32-5283: Recombinant Human ZW10 Interacting Kinetochore Protein

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32-5283: Recombinant Human ZW10 Interacting Kinetochore Protein

9853 Pacific Heights Blvd. Suite D. San Diego, CA 92121, USA Tel: 858-263-4982 Email: [email protected] 32-5283: Recombinant Human ZW10 Interacting Kinetochore Protein Alternative ZW10 Interacting Kinetochore Protein,ZWINT,ZW10 Interactor,HZwint-1,KNTC2AP,ZWINT1,Human ZW10 Name : Interacting Protein-1,ZW10 Interactor Kinetochore Protein zwint-1,Zwint-1,ZW10-Interacting Protein 1. Description Source : Escherichia Coli. ZWINT Human Recombinant produced in E.coli is a single, non-glycosylated polypeptide chain containing 253 amino acids (1-230) and having a molecular mass of 27.9 kDa (Molecular size on SDS-PAGE will appear higher).ZWINT is fused to a 23 amino acid His-tag at N-terminus & purified by proprietary chromatographic techniques. ZW10 Interacting Kinetochore Protein (ZWINT) is involved in kinetochore function. ZWINT is part of the MIS12 complex, which is essential for kinetochore formation and spindle checkpoint activity. ZWINT is localized to the cytoplasm during interphase and to kinetochores from late prophase to anaphase. ZWINT interacts with ZW10 (Zeste White 10) and regulates the association between ZW10 and kinetochores. ZWINT gene defects are linked with the pathogenesis of Roberts's syndrome, an autosomal recessive disorder characterized by growth retardation due to premature chromosome separation. Product Info Amount : 10 µg Purification : Greater than 85.0% as determined by SDS-PAGE. The ZWINT solution (0.25mg/ml) contains 20mM Tris-HCl buffer (pH 8.0), 0.15M NaCl, 20% Content : glycerol and 1mM DTT. Store at 4°C if entire vial will be used within 2-4 weeks. Store, frozen at -20°C for longer periods of Storage condition : time. For long term storage it is recommended to add a carrier protein (0.1% HSA or BSA).Avoid multiple freeze-thaw cycles. Amino Acid : MGSSHHHHHH SSGLVPRGSH MGSMEAAETE AEAAALEVLA EVAGILEPVG LQEEAELPAK ILVEFVVDSQ KKDKLLCSQL QVADFLQNIL AQEDTAKGLD PLASEDTSRQ KAIAAKEQWK ELKATYREHV EAIKIGLTKA LTQMEEAQRK RTQLREAFEQ LQAKKQMAME KRRAVQNQWQ LQQEKHLQHL AEVSAEGKLL FPEAEAEAEN LPDDKPQQPT RPQEQSTGDT MGRDPGVSFK AVGLQPAGDV NLP. For Research Use Only. Not for use in diagnostic/therapeutics procedures..

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PARSANA-DISSERTATION-2020.Pdf
DECIPHERING TRANSCRIPTIONAL PATTERNS OF GENE REGULATION: A COMPUTATIONAL APPROACH by Princy Parsana A dissertation submitted to The Johns Hopkins University in conformity with the requirements for the degree of Doctor of Philosophy Baltimore, Maryland July, 2020 © 2020 Princy Parsana All rights reserved Abstract With rapid advancements in sequencing technology, we now have the ability to sequence the entire human genome, and to quantify expression of tens of thousands of genes from hundreds of individuals. This provides an extraordinary opportunity to learn phenotype relevant genomic patterns that can improve our understanding of molecular and cellular processes underlying a trait. The high dimensional nature of genomic data presents a range of computational and statistical challenges. This dissertation presents a compilation of projects that were driven by the motivation to efficiently capture gene regulatory patterns in the human transcriptome, while addressing statistical and computational challenges that accompany this data. We attempt to address two major difficulties in this domain: a) artifacts and noise in transcriptomic data, andb) limited statistical power. First, we present our work on investigating the effect of artifactual variation in gene expression data and its impact on trans-eQTL discovery. Here we performed an in-depth analysis of diverse pre-recorded covariates and latent confounders to understand their contribution to heterogeneity in gene expression measurements. Next, we discovered 673 trans-eQTLs across 16 human tissues using v6 data from the Genotype Tissue Expression (GTEx) project. Finally, we characterized two trait-associated trans-eQTLs; one in Skeletal Muscle and another in Thyroid. Second, we present a principal component based residualization method to correct gene expression measurements prior to reconstruction of co-expression networks.
