The Cerebellum

, Volume 16, Issue 2, pp 462–472

Defining Trends in Global Gene Expression in Arabian Horses with Cerebellar Abiotrophy

Original Paper

Abstract

Equine cerebellar abiotrophy (CA) is a hereditary neurodegenerative disease that affects the Purkinje neurons of the cerebellum and causes ataxia in Arabian foals. Signs of CA are typically first recognized either at birth to any time up to 6 months of age. CA is inherited as an autosomal recessive trait and is associated with a single nucleotide polymorphism (SNP) on equine chromosome 2 (13074277G>A), located in the fourth exon of TOE1 and in proximity to MUTYH on the antisense strand. We hypothesize that unraveling the functional consequences of the CA SNP using RNA-seq will elucidate the molecular pathways underlying the CA phenotype. RNA-seq (100 bp PE strand-specific) was performed in cerebellar tissue from four CA-affected and five age-matched unaffected horses. Three pipelines for differential gene expression (DE) analysis were used (Tophat2/Cuffdiff2, Kallisto/EdgeR, and Kallisto/Sleuth) with 151 significant DE genes identified by all three pipelines in CA-affected horses. TOE1 (Log2(foldchange) = 0.92, p = 0.66) and MUTYH (Log2(foldchange) = 1.13, p = 0.66) were not differentially expressed. Among the major pathways that were differentially expressed, genes associated with calcium homeostasis and specifically expressed in Purkinje neurons, CALB1 (Log2(foldchange) = −1.7, p < 0.01) and CA8 (Log2(foldchange) = −0.97, p < 0.01), were significantly down-regulated, confirming loss of Purkinje neurons. There was also a significant up-regulation of markers for microglial phagocytosis, TYROBP (Log2(foldchange) = 1.99, p < 0.01) and TREM2 (Log2(foldchange) = 2.02, p < 0.01). These findings reaffirm a loss of Purkinje neurons in CA-affected horses along with a potential secondary loss of granular neurons and activation of microglial cells.

