Skip to main content

Advertisement

SpringerLink
Log in

Creation and characterization of BAC-transgenic mice with physiological overexpression of epitope-tagged RCAN1 (DSCR1)

  • Published:
Mammalian Genome Aims and scope Submit manuscript

Abstract

The chromosome 21 gene RCAN1, encoding a modulator of the calcineurin (CaN) phosphatase, is a candidate gene for contributing to cognitive disability in people with Down syndrome (DS; trisomy 21). To develop a physiologically relevant model for studying the biochemistry of RCAN1 and its contribution to DS, we generated bacterial artificial chromosome-transgenic (BAC-Tg) mouse lines containing the human RCAN1 gene with a C-terminal HA-FLAG epitope tag incorporated by recombineering. The BAC-Tg was expressed at levels only moderately higher than the native Rcan1 gene: approximately 1.5-fold in RCAN1 BAC-Tg1 and twofold in RCAN1 BAC-Tg2. Affinity purification of the RCAN1 protein complex from brains of these mice revealed a core complex of RCAN1 with CaN, glycogen synthase kinase 3-beta (Gsk3b), and calmodulin, with substoichiometric components, including LOC73419. The BAC-Tg mice are fully viable, but long-term synaptic potentiation is impaired in proportion to BAC-Tg dosage in hippocampal brain slices from these mice. RCAN1 can act as a tumor suppressor in some systems, but we found that the RCAN1 BAC-Tg did not reduce mammary cancer growth when present at a low copy number in Tp53;WAP-Cre mice. This work establishes a useful mouse model for investigating the biochemistry and dose-dependent functions of the RCAN1 protein in vivo.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Subscribe and save

Springer+ Basic
$34.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or eBook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

References

  • Ahn KJ, Jeong HK, Choi HS, Ryoo SR, Kim YJ, Goo JS, Choi SY, Han JS, Ha I, Song WJ (2006) DYRK1A BAC transgenic mice show altered synaptic plasticity with learning and memory defects. Neurobiol Dis 22:463–472

    Article  PubMed  CAS  Google Scholar 

  • Antonarakis SE, Lyle R, Dermitzakis ET, Reymond A, Deutsch S (2004) Chromosome 21 and Down syndrome: from genomics to pathophysiology. Nat Rev 5:725–738

    Article  CAS  Google Scholar 

  • Baek KH, Zaslavsky A, Lynch RC, Britt C, Okada Y, Siarey RJ, Lensch MW, Park IH, Yoon SS, Minami T, Korenberg JR, Folkman J, Daley GQ, Aird WC, Galdzicki Z, Ryeom S (2009) Down’s syndrome suppression of tumour growth and the role of the calcineurin inhibitor DSCR1. Nature 459:1126–1130

    Article  PubMed  CAS  Google Scholar 

  • Branchi I, Bichler Z, Minghetti L, Delabar JM, Malchiodi-Albedi F, Gonzalez MC, Chettouh Z, Nicolini A, Chabert C, Smith DJ, Rubin EM, Migliore-Samour D, Alleva E (2004) Transgenic mouse in vivo library of human Down syndrome critical region 1: association between DYRK1A overexpression, brain development abnormalities, and cell cycle protein alteration. J Neuropathol Exp Neurol 63:429–440

    PubMed  CAS  Google Scholar 

  • Chabert C, Jamon M, Cherfouh A, Duquenne V, Smith DJ, Rubin E, Roubertoux PL (2004) Functional analysis of genes implicated in Down syndrome: 1. Cognitive abilities in mice transpolygenic for Down syndrome chromosomal region-1 (DCR-1). Behav Genet 34:559–569

    Article  PubMed  Google Scholar 

  • Chan B, Greenan G, McKeon F, Ellenberger T (2005) Identification of a peptide fragment of DSCR1 that competitively inhibits calcineurin activity in vitro and in vivo. Proc Natl Acad Sci USA 102:13075–13080

    Article  PubMed  CAS  Google Scholar 

  • Chang KT, Shi YJ, Min KT (2003) The drosophila homolog of Down’s syndrome critical region 1 gene regulates learning: implications for mental retardation. Proc Natl Acad Sci USA 100:15794–15799

    Article  PubMed  CAS  Google Scholar 

  • Chen Z, Trotman LC, Shaffer D, Lin HK, Dotan ZA, Niki M, Koutcher JA, Scher HI, Ludwig T, Gerald W, Cordon-Cardo C, Pandolfi PP (2005) Crucial role of p53-dependent cellular senescence in suppression of Pten-deficient tumorigenesis. Nature 436:725–730