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TANGO1 Builds a Machine for Collagen Export by Recruiting And
RESEARCH ARTICLE TANGO1 builds a machine for collagen export by recruiting and spatially organizing COPII, tethers and membranes Ishier Raote1,2†, Maria Ortega-Bellido1,2†, Anto´ nio JM Santos1,2‡, Ombretta Foresti1,2, Chong Zhang3, Maria F Garcia-Parajo4,5, Felix Campelo4, Vivek Malhotra1,2,5* 1Centre for Genomic Regulation, The Barcelona Institute of Science and Technology, Barcelona, Spain; 2Universitat Pompeu Fabra, Barcelona, Spain; 3SIMBIOsys Group, Department of Information and Communication Technologies, Universitat Pompeu Fabra, Barcelona, Spain; 4ICFO-Institut de Ciencies Fotoniques, The Barcelona Institute of Science and Technology, Castelldefels, Spain; 5Institucio Catalana de Recerca i Estudis Avanc¸ats, Barcelona, Spain Abstract Collagen export from the endoplasmic reticulum (ER) requires TANGO1, COPII coats, and retrograde fusion of ERGIC membranes. How do these components come together to produce a transport carrier commensurate with the bulky cargo collagen? TANGO1 is known to form a ring that corrals COPII coats, and we show here how this ring or fence is assembled. Our data reveal that a TANGO1 ring is organized by its radial interaction with COPII, and lateral interactions with *For correspondence: cTAGE5, TANGO1-short or itself. Of particular interest is the finding that TANGO1 recruits ERGIC [email protected] membranes for collagen export via the NRZ (NBAS/RINT1/ZW10) tether complex. Therefore, †These authors contributed TANGO1 couples retrograde membrane flow to anterograde cargo transport. Without the NRZ equally to this work complex, the TANGO1 ring does not assemble, suggesting its role in nucleating or stabilising this Present address: ‡Division of process. Thus, coordinated capture of COPII coats, cTAGE5, TANGO1-short, and tethers by Hematology, Department of TANGO1 assembles a collagen export machine at the ER.
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Supplemental Information
Supplemental information Dissection of the genomic structure of the miR-183/96/182 gene. Previously, we showed that the miR-183/96/182 cluster is an intergenic miRNA cluster, located in a ~60-kb interval between the genes encoding nuclear respiratory factor-1 (Nrf1) and ubiquitin-conjugating enzyme E2H (Ube2h) on mouse chr6qA3.3 (1). To start to uncover the genomic structure of the miR- 183/96/182 gene, we first studied genomic features around miR-183/96/182 in the UCSC genome browser (http://genome.UCSC.edu/), and identified two CpG islands 3.4-6.5 kb 5’ of pre-miR-183, the most 5’ miRNA of the cluster (Fig. 1A; Fig. S1 and Seq. S1). A cDNA clone, AK044220, located at 3.2-4.6 kb 5’ to pre-miR-183, encompasses the second CpG island (Fig. 1A; Fig. S1). We hypothesized that this cDNA clone was derived from 5’ exon(s) of the primary transcript of the miR-183/96/182 gene, as CpG islands are often associated with promoters (2). Supporting this hypothesis, multiple expressed sequences detected by gene-trap clones, including clone D016D06 (3, 4), were co-localized with the cDNA clone AK044220 (Fig. 1A; Fig. S1). Clone D016D06, deposited by the German GeneTrap Consortium (GGTC) (http://tikus.gsf.de) (3, 4), was derived from insertion of a retroviral construct, rFlpROSAβgeo in 129S2 ES cells (Fig. 1A and C). The rFlpROSAβgeo construct carries a promoterless reporter gene, the β−geo cassette - an in-frame fusion of the β-galactosidase and neomycin resistance (Neor) gene (5), with a splicing acceptor (SA) immediately upstream, and a polyA signal downstream of the β−geo cassette (Fig.