Keywords

Cerebellar abiotrophy Horse Calcium Microglial activation RNA-seq 

Supplementary material

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References

  1. 1.
    de Lahunta A. Abiotrophy in domestic animals: a review. Can J Vet Res. 1990;54(1):65–76.PubMedPubMedCentralGoogle Scholar
  2. 2.
    Koehler JW, Newcomer BW, Holland M, Caldwell JM. A novel inherited cerebellar abiotrophy in a cohort of related goats. J Comp Pathol. 2015;153(2–3):135–9.CrossRefPubMedGoogle Scholar
  3. 3.
    Sato J, Yamada N, Kobayashi R, Tsuchitani M, Kobayashi Y. Morphometric analysis of progressive changes in hereditary cerebellar cortical degenerative disease (abiotrophy) in rabbits caused by abnormal synaptogenesis. J Toxicol Pathol. 2015;28(2):73–8.CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Shearman JR, Cook RW, McCowan C, Fletcher JL, Taylor RM, Wilton AN. Mapping cerebellar abiotrophy in Australian kelpies. Anim Genet. 2011;42(6):675–8.CrossRefPubMedGoogle Scholar
  5. 5.
    DeBowes RM, Leipold HW, Turner-Beatty M. Cerebellar abiotrophy. Vet Clin North Am Equine Pract. 1987;3(2):345–52.PubMedGoogle Scholar
  6. 6.
    Blanco A, Moyano R, Vivo J, Flores-Acuna R, Molina A, Blanco C, et al. Purkinje cell apoptosis in Arabian horses with cerebellar abiotrophy. J Vet Med A Physiol Pathol Clin Med. 2006;53(6):286–7.CrossRefPubMedGoogle Scholar
  7. 7.
    Cavalleri JM, Metzger J, Hellige M, Lampe V, Stuckenschneider K, Tipold A, et al. Morphometric magnetic resonance imaging and genetic testing in cerebellar abiotrophy in Arabian horses. BMC Vet Res. 2013;9:105.CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Palmer AC, Blakemore WF, Cook WR, Platt H, Whitwell KE. Cerebellar hypoplasia and degeneration in the young Arab horse: clinical and neuropathological features. Vet Rec. 1973;93(3):62–6.CrossRefPubMedGoogle Scholar
  9. 9.
    Brault LS, Cooper CA, Famula TR, Murray JD, Penedo MC. Mapping of equine cerebellar abiotrophy to ECA2 and identification of a potential causative mutation affecting expression of MUTYH. Genomics. 2011;97(2):121–9.CrossRefPubMedGoogle Scholar
  10. 10.
    Brault LS, Famula TR, Penedo MC. Inheritance of cerebellar abiotrophy in Arabians. Am J Vet Res. 2011;72(7):940–4.CrossRefPubMedGoogle Scholar
  11. 11.
    Wagner E, Clement SL, Lykke-Andersen J. An unconventional human Ccr4-Caf1 deadenylase complex in nuclear cajal bodies. Mol Cell Biol. 2007;27(5):1686–95.CrossRefPubMedGoogle Scholar
  12. 12.
    Zheng D, Ezzeddine N, Chen CY, Zhu W, He X, Shyu AB. Deadenylation is prerequisite for P-body formation and mRNA decay in mammalian cells. J Cell Biol. 2008;182(1):89–101.CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Machyna M, Heyn P, Neugebauer KM. Cajal bodies: where form meets function. Wiley Interdiscip Rev RNA. 2013;4(1):17–34.CrossRefPubMedGoogle Scholar
  14. 14.
    Parker R, Sheth UP. Bodies and the control of mRNA translation and degradation. Mol Cell. 2007;25(5):635–46.CrossRefPubMedGoogle Scholar
  15. 15.
    Cougot N, Bhattacharyya SN, Tapia-Arancibia L, Bordonne R, Filipowicz W, Bertrand E, et al. Dendrites of mammalian neurons contain specialized P-body-like structures that respond to neuronal activation. J Neurosci. 2008;28(51):13793–804.CrossRefPubMedGoogle Scholar
  16. 16.
    Baltanas FC, Casafont I, Weruaga E, Alonso JR, Berciano MT, Lafarga M. Nucleolar disruption and cajal body disassembly are nuclear hallmarks of DNA damage-induced neurodegeneration in purkinje cells. Brain Pathol. 2011;21(4):374–88.CrossRefPubMedGoogle Scholar
  17. 17.
    Oka S, Ohno M, Tsuchimoto D, Sakumi K, Furuichi M, Nakabeppu Y. Two distinct pathways of cell death triggered by oxidative damage to nuclear and mitochondrial DNAs. EMBO J. 2008;27(2):421–32.CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Lee HM, Hu Z, Ma H, Greeley Jr GH, Wang C, Englander EW. Developmental changes in expression and subcellular localization of the DNA repair glycosylase, MYH, in the rat brain. J Neurochem. 2004;88(2):394–400.Google Scholar
  19. 19.