    Article  PubMed  CAS  Google Scholar 

  • Chrast R, Scott HS, Madani R, Huber L, Wolfer DP, Prinz M, Aguzzi A, Lipp HP, Antonarakis SE (2000) Mice trisomic for a bacterial artificial chromosome with the single-minded 2 gene (Sim2) show phenotypes similar to some of those present in the partial trisomy 16 mouse models of Down syndrome. Hum Mol Genet 9:1853–1864

    Article  PubMed  CAS  Google Scholar 

  • Cook CN, Hejna MJ, Magnuson DJ, Lee JM (2005) Expression of calcipressin1, an inhibitor of the phosphatase calcineurin, is altered with aging and Alzheimer’s disease. J Alzheimers Dis 8:63–73

    PubMed  CAS  Google Scholar 

  • Davies KJ, Ermak G, Rothermel BA, Pritchard M, Heitman J, Ahnn J, Henrique-Silva F, Crawford D, Canaider S, Strippoli P, Carinci P, Min KT, Fox DS, Cunningham KW, Bassel-Duby R, Olson EN, Zhang Z, Williams RS, Gerber HP, Perez-Riba M, Seo H, Cao X, Klee CB, Redondo JM, Maltais LJ, Bruford EA, Povey S, Molkentin JD, McKeon FD, Duh EJ, Crabtree GR, Cyert MS, de la Luna S, Estivill X (2007) Renaming the DSCR1/Adapt78 gene family as RCAN: regulators of calcineurin. FASEB J 21:3023–3028

    Article  PubMed  CAS  Google Scholar 

  • Dunbier AK, Anderson H, Ghazoui Z, Lopez-Knowles E, Pancholi S, Ribas R, Drury S, Sidhu K, Leary A, Martin LA, Dowsett M (2011) ESR1 is co-expressed with closely adjacent uncharacterised genes spanning a breast cancer susceptibility locus at 6q25.1. PLoS Genet 7:e1001382

    Article  PubMed  CAS  Google Scholar 

  • Ermak G, Morgan TE, Davies KJ (2001) Chronic overexpression of the calcineurin inhibitory gene DSCR1 (Adapt78) is associated with Alzheimer’s disease. J Biol Chem 276:38787–38794

    Article  PubMed  CAS  Google Scholar 

  • Fuentes JJ, Pritchard MA, Estivill X (1997) Genomic organization, alternative splicing, and expression patterns of the DSCR1 (Down syndrome candidate region 1) gene. Genomics 44:358–361

    Article  PubMed  CAS  Google Scholar 

  • Fuentes JJ, Genesca L, Kingsbury TJ, Cunningham KW, Perez-Riba M, Estivill X, de la Luna S (2000) DSCR1, overexpressed in Down syndrome, is an inhibitor of calcineurin-mediated signaling pathways. Hum Mol Genet 9:1681–1690

    Article  PubMed  CAS  Google Scholar 

  • Genesca L, Aubareda A, Fuentes JJ, Estivill X, De La Luna S, Perez-Riba M (2003) Phosphorylation of calcipressin 1 increases its ability to inhibit calcineurin and decreases calcipressin half-life. Biochem J 374:567–575

    Article  PubMed  CAS  Google Scholar 

  • Gu W, Malik S, Ito M, Yuan CX, Fondell JD, Zhang X, Martinez E, Qin J, Roeder RG (1999) A novel human SRB/MED-containing cofactor complex, SMCC, involved in transcription regulation. Mol Cell 3:97–108

    Article  PubMed  CAS  Google Scholar 

  • Hasle H (2001) Pattern of malignant disorders in individuals with Down’s syndrome. Lancet Oncol 2:429–436

    Article  PubMed  CAS  Google Scholar 

  • Herault Y, Duchon A, Velot E, Marechal D, Brault V (2012) The in vivo Down syndrome genomic library in mouse. Prog Brain Res 197:169–197

    Article  PubMed  CAS  Google Scholar 

  • Hesser BA, Liang XH, Camenisch G, Yang S, Lewin DA, Scheller R, Ferrara N, Gerber HP (2004) Down syndrome critical region protein 1 (DSCR1), a novel VEGF target gene that regulates expression of inflammatory markers on activated endothelial cells. Blood 104:149–158

    Article  PubMed  CAS  Google Scholar 

  • Hilioti Z, Gallagher DA, Low-Nam ST, Ramaswamy P, Gajer P, Kingsbury TJ, Birchwood CJ, Levchenko A, Cunningham KW (2004) GSK-3 kinases enhance calcineurin signaling by phosphorylation of RCNs. Genes Dev 18:35–47