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Kinetochores, Microtubules, and Spindle Assembly Checkpoint
Review Joined at the hip: kinetochores, microtubules, and spindle assembly checkpoint signaling 1 1,2,3 Carlos Sacristan and Geert J.P.L. Kops 1 Molecular Cancer Research, University Medical Center Utrecht, 3584 CG Utrecht, The Netherlands 2 Center for Molecular Medicine, University Medical Center Utrecht, 3584 CG Utrecht, The Netherlands 3 Cancer Genomics Netherlands, University Medical Center Utrecht, 3584 CG Utrecht, The Netherlands Error-free chromosome segregation relies on stable and cell division. The messenger is the SAC (also known as connections between kinetochores and spindle microtu- the mitotic checkpoint) (Figure 1). bules. The spindle assembly checkpoint (SAC) monitors The transition to anaphase is triggered by the E3 ubiqui- such connections and relays their absence to the cell tin ligase APC/C, which tags inhibitors of mitotic exit cycle machinery to delay cell division. The molecular (CYCLIN B) and of sister chromatid disjunction (SECURIN) network at kinetochores that is responsible for microtu- for proteasomal degradation [2]. The SAC has a one-track bule binding is integrated with the core components mind, inhibiting APC/C as long as incorrectly attached of the SAC signaling system. Molecular-mechanistic chromosomes persist. It goes about this in the most straight- understanding of how the SAC is coupled to the kineto- forward way possible: it assembles a direct and diffusible chore–microtubule interface has advanced signiﬁcantly inhibitor of APC/C at kinetochores that are not connected in recent years. The latest insights not only provide a to spindle microtubules. This inhibitor is named the striking view of the dynamics and regulation of SAC mitotic checkpoint complex (MCC) (Figure 1).
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Differential Expression of ZWINT in Cancers of the Breast
Differential expression of ZW10 interacting kinetochore protein in cancers of the breast. Shahan Mamoor, MS1 [email protected] East Islip, NY 11730 Breast cancer affects women at relatively high frequency1. We mined published microarray datasets2,3 to determine in an unbiased fashion and at the systems level genes most differentially expressed in the primary tumors of patients with breast cancer. We report here significant differential expression of the gene encoding the ZW10 interacting kinetochore protein, ZWINT, when comparing primary tumors of the breast to the tissue of origin, the normal breast. ZWINT was also differentially expressed in the tumor cells of patients with triple negative breast cancer. ZWINT mRNA was present at significantly higher quantities in tumors of the breast as compared to normal breast tissue. Analysis of human survival data revealed that expression of ZWINT in primary tumors of the breast was correlated with overall survival in patients with basal and luminal A subtype cancer, but in a contrary manner, demonstrating a complex relationship between correlation of primary tumor expression with overall survival based on molecular subtype. ZWINT may be of relevance to initiation, maintenance or progression of cancers of the female breast. Keywords: breast cancer, ZWINT, ZW10 interacting kinetochore protein, systems biology of breast cancer, targeted therapeutics in breast cancer. 1 Invasive breast cancer is diagnosed in over a quarter of a million women in the United States each year1 and in 2018, breast cancer was the leading cause of cancer death in women worldwide4. While patients with localized breast cancer are provided a 99% 5-year survival rate, patients with regional breast cancer, cancer that has spread to lymph nodes or nearby structures, are provided an 86% 5-year survival rate5,6.
