, Volume 9, Issue 2, pp 83–94 | Cite as

RNA interference of LRRK2–microarray expression analysis of a Parkinson’s disease key player

  • K. Häbig
  • M. Walter
  • S. Poths
  • O. Riess
  • M. Bonin
Original Article


The protein leucine-rich repeat kinase 2 (LRRK2) is a key player in the pathogenesis of Parkinson’s disease (PD). Mutations in the LRRK2 gene account for up to 10% of all autosomal dominant forms of familiar and for approximately 1–3% of sporadic PD patients. Although the LRRK2 protein has many functional domains like a leucine-rich repeat domain, a Roc-GTPase domain, a kinase domain of the tyrosine kinase-like subfamily and multiple protein interaction domains (armadillo, ankyrin, WD40), the exact biological role of LRRK2 in the human brain is elusive. To gain more insight into the biological function of this protein, we monitored the changes in the expression profiles of SH-SY5Y cells, a dopaminergic neuroblastoma cell line, induced by a depletion of LRRK2 levels by RNA interference (RNAi) with Affymetrix U133 Plus 2.0 microarrays. A total of 187 genes were differentially regulated by at least a 1.5-fold change with 94 transcripts being upregulated and 93 transcripts being downregulated compared to scrambled control siRNA transfected cells. Key players of the interaction networks were independently verified by qRT-PCR. The differentially expressed gene products are involved in axonal guidance, nervous system development, cell cycle, cell growth, cell differentiation, cell communication, MAPKKK cascade, and Ras protein signal transduction. Defined gene expression networks will now serve to look more closely for candidates affected by LRRK2 reduction and how they might be altered in other forms of familial or sporadic PD.


LRRK2 Microarray analysis Parkinson’s disease PARK8 RNA interference 



We thank S. Biskup and P. Kahle for helpful discussion and reading the manuscript. This work was supported by IZKF (01KS 9602/4)/BMBF and an NGFN2 grant to OR. Experiments comply with the current laws in Germany.

Supplementary material

10048_2007_114_MOESM1_ESM.doc (476 kb)
Supplementary material 1 Differentially expressed transcripts of the LRRK2 RNAi microarray experiment. (DOC 487 KB)
10048_2007_114_MOESM2_ESM.doc (234 kb)
Supplementary material 2 Differentially expressed transcripts of the 16 presented GO-categories. (DOC 239 KB)