    Article  PubMed  CAS  Google Scholar 

  • Hogan PG, Chen L, Nardone J, Rao A (2003) Transcriptional regulation by calcium, calcineurin, and NFAT. Genes Dev 17:2205–2232

    Article  PubMed  CAS  Google Scholar 

  • Iizuka M, Abe M, Shiiba K, Sasaki I, Sato Y (2004) Down syndrome candidate region 1, a downstream target of VEGF, participates in endothelial cell migration and angiogenesis. J Vasc Res 41:334–344

    Article  PubMed  CAS  Google Scholar 

  • Kingsbury TJ, Cunningham KW (2000) A conserved family of calcineurin regulators. Genes Dev 14:1595–1604

    PubMed  CAS  Google Scholar 

  • Kulnane LS, Lamb BT (2001) Neuropathological characterization of mutant amyloid precursor protein yeast artificial chromosome transgenic mice. Neurobiol Dis 8:982–992

    Article  PubMed  CAS  Google Scholar 

  • Kwak HI, Gustafson T, Metz RP, Laffin B, Schedin P, Porter WW (2007) Inhibition of breast cancer growth and invasion by single-minded 2s. Carcinogenesis 28:259–266

    Article  PubMed  CAS  Google Scholar 

  • Lamb BT, Sisodia SS, Lawler AM, Slunt HH, Kitt CA, Kearns WG, Pearson PL, Price DL, Gearhart JD (1993) Introduction and expression of the 400 kilobase amyloid precursor protein gene in transgenic mice [corrected]. Nat Genet 5:22–30

    Article  PubMed  CAS  Google Scholar 

  • Lange AW, Molkentin JD, Yutzey KE (2004) DSCR1 gene expression is dependent on NFATc1 during cardiac valve formation and colocalizes with anomalous organ development in trisomy 16 mice. Dev Biol 266:346–360

    Article  PubMed  CAS  Google Scholar 

  • Lange AW, Rothermel BA, Yutzey KE (2005) Restoration of DSCR1 to disomy in the trisomy 16 mouse model of Down syndrome does not correct cardiac or craniofacial development anomalies. Dev Dyn 233:954–963

    Article  PubMed  CAS  Google Scholar 

  • Lee EC, Yu D, Martinez de Velasco J, Tessarollo L, Swing DA, Court DL, Jenkins NA, Copeland NG (2001) A highly efficient Escherichia coli-based chromosome engineering system adapted for recombinogenic targeting and subcloning of BAC DNA. Genomics 73:56–65

    Article  PubMed  CAS  Google Scholar 

  • Lehman EJ, Kulnane LS, Gao Y, Petriello MC, Pimpis KM, Younkin L, Dolios G, Wang R, Younkin SG, Lamb BT (2003) Genetic background regulates beta-amyloid precursor protein processing and beta-amyloid deposition in the mouse. Hum Mol Genet 12:2949–2956

    Article  PubMed  CAS  Google Scholar 

  • Liu C, Belichenko PV, Zhang L, Fu D, Kleschevnikov AM, Baldini A, Antonarakis SE, Mobley WC, Yu YE (2011a) Mouse models for Down syndrome-associated developmental cognitive disabilities. Dev Neurosci 33:404–413

    Article  PubMed  CAS  Google Scholar 

  • Liu C, Morishima M, Yu T, Matsui S, Zhang L, Fu D, Pao A, Costa AC, Gardiner KJ, Cowell JK, Nowak NJ, Parmacek MS, Liang P, Baldini A, Yu YE (2011b) Genetic analysis of Down syndrome-associated heart defects in mice. Hum Genet 130:623–632

    Article  PubMed  CAS  Google Scholar 

  • Ludwig T, Fisher P, Murty V, Efstratiadis A (2001) Development of mammary adenocarcinomas by tissue-specific knockout of Brca2 in mice. Oncogene 20:3937–3948

    Article  PubMed  CAS  Google Scholar 

  • Ma H, Xiong H, Liu T, Zhang L, Godzik A, Zhang Z (2004) Aggregate formation and synaptic abnormality induced by DSCR1. J Neurochem 88:1485–1496

    Article  PubMed  CAS  Google Scholar 

  • Macian F (2005) NFAT proteins: key regulators of T-cell development and function. Nat Rev Immunol 5:472–484

    Article  PubMed  CAS  Google Scholar 

  • Minami T, Horiuchi K, Miura M, Abid MR, Takabe W, Noguchi N, Kohro T, Ge X, Aburatani H, Hamakubo T, Kodama T, Aird WC (2004) Vascular endothelial growth factor-and thrombin-induced termination factor, Down syndrome critical region-1, attenuates endothelial cell proliferation and angiogenesis. J Biol Chem 279:50537–50554