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Product Name: ZW10 Peptide Polyclonal Antibody, ALEXA FLUOR® 594 Conjugated Catalog No
Product Name: ZW10 peptide Polyclonal Antibody, ALEXA FLUOR® 594 Conjugated Catalog No. : TAP01-98062R-A594 Intended Use: For Research Use Only. Not for used in diagnostic procedures. Size 100ul Concentration 1ug/ul Gene ID 9183 ISO Type Rabbit IgG Clone N/A Immunogen Range Conjugation ALEXA FLUOR® 594 Subcellular Locations Applications IF(IHC-P) Cross Reactive Species Human, Mouse, Rat Source KLH conjugated synthetic peptide derived from mouse ZW10 Applications with IF(IHC-P)(1:50-200) Dilutions Purification Purified by Protein A. Background The mitotic checkpoint ensures that chromosomes are divided equally between daughter cells and is a primary mechanism preventing the chromosome instability often seen in aneuploid human tumors. This gene encodes a protein that is one of many involved in mechanisms to ensure proper chromosome segregation during cell division. The encoded protein binds to centromeres during the prophase, metaphase, and early anaphase cell division stages and to kinetochore microtubules during metaphase. It is part of the MIS12 complex, which may be fundamental for kinetochore formation and proper chromosome segregation during mitosis. In mitotic human cells ZW10 resides in a complex with Rod and Zwilch, whereas another ZW10 partner, Zwint-1, is part of a separate complex ofstructural kinetochore components including Mis12 and Ndc80-Hec1. Zwint-1 is critical for recruiting ZW10 to unattached kinetochores. Depletion from human cells demonstrates that the ZW10 complex is essential for stable binding of a Mad1-Mad2 complex tounattached kinetochores. Thus, ZW10 functions as a linker between the core structural elements of the outer kinetochore and components that catalyze generation of the mitotic checkpoint-derived "stop anaphase" inhibitor.
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Team Publications Biology of Centrosomes and Genetic Instability
Team Publications Biology of centrosomes and genetic instability Year of publication 2001 E Wojcik, R Basto, M Serr, F Scaërou, R Karess, T Hays (2001 Nov 21) Kinetochore dynein: its dynamics and role in the transport of the Rough deal checkpoint protein. Nature cell biology : 1001-7 Summary We describe the dynamics of kinetochore dynein-dynactin in living Drosophila embryos and examine the effect of mutant dynein on the metaphase checkpoint. A functional conjugate of dynamitin with green fluorescent protein accumulates rapidly at prometaphase kinetochores, and subsequently migrates off kinetochores towards the poles during late prometaphase and metaphase. This behaviour is seen for several metaphase checkpoint proteins, including Rough deal (Rod). In neuroblasts, hypomorphic dynein mutants accumulate in metaphase and block the normal redistribution of Rod from kinetochores to microtubules. By transporting checkpoint proteins away from correctly attached kinetochores, dynein might contribute to shutting off the metaphase checkpoint, allowing anaphase to ensue. R Basto, R Gomes, R E Karess (2001 Jan 9) Rough deal and Zw10 are required for the metaphase checkpoint in Drosophila. Nature cell biology : 939-43 Summary The metaphase-anaphase transition during mitosis is carefully regulated in order to assure high-fidelity transmission of genetic information to the daughter cells. A surveillance mechanism known as the metaphase checkpoint (or spindle-assembly checkpoint) monitors the attachment of kinetochores to the spindle microtubules, and inhibits anaphase onset until all chromosomes have achieved a proper bipolar orientation on the spindle. Defects in this checkpoint lead to premature anaphase onset, and consequently to greatly increased rates of aneuploidy. Here we show that the Drosophila kinetochore components Rough deal (Rod) and Zeste-White 10 (Zw10) are required for the proper functioning of the metaphase checkpoint in flies.
