing the selective loss of XAF1 expression in the tumor tissue.
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http://dx.doi.org/10.1111/j.0022-202X.2004.23467.xDOI Listing
December 2004

Mutations of the ING1 tumor suppressor gene detected in human melanoma abrogate nucleotide excision repair.

Int J Oncol 2004 Jul;25(1):73-80

Department of Medicine, Division of Dermatology, University of British Columbia, Vancouver, British Columbia, Canada.

Epidemiological evidence indicates that ultraviolet radiation (UVR) is the primary environmental cause of the rapid increase in the incidence of human cutaneous melanoma observed in the past decades. However, the genetic changes caused by UVR that lead to melanoma formation remain unclear. The ING1 (inhibitor of growth 1) tumor suppressor plays an important role in cellular stress response to UVR. To further investigate whether ING1 is involved in melanoma development, we examined the mutational status of the ING1 gene in 46 human cutaneous melanoma biopsies and characterized the biological importance of ING1 mutations in nucleotide excision repair. Single-strand conformation polymorphism and DNA sequencing were used to detect the mutational status of the ING1 gene. The host-cell-reactivation assay and radioimmunoassay were used to determine the role of ING1 mutations in nucleotide excision repair. We show that 20% of the melanoma primaries contained missense mutations in the SAP30-interacting domain and PHD finger motif of the ING1 gene with the R102L and N260S alterations observed more than once. Furthermore, our data indicate that patients that harbor ING1 mutations in the tumors have a higher risk to die from the disease within 5 years (50%) compared to patients with no ING1 mutation (18%). Moreover, we demonstrated that mutations at codon 102 or 260 as well as deletion of the PHD finger motif are detrimental to p33ING1-mediated enhancement of DNA repair. Taken together, our data indicate that ING1 mutations abrogate its enhancement in nucleotide excision repair.
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July 2004

Increased expression of integrin-linked kinase is correlated with melanoma progression and poor patient survival.

Clin Cancer Res 2003 Oct;9(12):4409-14

Department of Medicine, Division of Dermatology, University of British Columbia, Vancouver, British Columbia, V6H 3Z6 Canada.

Purpose: Integrin-linked kinase (ILK), a key component of the extracellular matrix adhesion, has been studied extensively in recent years. Overexpression of ILK in epithelial cells results in anchorage-independent cell growth with increased cell cycle progression. Furthermore, increased ILK expression is correlated with progression of several human tumor types, including breast, prostate, and colon carcinomas. However, the role of ILK overexpression in human melanoma pathogenesis is not known. To investigate whether ILK plays a role in melanoma progression, we measured ILK expression in primary melanoma biopsies at various stages of invasion and evaluated the prognostic value of ILK expression in human melanoma.

Experimental Design: We used tissue microarray and immunohistochemistry to determine ILK expression in 67 primary melanomas and analyzed the correlation between ILK expression and melanoma progression and 5-year patient survival.

Results: We show that strong ILK expression is significantly associated with melanoma thickness. Strong ILK expression was observed in 0, 22, 33, and 63% in melanoma biopsies 3.0 mm in thickness, respectively. Furthermore, strong ILK expression was detected in 83% of the tumors with lymph node invasion compared with only 18% for tumors without lymph node invasion (P < 0.01). Strikingly, our data revealed that strong ILK expression is inversely correlated with 5-year patient survival (P < 0.05).

Conclusion: ILK expression increases dramatically with melanoma invasion and progression and is inversely correlated with patient survival.
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October 2003

Analyses of the tumour suppressor ING1 expression and gene mutation in human basal cell carcinoma.

Int J Oncol 2003 Apr;22(4):927-31

Department of Dermatology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, P.R. China.

The incidence of basal cell carcinoma is the highest among all human malignancies. Epidemiological evidence indicates that ultraviolet radiation is the primary environmental cause for the pathogenesis of basal cell carcinoma. However, the genetic changes caused by ultraviolet radiation that lead to basal cell carcinoma formation remain unclear. We and others have demonstrated that the ING1 (inhibitor of growth 1) tumour suppressor plays an important role in cellular stress response to ultraviolet irradiation, such as DNA repair and apoptosis. This study was designed to investigate whether ING1 is overexpressed and/or mutated in human basal cell carcinoma. Immunohistochemistry, single-strand conformation polymorphism, and DNA sequencing were used to determine the expression and mutational status of the ING1 gene in 54 basal cell carcinoma biopsies. Immunohistochemical staining demonstrated that ING1 is overexpressed in 25% (6/24) human basal cell carcinomas. Single-strand conformation polymorphism and DNA sequencing revealed that only 1 in 54 (1.8%) basal cell carcinoma primaries contained a missense mutation in the ING1 gene. The mutation is located in exon 2 and could thus potentially interfere with the structure of every ING1 isoforms and the functions of the PHD zinc finger motif. Our data indicate that overexpression and mutation of the ING1 gene are infrequent in human basal cell carcinoma.
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April 2003