  1. 1.
    Lang AE, Lozano AM (1998) Parkinson's disease. First of two parts. N Engl J Med 339:1044–10053Google Scholar
  2. 2.
    Taylor JP, Mata IF, Farrer MJ (2006) LRRK2: a common pathway for parkinsonism, pathogenesis and prevention. Trends Mol Med 12:76–82PubMedCrossRefGoogle Scholar
  3. 3.
    Zimprich A et al (2004) Mutations in LRRK2 cause autosomal-dominant parkinsonism with pleomorphic pathology. Neuron 44:601–607PubMedCrossRefGoogle Scholar
  4. 4.
    Paisan-Ruiz C et al (2004) Cloning of the gene containing mutations that cause PARK8-linked Parkinson's disease. Neuron 44:595–600PubMedCrossRefGoogle Scholar
  5. 5.
    Hardy J et al (2006) Genetics of Parkinson's disease and parkinsonism. Ann Neurol 60:389–398PubMedCrossRefGoogle Scholar
  6. 6.
    Lesage S et al (2005) G2019S LRRK2 mutation in French and North African families with Parkinson's disease. Ann Neurol 58:784–787PubMedCrossRefGoogle Scholar
  7. 7.
    Gasser T (2005) Genetics of Parkinson's disease. Curr Opin Neurol 18:363–369PubMedCrossRefGoogle Scholar
  8. 8.
    Bonifati V (2005) Genetics of Parkinson's disease. Minerva Med 96:175–186PubMedGoogle Scholar
  9. 9.
    Di Fonzo A et al (2005) A frequent LRRK2 gene mutation associated with autosomal dominant Parkinson's disease. Lancet 365:412–415PubMedGoogle Scholar
  10. 10.
    Gilks WP et al (2005) A common LRRK2 mutation in idiopathic Parkinson's disease. Lancet 365:415–416PubMedGoogle Scholar
  11. 11.
    Kachergus J et al (2005) Identification of a novel LRRK2 mutation linked to autosomal dominant parkinsonism: evidence of a common founder across European populations. Am J Hum Genet 76:672–680PubMedCrossRefGoogle Scholar
  12. 12.
    Ozelius LJ et al (2006) LRRK2 G2019S as a cause of Parkinson's disease in Ashkenazi Jews. N Engl J Med 354:424–425PubMedCrossRefGoogle Scholar
  13. 13.
    Mata IF et al (2006) LRRK2 in Parkinson's disease: protein domains and functional insights. Trends Neurosci 29:286–293PubMedCrossRefGoogle Scholar
  14. 14.
    Kobe B, Deisenhofer J (1994) The leucine-rich repeat: a versatile binding motif. Trends Biochem Sci 19:415–421PubMedCrossRefGoogle Scholar
  15. 15.
    Stenmark H, Olkkonen VM (2001) The Rab GTPase family. Genome Biol 2:REVIEWS3007PubMedCrossRefGoogle Scholar
  16. 16.
    Marin I (2006) The Parkinson disease gene LRRK2: evolutionary and structural insights. Mol Biol Evol 23:2423–2433PubMedCrossRefGoogle Scholar
  17. 17.
    Li D, Roberts R (2001) WD-repeat proteins: structure characteristics, biological function, and their involvement in human diseases. Cell Mol Life Sci 58:2085–2097PubMedCrossRefGoogle Scholar
  18. 18.
    Dykxhoorn DM, Novina CD, Sharp PA (2003) Killing the messenger: short RNAs that silence gene expression. Nat Rev Mol Cell Biol 4:457–467PubMedCrossRefGoogle Scholar
  19. 19.
    Goossens K et al (2005) Selection of reference genes for quantitative real-time PCR in bovine preimplantation embryos. BMC Dev Biol 5:27PubMedCrossRefGoogle Scholar
  20. 20.
    Hellemans J et al (2007) qBase relative quantification framework and software for management and automated analysis of real-time quantitative PCR data. Genome Biol 8:R19PubMedCrossRefGoogle Scholar
  21. 21.
    Pfaffl MW (2001) A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res 29:e45PubMedCrossRefGoogle Scholar
  22. 22.
    Pfaffl MW, Horgan GW, Dempfle L (2002) Relative expression software tool (REST) for group-wise comparison and statistical analysis of relative expression results in real-time PCR. Nucleic Acids Res 30:e36PubMedCrossRefGoogle Scholar
  23. 23.
    Farrer M et al (2005) LRRK2 mutations in Parkinson disease. Neurology 65:738–740PubMedCrossRefGoogle Scholar
  24. 24.
    Holen T et al (2002) Positional effects of short interfering RNAs targeting the human coagulation trigger Tissue Factor. Nucleic Acids Res 30:1757–1766PubMedCrossRefGoogle Scholar
  25. 25.
    Luu-The V et al (2005) Improved real-time RT-PCR method for high-throughput measurements using second derivative calculation and double correction. Biotechniques 38:287–293PubMedCrossRefGoogle Scholar