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Bipolar Spindle Attachments Affect Redistributions of ZW10, a Drosophila Centromere/Kinetochore Component Required for Accurate Chromosome Segregation Byron C
Bipolar Spindle Attachments Affect Redistributions of ZW10, a Drosophila Centromere/Kinetochore Component Required for Accurate Chromosome Segregation Byron C. Williams,* Maurizio Gatti,* and Michael L. Goldberg* *Section of Genetics and Development, Cornell University, Ithaca, New York 14853-2703; andHstituto Pasteur-Fondazione Cenci Bolognetti, Dipartimento di Genetica e Biologia Molecolare, Universit~ di Roma "La Sapienza," 00185 Rome, Italy Abstract. Previous efforts have shown that mutations (during meiosis II). During metaphase of both divi- in the Drosophila zwlO gene cause massive chromo- sions, ZW10 appears to move from the kinetochores some missegregation during mitotic divisions in several and to spread toward the poles along what appear to be tissues. Here we demonstrate that mutations in zwlO kinetochore microtubules. Redistributions of ZWl0 at also disrupt chromosome behavior in male meiosis I metaphase require bipolar attachments of individual and meiosis II, indicating that ZW10 function is com- chromosomes or paired bivalents to the spindle. At the mon to both equational and reductional divisions. Divi- onset of anaphase I or anaphase II, ZWl0 rapidly relo- sions are apparently normal before anaphase onset, but calizes to the kinetochore regions of the separating ZW10 mutants exhibit lagging chromosomes and irreg- chromosomes. In other mutant backgrounds in which ular chromosome segregation at anaphase. Chromo- chromosomes lag during anaphase, the presence or ab- some missegregation during meiosis I of these mutants sence
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Biological Role and Disease Impact of Copy Number Variation in Complex Disease
University of Pennsylvania ScholarlyCommons Publicly Accessible Penn Dissertations 2014 Biological Role and Disease Impact of Copy Number Variation in Complex Disease Joseph Glessner University of Pennsylvania, [email protected] Follow this and additional works at: https://repository.upenn.edu/edissertations Part of the Bioinformatics Commons, and the Genetics Commons Recommended Citation Glessner, Joseph, "Biological Role and Disease Impact of Copy Number Variation in Complex Disease" (2014). Publicly Accessible Penn Dissertations. 1286. https://repository.upenn.edu/edissertations/1286 This paper is posted at ScholarlyCommons. https://repository.upenn.edu/edissertations/1286 For more information, please contact [email protected]. Biological Role and Disease Impact of Copy Number Variation in Complex Disease Abstract In the human genome, DNA variants give rise to a variety of complex phenotypes. Ranging from single base mutations to copy number variations (CNVs), many of these variants are neutral in selection and disease etiology, making difficult the detection of true common orar r e frequency disease-causing mutations. However, allele frequency comparisons in cases, controls, and families may reveal disease associations. Single nucleotide polymorphism (SNP) arrays and exome sequencing are popular assays for genome-wide variant identification. oT limit bias between samples, uniform testing is crucial, including standardized platform versions and sample processing. Bases occupy single points while copy variants occupy segments.
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A High-Throughput Approach to Uncover Novel Roles of APOBEC2, a Functional Orphan of the AID/APOBEC Family
Rockefeller University Digital Commons @ RU Student Theses and Dissertations 2018 A High-Throughput Approach to Uncover Novel Roles of APOBEC2, a Functional Orphan of the AID/APOBEC Family Linda Molla Follow this and additional works at: https://digitalcommons.rockefeller.edu/ student_theses_and_dissertations Part of the Life Sciences Commons A HIGH-THROUGHPUT APPROACH TO UNCOVER NOVEL ROLES OF APOBEC2, A FUNCTIONAL ORPHAN OF THE AID/APOBEC FAMILY A Thesis Presented to the Faculty of The Rockefeller University in Partial Fulfillment of the Requirements for the degree of Doctor of Philosophy by Linda Molla June 2018 © Copyright by Linda Molla 2018 A HIGH-THROUGHPUT APPROACH TO UNCOVER NOVEL ROLES OF APOBEC2, A FUNCTIONAL ORPHAN OF THE AID/APOBEC FAMILY Linda Molla, Ph.D. The Rockefeller University 2018 APOBEC2 is a member of the AID/APOBEC cytidine deaminase family of proteins. Unlike most of AID/APOBEC, however, APOBEC2’s function remains elusive. Previous research has implicated APOBEC2 in diverse organisms and cellular processes such as muscle biology (in Mus musculus), regeneration (in Danio rerio), and development (in Xenopus laevis). APOBEC2 has also been implicated in cancer. However the enzymatic activity, substrate or physiological target(s) of APOBEC2 are unknown. For this thesis, I have combined Next Generation Sequencing (NGS) techniques with state-of-the-art molecular biology to determine the physiological targets of APOBEC2. Using a cell culture muscle differentiation system, and RNA sequencing (RNA-Seq) by polyA capture, I demonstrated that unlike the AID/APOBEC family member APOBEC1, APOBEC2 is not an RNA editor. Using the same system combined with enhanced Reduced Representation Bisulfite Sequencing (eRRBS) analyses I showed that, unlike the AID/APOBEC family member AID, APOBEC2 does not act as a 5-methyl-C deaminase.