    Sheng Z, Oka S, Tsuchimoto D, Abolhassani N, Nomaru H, Sakumi K, et al. 8-Oxoguanine causes neurodegeneration during MUTYH-mediated DNA base excision repair. J Clin Invest. 2012;122(12):4344–61.CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Plotz G, Casper M, Raedle J, Hinrichsen I, Heckel V, Brieger A, et al. MUTYH gene expression and alternative splicing in controls and polyposis patients. Hum Mutat. 2012;33(7):1067–74.CrossRefPubMedGoogle Scholar
  21. 21.
    Langfelder P, Horvath S. WGCNA: an R package for weighted correlation network analysis. BMC Bioinformatics. 2008;9:559.CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Wu D, Lim E, Vaillant F, Asselin-Labat ML, Visvader JE, Smyth GK. ROAST: rotation gene set tests for complex microarray experiments. Bioinformatics. 2010;26(17):2176–82.CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Okazaki Y, Furuno M, Kasukawa T, Adachi J, Bono H, Kondo S, et al. Analysis of the mouse transcriptome based on functional annotation of 60,770 full-length cDNAs. Nature. 2002;420(6915):563–73.CrossRefPubMedGoogle Scholar
  24. 24.
    Joshi NAJNF. Sickle: A sliding-window, adaptive, quality-based trimming tool for FastQ files 2011 [cited (Version 1.33)]. Software]. Available from: https://github.com/najoshi/sickle.
  25. 25.
    Kim D, Pertea G, Trapnell C, Pimentel H, Kelley R, Salzberg SL. TopHat2: accurate alignment of transcriptomes in the presence of insertions, deletions and gene fusions. Genome Biol. 2013;14(4):R36.CrossRefPubMedPubMedCentralGoogle Scholar
  26. 26.
    Bray N, Pimentel, H., Melsted, P. & Lior, Pachter. Near-optimal RNA-Seq quantification. 2015;arXiv:1505.02710.Google Scholar
  27. 27.
    Trapnell C, Hendrickson DG, Sauvageau M, Goff L, Rinn JL, Pachter L. Differential analysis of gene regulation at transcript resolution with RNA-seq. Nat Biotechnol. 2013;31(1):46–53.CrossRefPubMedGoogle Scholar
  28. 28.
    Robinson MD, McCarthy DJ, Smyth GK. edgeR: a Bioconductor package for differential expression analysis of digital gene expression data. Bioinformatics. 2010;26(1):139–40.CrossRefPubMedGoogle Scholar
  29. 29.
    Team RDC. R: A language and environment for statistical computing. Vienna, Austria: R Foundation for Statistcal Computing; 2010.Google Scholar
  30. 30.
    Kuhn A, Kumar A, Beilina A, Dillman A, Cookson MR, Singleton AB. Cell population-specific expression analysis of human cerebellum. BMC Genomics. 2012:13:610.Google Scholar
  31. 31.
    Kirsch L, Liscovitch N, Chechik G. Localizing genes to cerebellar layers by classifying ISH images. PLoS Comput Biol. 2012;8(12):e1002790.CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Bettencourt C, Ryten M, Forabosco P, Schorge S, Hersheson J, Hardy J, et al. Insights from cerebellar transcriptomic analysis into the pathogenesis of ataxia. JAMA Neurol. 2014;71(7):831–9.CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Mi H, Poudel S, Muruganujan A, Casagrande JT, Thomas PD. PANTHER version 10: expanded protein families and functions, and analysis tools. Nucleic Acids Res. 2016;44(D1):D336–42.CrossRefPubMedGoogle Scholar
  34. 34.
    Mi H, Muruganujan A, Casagrande JT, Thomas PD. Large-scale gene function analysis with the PANTHER classification system. Nat Protoc. 2013;8(8):1551–66.CrossRefPubMedGoogle Scholar
  35. 35.
    D'Souza CA, Chopra V, Varhol R, Xie YY, Bohacec S, Zhao Y, et al. Identification of a set of genes showing regionally enriched expression in the mouse brain. BMC Neurosci. 2008;9:66.CrossRefPubMedPubMedCentralGoogle Scholar
  36. 36.
    Shen EH, Overly CC, Jones AR. The Allen human brain atlas: comprehensive gene expression mapping of the human brain. Trends Neurosci. 2012;35(12):711–4.CrossRefPubMedGoogle Scholar
  37. 37.
    Biolatti C, Gianella P, Capucchio MT, Borrelli A, D'Angelo A. Late onset and rapid progression of cerebellar abiotrophy in a domestic shorthair cat. J Small Anim Pract. 2010;51(2):123–6.CrossRefPubMedGoogle Scholar
  38. 38.
    Forman OP, De Risio L, Matiasek K, Platt S, Mellersh C. Spinocerebellar ataxia in the Italian Spinone dog is associated with an intronic GAA repeat expansion in ITPR1. Mamm Genome. 2015;26(1–2):108–17.CrossRefPubMedGoogle Scholar
  39. 39.
    Sato J, Sasaki S, Yamada N, Tsuchitani M. Hereditary cerebellar degenerative disease (cerebellar cortical abiotrophy) in rabbits. Vet Pathol. 2012;49(4):621–8.CrossRefPubMedGoogle Scholar
  40. 40.
