Parkin promotes proteasomal degradation of p62: implication of selective vulnerability of neuronal cells in the pathogenesis of Parkinson’s disease

Abstract

Mutations or inactivation of parkin, an E3 ubiquitin ligase, are associated with familial form or sporadic Parkinson’s disease (PD), respectively, which manifested with the selective vulnerability of neuronal cells in substantia nigra (SN) and striatum (STR) regions. However, the underlying molecular mechanism linking parkin with the etiology of PD remains elusive. Here we report that p62, a critical regulator for protein quality control, inclusion body formation, selective autophagy and diverse signaling pathways, is a new substrate of parkin. P62 levels were increased in the SN and STR regions, but not in other brain regions in parkin knockout mice. Parkin directly interacts with and ubiquitinates p62 at the K13 to promote proteasomal degradation of p62 even in the absence of ATG5. Pathogenic mutations, knockdown of parkin or mutation of p62 at K13 prevented the degradation of p62. We further showed that parkin deficiency mice have pronounced loss of tyrosine hydroxylase positive neurons and have worse performance in motor test when treated with 6-hydroxydopamine hydrochloride in aged mice. These results suggest that, in addition to their critical role in regulating autophagy, p62 are subjected to parkin mediated proteasomal degradation and implicate that the dysregulation of parkin/p62 axis may involve in the selective vulnerability of neuronal cells during the onset of PD pathogenesis.

Key words： parkin; sequestosome1/p62; ubiquitin; substantia nigra

References

1 Babu JR, Geetha T, Wooten MW (2005) Sequestosome 1/p62 shuttles polyubiquitinated tau for proteasomal degradation . J Neurochem 94:192–203

2 Bartlett BJ, Isakson P, Lewerenz J, Sanchez H, Kotzebue RW, Cumming RC, Harris GL, Nezis IP, Schubert DR, Simonsen A (2011) p62, Ref(2)P and ubiquitinated proteins are conserved markers of neuronal aging, aggregate formation and progressive autophagic defects . Autophagy 7:572–583

3 Bingol B, Tea JS, Phu L, Reichelt M, Bakalarski CE, Song Q, Foreman O, Kirkpatrick DS, Sheng M (2014) The mitochondrial deubiquitinase USP30 opposes parkin-mediated mitophagy . Nature 510:370–375

4 Biskup S, Gerlach M, Kupsch A, Reichmann H, Riederer P, Vieregge P, Wullner U, Gasser T (2008) Genes associatedwith Parkinson syndrome . J Neurol 255(Suppl 5):8–17

5 Bjorkoy G, Lamark T, Johansen T (2006) p62/SQSTM1: a missing link between protein aggregates and the autophagy machinery . Autophagy 2:138–139

6 Bossy-Wetzel E, Schwarzenbacher R, Lipton SA (2004) Molecular pathways to neurodegeneration . Nat Med 10(Suppl):S2–S9

7 Burchell VS, Nelson DE, Sanchez-Martinez A, Delgado-Camprubi M, Ivatt RM, Pogson JH, Randle SJ, Wray S, Lewis PA, Houlden H (2013) The Parkinson’s disease-linked proteins Fbxo7 and Parkin interact to mediate mitophagy . Nat Neurosci 16:1257–1265

8 Chen Y, Dorn GW 2nd (2013) PINK1-phosphorylated mitofusin2isa Parkin receptor for culling damaged mitochondria . Science 340:471–475

9 Chung KK, Thomas B, Li X, Pletnikova O, Troncoso JC, Marsh L, Dawson VL, Dawson TM (2004) S-nitrosylation of parkin regulates ubiquitination and compromises parkin’s protective function . Science 304:1328–1331

10 Dawson TM (2007) Unraveling the role of defective genes in Parkinson’s disease . Parkinsonism Relat Disord 13(Suppl 3): S248–S249

11 Dawson TM, Dawson VL (2003) Molecular pathways of neurodegeneration in Parkinson’s disease . Science 302:819–822

12 Dawson TM, Dawson VL (2010) The role of parkin in familial and sporadic Parkinson’s disease . Mov Disord 25(Suppl 1):S32–S39

13 Farrer MJ (2006) Genetics of Parkinson disease: paradigm shifts and future prospects . Nat Rev Genet 7:306–318

14 Fecto F, Yan J, Vemula SP, Liu E, Yang Y, Chen W, Zheng JG, Shi Y, Siddique N, Arrat H (2011) SQSTM1 mutations in familial and sporadic amyotrophic lateral sclerosis . Arch Neurol 68:1440–1446

15 Gegg ME, Schapira AH (2011) PINK1-parkin-dependent mitophagy involves ubiquitination of mitofusins 1 and 2: implications for Parkinson disease pathogenesis . Autophagy 7:243–245