  26. 26.
    Erickson JW et al (1996) Mammalian Cdc42 is a brefeldin A-sensitive component of the Golgi apparatus. J Biol Chem 271:26850–26854PubMedCrossRefGoogle Scholar
  27. 27.
    Yang L, Wang L, Zheng Y (2006) Gene targeting of Cdc42 and Cdc42GAP affirms the critical involvement of Cdc42 in filopodia induction, directed migration, and proliferation in primary mouse embryonic fibroblasts. Mol Biol Cell 17:4675–4685PubMedCrossRefGoogle Scholar
  28. 28.
    Meyer G, Feldman EL (2002) Signaling mechanisms that regulate actin-based motility processes in the nervous system. J Neurochem 83:490–503PubMedCrossRefGoogle Scholar
  29. 29.
    MacLeod D et al (2006) The familial Parkinsonism gene LRRK2 regulates neurite process morphology. Neuron 52:587–593PubMedCrossRefGoogle Scholar
  30. 30.
    West AB et al (2007) Parkinson's disease-associated mutations in LRRK2 link enhanced GTP-binding and kinase activities to neuronal toxicity. Hum Mol Genet 16:223–232PubMedCrossRefGoogle Scholar
  31. 31.
    Li BS et al (2001) Activation of phosphatidylinositol-3 kinase (PI-3K) and extracellular regulated kinases (Erk1/2) is involved in muscarinic receptor-mediated DNA synthesis in neural progenitor cells. J Neurosci 21:1569–1579PubMedGoogle Scholar
  32. 32.
    Enslen H et al (1996) Regulation of mitogen-activated protein kinases by a calcium/calmodulin-dependent protein kinase cascade. Proc Natl Acad Sci USA 93:10803–10808PubMedCrossRefGoogle Scholar
  33. 33.
    Yang JJ (2002) Mixed lineage kinase ZAK utilizing MKK7 and not MKK4 to activate the c-Jun N-terminal kinase and playing a role in the cell arrest. Biochem Biophys Res Commun 297:105–110PubMedCrossRefGoogle Scholar
  34. 34.
    Di Cunto F et al (1998) Citron rho-interacting kinase, a novel tissue-specific ser/thr kinase encompassing the Rho-Rac-binding protein Citron. J Biol Chem 273:29706–29711PubMedCrossRefGoogle Scholar
  35. 35.
    Mogi M et al (2007) p53 protein, interferon-gamma, and NF-kappaB levels are elevated in the parkinsonian brain. Neurosci Lett 414:94–97PubMedCrossRefGoogle Scholar
  36. 36.
    Nair VD et al (2006) p53 mediates nontranscriptional cell death in dopaminergic cells in response to proteasome inhibition. J Biol Chem 281:39550–39560PubMedCrossRefGoogle Scholar
  37. 37.
    Rinaldo C et al (2007) MDM2-regulated degradation of HIPK2 prevents p53Ser46 phosphorylation and DNA damage-induced apoptosis. Mol Cell 25:739–750PubMedCrossRefGoogle Scholar
  38. 38.
    Dachsel JC et al (2007) Identification of potential protein interactors of Lrrk2. Parkinsonism Relat Disord 13(7):382–385PubMedCrossRefGoogle Scholar
  39. 39.
    Kaul SC et al (2005) Activation of wild type p53 function by its mortalin-binding, cytoplasmically localizing carboxyl terminus peptides. J Biol Chem 280:39373–39379PubMedCrossRefGoogle Scholar
  40. 40.
    Nihei T et al (1993) Demonstration of selective protein complexes of p53 with 73 kDa heat shock cognate protein, but not with 72 kDa heat shock protein in human tumor cells. Cancer Lett 73:181–189PubMedCrossRefGoogle Scholar
  41. 41.
    Zhang Y et al (2004) Repression of hsp90beta gene by p53 in UV irradiation-induced apoptosis of Jurkat cells. J Biol Chem 279:42545–42551PubMedCrossRefGoogle Scholar
  42. 42.
    Ceballos E et al (2005) Inhibitory effect of c-Myc on p53-induced apoptosis in leukemia cells. Microarray analysis reveals defective induction of p53 target genes and upregulation of chaperone genes. Oncogene 24:4559–45571PubMedCrossRefGoogle Scholar
  43. 43.
    Galy B et al (2001) p53 directs conformational change and translation initiation blockade of human fibroblast growth factor 2 mRNA. Oncogene 20:4613–4620PubMedCrossRefGoogle Scholar
  44. 44.
    Romanov VV et al (2005) Basic fibroblast growth factor suppresses p53 activation in the neoplastic cells of a proportion of patients with chronic lymphocytic leukaemia. Oncogene 24:6855–6860PubMedCrossRefGoogle Scholar
  45. 45.