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ZWINT Antibody (Monoclonal) (M04) Mouse Monoclonal Antibody Raised Against a Full-Length Recombinant ZWINT
10320 Camino Santa Fe, Suite G San Diego, CA 92121 Tel: 858.875.1900 Fax: 858.622.0609 ZWINT Antibody (monoclonal) (M04) Mouse monoclonal antibody raised against a full-length recombinant ZWINT. Catalog # AT4652a Specification ZWINT Antibody (monoclonal) (M04) - Product Information Application WB, E Primary Accession O95229 Other Accession BC020979 Reactivity Human Host mouse Clonality Monoclonal Isotype IgG1 Kappa Calculated MW 31293 ZWINT Antibody (monoclonal) (M04) - Additional Information Antibody Reactive Against Recombinant Protein.Western Blot detection against Gene ID 11130 Immunogen (56.21 KDa) . Other Names ZW10 interactor, ZW10-interacting protein 1, Zwint-1, ZWINT Target/Specificity ZWINT (AAH20979, 1 a.a. ~ 277 a.a) full-length recombinant protein with GST tag. MW of the GST tag alone is 26 KDa. Dilution WB~~1:500~1000 Format Clear, colorless solution in phosphate buffered saline, pH 7.2 . ZWINT monoclonal antibody (M04), clone 1B7. Western Blot analysis of ZWINT Storage expression in K-562 ( (Cat # AT4652a ) Store at -20°C or lower. Aliquot to avoid repeated freezing and thawing. Precautions ZWINT Antibody (monoclonal) (M04) is for research use only and not for use in diagnostic or therapeutic procedures. ZWINT Antibody (monoclonal) (M04) - Protocols Detection limit for recombinant GST tagged Page 1/2 10320 Camino Santa Fe, Suite G San Diego, CA 92121 Tel: 858.875.1900 Fax: 858.622.0609 Provided below are standard protocols that you ZWINT is 0.03 ng/ml as a capture antibody. may find useful for product applications. • Western Blot ZWINT Antibody (monoclonal) (M04) - • Blocking Peptides Background • Dot Blot • Immunohistochemistry This gene encodes a protein that is clearly • Immunofluorescence involved in kinetochore function although an • Immunoprecipitation exact role is not known.
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Supplementary Data
Supplemental figures Supplemental figure 1: Tumor sample selection. A total of 98 thymic tumor specimens were stored in Memorial Sloan-Kettering Cancer Center tumor banks during the study period. 64 cases corresponded to previously untreated tumors, which were resected upfront after diagnosis. Adjuvant treatment was delivered in 7 patients (radiotherapy in 4 cases, cyclophosphamide- doxorubicin-vincristine (CAV) chemotherapy in 3 cases). 34 tumors were resected after induction treatment, consisting of chemotherapy in 16 patients (cyclophosphamide-doxorubicin- cisplatin (CAP) in 11 cases, cisplatin-etoposide (PE) in 3 cases, cisplatin-etoposide-ifosfamide (VIP) in 1 case, and cisplatin-docetaxel in 1 case), in radiotherapy (45 Gy) in 1 patient, and in sequential chemoradiation (CAP followed by a 45 Gy-radiotherapy) in 1 patient. Among these 34 patients, 6 received adjuvant radiotherapy. 1 Supplemental Figure 2: Amino acid alignments of KIT H697 in the human protein and related orthologs, using (A) the Homologene database (exons 14 and 15), and (B) the UCSC Genome Browser database (exon 14). Residue H697 is highlighted with red boxes. Both alignments indicate that residue H697 is highly conserved. 2 Supplemental Figure 3: Direct comparison of the genomic profiles of thymic squamous cell carcinomas (n=7) and lung primary squamous cell carcinomas (n=6). (A) Unsupervised clustering analysis. Gains are indicated in red, and losses in green, by genomic position along the 22 chromosomes. (B) Genomic profiles and recurrent copy number alterations in thymic carcinomas and lung squamous cell carcinomas. Gains are indicated in red, and losses in blue. 3 Supplemental Methods Mutational profiling The exonic regions of interest (NCBI Human Genome Build 36.1) were broken into amplicons of 500 bp or less, and specific primers were designed using Primer 3 (on the World Wide Web for general users and for biologist programmers (see Supplemental Table 2) [1].
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