16 Gegg ME, Cooper JM, Chau KY, Rojo M, Schapira AH, Taanman JW (2010) Mitofusin 1 and mitofusin 2 are ubiquitinated in a PINK1/parkin-dependent manner upon induction of mitophagy . Hum Mol Genet19:4861–4870

17 Goldberg MS, Fleming SM, Palacino JJ, Cepeda C, Lam HA, Bhatnagar A, Meloni EG, Wu N, Ackerson LC, Klapstein GJ (2003) Parkin-deficient mice exhibit nigrostriatal deficits but not loss of dopaminergic neurons . J Biol Chem 278:43628–43635

18 Ichimura Y, Komatsu M (2010) Selective degradation of p62 by autophagy . Semin Immunopathol 32:431–436

19 Ishikawa A, Tsuji S (1996) Clinical analysis of 17 patients in 12 Japanese families with autosomal-recessive type juvenile parkinsonism . Neurology 47:160–166

20 Itier JM, Ibanez P, Mena MA, Abbas N, Cohen-Salmon C, Bohme GA, Laville M, Pratt J, Corti O, Pradier L (2003) Parkin gene inactivation alters behaviour and dopamine neurotransmission in the mouse . Hum Mol Genet 12:2277–2291

21 Jain A, Lamark T, Sjottem E, Larsen KB, Awuh JA, Overvatn A, McMahon M, Hayes JD, Johansen T (2010) p62/SQSTM1 is a target gene for transcription factor NRF2 and creates a positive feedback loop by inducing antioxidant response element-driven gene transcription . J Biol Chem 285:22576–22591

22 Johnson BN, Berger AK, Cortese GP, Lavoie MJ (2012) The ubiquitin E3 ligase parkin regulates the proapoptotic function of Bax . Proc Natl Acad Sci USA 109:6283–6288

23 Kahle PJ, Haass C (2004) How does parkin ligate ubiquitin to Parkinson’s disease ? EMBO Rep 5:681–685

24 Kane LA, Lazarou M, Fogel AI, Li Y, Yamano K, Sarraf SA, Banerjee S, Youle RJ (2014) PINK1 phosphorylates ubiquitin to activate Parkin E3 ubiquitin ligase activity . J Cell Biol 205:143–153

25 Kawahara K, Hashimoto M, Bar-On P, Ho GJ, Crews L, Mizuno H, Rockenstein E, Imam SZ, Masliah E (2008) alpha-Synuclein aggregates interfere with Parkin solubility and distribution: role in the pathogenesis of Parkinson disease . J Biol Chem 283:6979–6987

26 Kazlauskaite A, Kondapalli C, Gourlay R, Campbell DG, Ritorto MS, Hofmann K, Alessi DR, Knebel A, Trost M, Muqit MM (2014) Parkin is activated by PINK1-dependent phosphorylation of ubiquitin at Ser65 . Biochem J460:127–139

27 Kirkin V, McEwan DG, Novak I, Dikic I (2009) A role for ubiquitin in selective autophagy . Mol Cell 34:259–269

28 Kitada T, Asakawa S, Hattori N, Matsumine H, Yamamura Y, Minoshima S, Yokochi M, Mizuno Y, Shimizu N(1998) Mutations in the parkin gene cause autosomal recessive juvenile parkinsonism . Nature392:605–608

29 Ko HS, Kim SW, Sriram SR, Dawson VL, Dawson TM (2006) Identification of far upstream element-binding protein-1 as an authentic Parkin substrate . J Biol Chem 281:16193–16196

30 Komatsu M, Ichimura Y (2010) Physiological significance of selective degradation of p62 by autophagy . FEBS Lett 584:1374–1378

31 Komatsu M, Waguri S, Koike M, Sou YS, Ueno T, Hara T, Mizushima N, Iwata J, Ezaki J, Murata S (2007) Homeostatic levels of p62 control cytoplasmic inclusion body formation in autophagydeficient mice . Cell 131:1149–1163

32 Koyano F, Okatsu K, Kosako H, Tamura Y, Go E, Kimura M, Kimura Y, Tsuchiya H, Yoshihara H, Hirokawa T (2014) Ubiquitin is phosphorylated by PINK1 to activate parkin . Nature 510:162–166

33 LaVoie MJ, Ostaszewski BL, Weihofen A, Schlossmacher MG, Selkoe DJ (2005) Dopamine covalently modifies and functionally inactivates parkin . Nat Med 11:1214–1221

34 LaVoie MJ, Cortese GP, Ostaszewski BL, Schlossmacher MG (2007) The effects of oxidative stress on parkin and other E3 ligases . J Neurochem 103:2354–2368