    Article  PubMed  CAS  Google Scholar 

  • Nikolaev AY, Li M, Puskas N, Qin J, Gu W (2003) Parc: a cytoplasmic anchor for p53. Cell 112:29–40

    Article  PubMed  CAS  Google Scholar 

  • Osoegawa K, Mammoser AG, Wu C, Frengen E, Zeng C, Catanese JJ, de Jong PJ (2001) A bacterial artificial chromosome library for sequencing the complete human genome. Genome Res 11:483–496

    Article  PubMed  CAS  Google Scholar 

  • Puzzo D, Privitera L, Leznik E, Fa M, Staniszewski A, Palmeri A, Arancio O (2008) Picomolar amyloid-beta positively modulates synaptic plasticity and memory in hippocampus. J Neurosci 28:14537–14545

    Article  PubMed  CAS  Google Scholar 

  • Qin L, Zhao D, Liu X, Nagy JA, Hoang MV, Brown LF, Dvorak HF, Zeng H (2006) Down syndrome candidate region 1 isoform 1 mediates angiogenesis through the calcineurin-NFAT pathway. Mol Cancer Res 4:811–820

    Article  PubMed  CAS  Google Scholar 

  • Reynolds LE, Watson AR, Baker M, Jones TA, D’Amico G, Robinson SD, Joffre C, Garrido-Urbani S, Rodriguez-Manzaneque JC, Martino-Echarri E, Aurrand-Lions M, Sheer D, Dagna-Bricarelli F, Nizetic D, McCabe CJ, Turnell AS, Kermorgant S, Imhof BA, Adams R, Fisher EM, Tybulewicz VL, Hart IR, Hodivala-Dilke KM (2010) Tumour angiogenesis is reduced in the Tc1 mouse model of Down’s syndrome. Nature 465:813–817

    Article  PubMed  CAS  Google Scholar 

  • Rothermel B, Vega RB, Yang J, Wu H, Bassel-Duby R, Williams RS (2000) A protein encoded within the Down syndrome critical region is enriched in striated muscles and inhibits calcineurin signaling. J Biol Chem 275:8719–8725

    Article  PubMed  CAS  Google Scholar 

  • Ryeom S, Baek KH, Rioth MJ, Lynch RC, Zaslavsky A, Birsner A, Yoon SS, McKeon F (2008) Targeted deletion of the calcineurin inhibitor DSCR1 suppresses tumor growth. Cancer Cell 13:420–431

    Article  PubMed  CAS  Google Scholar 

  • Sanna B, Brandt EB, Kaiser RA, Pfluger P, Witt SA, Kimball TR, van Rooij E, De Windt LJ, Rothenberg ME, Tschop MH, Benoit SC, Molkentin JD (2006) Modulatory calcineurin-interacting proteins 1 and 2 function as calcineurin facilitators in vivo. Proc Natl Acad Sci USA 103:7327–7332

    Article  PubMed  CAS  Google Scholar 

  • Smith DJ, Rubin EM (1997) Functional screening and complex traits: human 21q22.2 sequences affecting learning in mice. Hum Mol Genet 6:1729–1733

    Article  PubMed  CAS  Google Scholar 

  • Smith DJ, Stevens ME, Sudanagunta SP, Bronson RT, Makhinson M, Watabe AM, O’Dell TJ, Fung J, Weier HU, Cheng JF, Rubin EM (1997) Functional screening of 2 Mb of human chromosome 21q22.2 in transgenic mice implicates minibrain in learning defects associated with Down syndrome. Nat Genet 16:28–36

    Article  PubMed  CAS  Google Scholar 

  • Stewart MP, McDowall A, Hogg N (1998) LFA-1-mediated adhesion is regulated by cytoskeletal restraint and by a Ca2 + -dependent protease, calpain. J Cell Biol 140:699–707

    Article  PubMed  CAS  Google Scholar 

  • Sussan TE, Yang A, Li F, Ostrowski MC, Reeves RH (2008) Trisomy represses Apc(Min)-mediated tumours in mouse models of Down’s syndrome. Nature 451:73–75

    Article  PubMed  CAS  Google Scholar 

  • Vega RB, Rothermel BA, Weinheimer CJ, Kovacs A, Naseem RH, Bassel-Duby R, Williams RS, Olson EN (2003) Dual roles of modulatory calcineurin-interacting protein 1 in cardiac hypertrophy. Proc Natl Acad Sci USA 100:669–674