    Whittington RJ, Morton AG, Kennedy DJ. Cerebellar abiotrophy in crossbred cattle. Aust Vet J. 1989;66(1):12–5.CrossRefPubMedGoogle Scholar
  41. 41.
    Forabosco P, Ramasamy A, Trabzuni D, Walker R, Smith C, Bras J, et al. Insights into TREM2 biology by network analysis of human brain gene expression data. Neurobiol Aging. 2013;34(12):2699–714.CrossRefPubMedPubMedCentralGoogle Scholar
  42. 42.
    Block ML, Hong JS. Microglia and inflammation-mediated neurodegeneration: multiple triggers with a common mechanism. Prog Neurobiol. 2005;76(2):77–98.CrossRefPubMedGoogle Scholar
  43. 43.
    Cvetanovic M, Ingram M, Orr H, Opal P. Early activation of microglia and astrocytes in mouse models of spinocerebellar ataxia type 1. Neuroscience. 2015;289:289–99.CrossRefPubMedPubMedCentralGoogle Scholar
  44. 44.
    Guillot-Sestier MV, Doty KR, Gate D, Rodriguez Jr J, Leung BP, Rezai-Zadeh K, et al. Il10 deficiency rebalances innate immunity to mitigate Alzheimer-like pathology. Neuron. 2015;85(3):534–48.Google Scholar
  45. 45.
    LE F, Tirolo C, Testa N, Caniglia S, Morale MC, Marchetti B. Glia as a turning point in the therapeutic strategy of Parkinson's disease. CNS Neurol Disord Drug Targets. 2010;9(3):349–72.CrossRefGoogle Scholar
  46. 46.
    Sultan M, Amstislavskiy V, Risch T, Schuette M, Dokel S, Ralser M, et al. Influence of RNA extraction methods and library selection schemes on RNA-seq data. BMC Genomics. 2014;15:675.CrossRefPubMedPubMedCentralGoogle Scholar
  47. 47.
    Papadimitriou D, Le Verche V, Jacquier A, Ikiz B, Przedborski S, Re DB. Inflammation in ALS and SMA: sorting out the good from the evil. Neurobiol Dis. 2010;37(3):493–502.CrossRefPubMedGoogle Scholar
  48. 48.
    Kaya N, Aldhalaan H, Al-Younes B, Colak D, Shuaib T, Al-Mohaileb F, et al. Phenotypical spectrum of cerebellar ataxia associated with a novel mutation in the CA8 gene, encoding carbonic anhydrase (CA) VIII. Am J Med Genet B Neuropsychiatr Genet. 2011;156B(7):826–34.CrossRefPubMedGoogle Scholar
  49. 49.
    Hirota J, Ando H, Hamada K, Mikoshiba K. Carbonic anhydrase-related protein is a novel binding protein for inositol 1,4,5-trisphosphate receptor type 1. Biochem J. 2003;372(Pt 2):435–41.CrossRefPubMedPubMedCentralGoogle Scholar
  50. 50.
    Okubo Y, Suzuki J, Kanemaru K, Nakamura N, Shibata T, Iino M. Visualization of Ca2+ filling mechanisms upon synaptic inputs in the endoplasmic reticulum of cerebellar Purkinje cells. J Neurosci. 2015;35(48):15837–46.CrossRefPubMedGoogle Scholar
  51. 51.
    Paxinos G. Cerebellum and Cerebellar Connections. In: Science E, editor. The Rat Nervous System. 4th ed. Burlington: Elsevier Science; 2014. p. 1053.Google Scholar
  52. 52.
    Anderson WA, Flumerfelt BA. Long-term effects of parallel fiber loss in the cerebellar cortex of the adult and weanling rat. Brain Res. 1986;383(1–2):245–61.CrossRefPubMedGoogle Scholar
  53. 53.
    Neuman T, Keen A, Zuber MX, Kristjansson GI, Gruss P, Nornes HO. Neuronal expression of regulatory helix-loop-helix factor Id2 gene in mouse. Dev Biol. 1993;160(1):186–95.CrossRefPubMedGoogle Scholar
  54. 54.
    Sullivan JM, Havrda MC, Kettenbach AN, Paolella BR, Zhang Z, Gerber SA, et al. Phosphorylation regulates Id2 degradation and mediates the proliferation of neural precursor cells. Stem Cells. 2016;34(5):1321–31.CrossRefPubMedPubMedCentralGoogle Scholar
  55. 55.
    Cho DH, Hong YM, Lee HJ, Woo HN, Pyo JO, Mak TW, et al. Induced inhibition of ischemic/hypoxic injury by APIP, a novel Apaf-1-interacting protein. J Biol Chem. 2004;279(38):39942–50.CrossRefPubMedGoogle Scholar
  56. 56.
    Ko DC, Gamazon ER, Shukla KP, Pfuetzner RA, Whittington D, Holden TD, et al. Functional genetic screen of human diversity reveals that a methionine salvage enzyme regulates inflammatory cell death. Proc Natl Acad Sci U S A. 2012;109(35):E2343–52.CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  1. 1.Department of Animal ScienceUniversity of CaliforniaDavisUSA
  2. 2.Veterinary Genetics Laboratory, School of Veterinary MedicineUniversity of CaliforniaDavisUSA
  3. 3.Department of Population Health and Reproduction, School of Veterinary MedicineUniversity of CaliforniaDavisUSA

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