    Lee HT, Kay EP (2003) FGF-2 induced reorganization and disruption of actin cytoskeleton through PI 3-kinase, Rho, and Cdc42 in corneal endothelial cells. Mol Vis 9:624–634PubMedGoogle Scholar
  46. 46.
    Dell'Era P et al (2002) Gene expression profile in fibroblast growth factor 2-transformed endothelial cells. Oncogene 21:2433–2440PubMedCrossRefGoogle Scholar
  47. 47.
    Thomas EC et al (2006) The subcellular fractionation properties and function of insulin receptor substrate-1 (IRS-1) are independent of cytoskeletal integrity. Int J Biochem Cell Biol 38:1686–1699PubMedCrossRefGoogle Scholar
  48. 48.
    Zhang CC et al (1998) The role of MAP4 expression in the sensitivity to paclitaxel and resistance to vinca alkaloids in p53 mutant cells. Oncogene 16:1617–1624PubMedCrossRefGoogle Scholar
  49. 49.
    Marino G et al (2003) Human autophagins, a family of cysteine proteinases potentially implicated in cell degradation by autophagy. J Biol Chem 278:3671–3678PubMedCrossRefGoogle Scholar
  50. 50.
    Marszalek JR et al (1999) Novel dendritic kinesin sorting identified by different process targeting of two related kinesins: KIF21A and KIF21B. J Cell Biol 145:469–479PubMedCrossRefGoogle Scholar
  51. 51.
    Jaleel M et al (2007) LRRK2 phosphorylates moesin at threonine-558: characterization of how Parkinson's disease mutants affect kinase activity. Biochem J 405:307–317PubMedCrossRefGoogle Scholar
  52. 52.
    Muthuchamy M et al (1998) Beta-tropomyosin overexpression induces severe cardiac abnormalities. J Mol Cell Cardiol 30:1545–1557PubMedCrossRefGoogle Scholar
  53. 53.
    Sultana S et al (1998) Effects of growth factors and basement membrane proteins on the phenotype of U-373 MG glioblastoma cells as determined by the expression of intermediate filament proteins. Am J Pathol 153:1157–1168PubMedGoogle Scholar
  54. 54.
    Izumi M et al (2001) Bone morphogenetic protein-2 inhibits serum deprivation-induced apoptosis of neonatal cardiac myocytes through activation of the Smad1 pathway. J Biol Chem 276:31133–31141PubMedCrossRefGoogle Scholar
  55. 55.
    Hammond PW et al (2001) In vitro selection and characterization of Bcl-X(L)-binding proteins from a mix of tissue-specific mRNA display libraries. J Biol Chem 276:20898–20906PubMedCrossRefGoogle Scholar
  56. 56.
    Shorter J et al (2002) Sequential tethering of Golgins and catalysis of SNAREpin assembly by the vesicle-tethering protein p115. J Cell Biol 157:45–62PubMedCrossRefGoogle Scholar
  57. 57.
    Chu CY, Rana TM (2006) Translation repression in human cells by microRNA-induced gene silencing requires RCK/p54. PLoS Biol 4:e210PubMedCrossRefGoogle Scholar
  58. 58.
    Nathan CA et al (1997) Elevated expression of eIF4E and FGF-2 isoforms during vascularization of breast carcinomas. Oncogene 15:1087–1094PubMedCrossRefGoogle Scholar
  59. 59.
    Smith WW et al (2005) Leucine-rich repeat kinase 2 (LRRK2) interacts with parkin, and mutant LRRK2 induces neuronal degeneration. Proc Natl Acad Sci USA 102:18676–18681PubMedCrossRefGoogle Scholar
  60. 60.
    Imai Y et al (2003) A product of the human gene adjacent to parkin is a component of Lewy bodies and suppresses Pael receptor-induced cell death. J Biol Chem 278:51901–51910PubMedCrossRefGoogle Scholar
  61. 61.
    Bryce NS et al (2003) Specification of actin filament function and molecular composition by tropomyosin isoforms. Mol Biol Cell 14:1002–10016PubMedCrossRefGoogle Scholar
  62. 62.
    Kishimoto K et al (2005) Endogenous angiogenin in endothelial cells is a general requirement for cell proliferation and angiogenesis. Oncogene 24:445–456PubMedCrossRefGoogle Scholar
  63. 63.
    Sakaguchi-Nakashima A et al (2007) LRK-1, a C. elegans PARK8-related kinase, regulates axonal-dendritic polarity of SV proteins. Curr Biol 17:592–598PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2007

Authors and Affiliations

  • K. Häbig
    • 1
  • M. Walter
    • 1
  • S. Poths
    • 1
  • O. Riess
    • 1
  • M. Bonin
    • 1
  1. 1.Department of Medical Genetics, Microarray FacilityUniversity of TübingenTübingenGermany

Personalised recommendations