    Article  PubMed  CAS  Google Scholar 

  • Voronov SV, Frere SG, Giovedi S, Pollina EA, Borel C, Zhang H, Schmidt C, Akeson EC, Wenk MR, Cimasoni L, Arancio O, Davisson MT, Antonarakis SE, Gardiner K, De Camilli P, Di Paolo G (2008) Synaptojanin 1-linked phosphoinositide dyshomeostasis and cognitive deficits in mouse models of Down’s syndrome. Proc Natl Acad Sci USA 105:9415–9420

    Article  PubMed  CAS  Google Scholar 

  • Winder DG, Sweatt JD (2001) Roles of serine/threonine phosphatases in hippocampal synaptic plasticity. Nat Rev Neurosci 2:461–474

    Article  PubMed  CAS  Google Scholar 

  • Wiseman FK, Alford KA, Tybulewicz VL, Fisher EM (2009) Down syndrome––recent progress and future prospects. Hum Mol Genet 18:R75–R83

    Article  PubMed  CAS  Google Scholar 

  • Wolvetang EJ, Wilson TJ, Sanij E, Busciglio J, Hatzistavrou T, Seth A, Hertzog PJ, Kola I (2003) ETS2 overexpression in transgenic models and in Down syndrome predisposes to apoptosis via the p53 pathway. Hum Mol Genet 12:247–255

    Article  PubMed  CAS  Google Scholar 

  • Xing L, Salas M, Lin CS, Zigman W, Silverman W, Subramaniyam S, Murty VV, Tycko B (2007) Faithful tissue-specific expression of the human chromosome 21-linked COL6A1 gene in BAC-transgenic mice. Mamm Genome 18:113–122

    Article  PubMed  CAS  Google Scholar 

  • Yang A, Reeves RH (2011) Increased survival following tumorigenesis in Ts65Dn mice that model Down syndrome. Cancer Res 71:3573–3581

    Article  PubMed  CAS  Google Scholar 

  • Yang J, Rothermel B, Vega RB, Frey N, McKinsey TA, Olson EN, Bassel-Duby R, Williams RS (2000) Independent signals control expression of the calcineurin inhibitory proteins MCIP1 and MCIP2 in striated muscles. Circ Res 87:E61–E68

    Article  PubMed  CAS  Google Scholar 

  • Zeng H, Chattarji S, Barbarosie M, Rondi-Reig L, Philpot BD, Miyakawa T, Bear MF, Tonegawa S (2001) Forebrain-specific calcineurin knockout selectively impairs bidirectional synaptic plasticity and working/episodic-like memory. Cell 107:617–629

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

This work was supported by grants from the NIH (P01-HD035897 to BT and WS, and R01-AG034248 to OA).

Author information

Author notes

  1. Thomas Ludwig

    Present address: Department of Molecular & Cellular Biochemistry, Ohio State University Medical Center, Columbus, OH, USA

Authors and Affiliations

  1. Institute for Cancer Genetics, Columbia University Medical Center, Herbert Irving Cancer Research Building, Room 607, New York, NY, 10032, USA

    Luzhou Xing, Martha Salas, Julia Gittler, Thomas Ludwig, Vundavalli V. Murty & Benjamin Tycko

  2. Department of Pathology, Columbia University Medical Center, New York, NY, 10032, USA

    Hong Zhang, Vundavalli V. Murty, Ottavio Arancio & Benjamin Tycko

  3. Herbert Irving Comprehensive Cancer Center, Columbia University Medical Center, New York, NY, 10032, USA

    Chyuan-Sheng Lin, Vundavalli V. Murty & Benjamin Tycko

  4. Kennedy-Krieger Institute, Baltimore, MD, 21202, USA

    Wayne Silverman

  5. Taub Institute for Research on Alzheimer’s Disease and the Aging Brain, Columbia University Medical Center, New York, NY, 10032, USA

    Ottavio Arancio & Benjamin Tycko

Authors
  1. Luzhou Xing
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Martha Salas
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Hong Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Julia Gittler
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Thomas Ludwig
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Chyuan-Sheng Lin
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Vundavalli V. Murty
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Wayne Silverman
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Ottavio Arancio
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Benjamin Tycko
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Benjamin Tycko.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (PDF 712 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Xing, L., Salas, M., Zhang, H. et al. Creation and characterization of BAC-transgenic mice with physiological overexpression of epitope-tagged RCAN1 (DSCR1). Mamm Genome 24, 30–43 (2013). https://doi.org/10.1007/s00335-012-9436-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00335-012-9436-9

Keywords

Search

Navigation