35 Lazarou M, Narendra DP, Jin SM, Tekle E, Banerjee S, Youle RJ (2013) PINK1 drives Parkin self-association and HECT-like E3 activity upstreamof mitochondrial binding. J CellBiol 200:163–172

36 Lazarou M, Sliter DA, Kane LA, Sarraf SA, Wang CX, Burman JL, Sideris DP, Fogel AI, Youle RJ (2015) Theubiquitin kinase PINK1recruits autophagy receptorstoinduce mitophagy . Nature 524:309–314

37 Lee JT, Wheeler TC, Li L, Chin LS (2008) Ubiquitination of alphasynuclein by Siah-1 promotes alpha-synuclein aggregation and apoptotic cell death . Hum Mol Genet 17:906–917

38 Lee J, Kim HR, Quinley C, Kim J, Gonzalez-Navajas J, Xavier R, Raz E(2012) Autophagy suppresses interleukin-1beta (IL-1beta) signaling by activation of p62 degradation via lysosomal and proteasomal pathways . J Biol Chem 287:4033–4040

39 Lesage S, Brice A(2009) Parkinson’sdisease: from monogenic forms to genetic susceptibility factors . Hum Mol Genet 18:R48–R59

40 Li H, Guo M (2009) Protein degradation in Parkinson disease revisited: it’s complex . J Clin Investig 119:442–445

41 Li J, Qi W, Chen G, Feng D, Liu JH, Ma B, Zhou CQ, Mu CL, Zhang WL, Chen Q (2015) Mitochondrial outer-membrane E3 ligase MUL1 ubiquitinates ULK1 and regulates selenite-induced mitophagy . Autophagy 11:1216–1229

42 Lowe J, Blanchard A, Morrell K, Lennox G, Reynolds L, Billett M, Landon M, Mayer RJ (1988) Ubiquitin is a common factor in intermediate filament inclusion bodies of diverse type in man, including those of Parkinson’s disease, Pick’s disease, and Alzheimer’s disease, as well as Rosenthal fibres in cerebellar astrocytomas, cytoplasmic bodies in muscle, and mallory bodies in alcoholic liver disease . J Pathol 155:9–15

43 Matsuda N, Sato S, Shiba K, Okatsu K, Saisho K, Gautier CA, Sou YS, Saiki S, Kawajiri S, Sato F (2010) PINK1 stabilized by mitochondrial depolarization recruits Parkin to damaged mitochondria and activateslatent Parkinformitophagy . J CellBiol 189:211–221

44 Nakaso K, Yoshimoto Y, Nakano T, Takeshima T, Fukuhara Y, Yasui K, Araga S, Yanagawa T, Ishii T, Nakashima K (2004) Transcriptional activation of p62/A170/ZIP during the formation of the aggregates: possible mechanisms and the role in Lewy body formation in Parkinson’s disease . Brain Res1012–42–51

45 Narendra D, Tanaka A, Suen DF, Youle RJ (2008) Parkinis recruited selectively to impaired mitochondria and promotes their autophagy . J Cell Biol 183:795–803

46 Pankiv S, Clausen TH, Lamark T, Brech A, Bruun JA, Outzen H, Overvatn A, Bjorkoy G, Johansen T (2007) p62/SQSTM1 binds directly to Atg8/LC3 to facilitate degradation of ubiquitinated protein aggregatesby autophagy . J Biol Chem 282:24131–24145

47 Pankiv S, Lamark T, Bruun JA, Overvatn A, Bjorkoy G, Johansen T (2010) Nucleocytoplasmic shuttling of p62/SQSTM1 and its role in recruitment of nuclear polyubiquitinated proteins to promyelocytic leukemia bodies . J Biol Chem 285:5941–5953

48 Perez FA, Palmiter RD (2005) Parkin-deficient mice are not a robust model of parkinsonism . Proc Natl Acad Sci USA 102:2174–2179

49 Perez FA, Curtis WR, Palmiter RD (2005) Parkin-deficient mice are not more sensitive to 6-hydroxydopamine or methamphetamine neurotoxicity . BMC neuroscience 6:71

50 Pickrell AM, Youle RJ (2015) The roles of PINK1, Parkin, and mitochondrial fidelity in Parkinson’s disease . Neuron 85:257–273

51 Rott R, Szargel R, Haskin J, Shani V, Shainskaya A, Manov I, Liani E, Avraham E, Engelender S(2008) Monoubiquitylationof alphasynuclein by seven in absentia homolog (SIAH) promotes its aggregation in dopaminergic cells . J Biol Chem 283:3316–3328

52 Rubino E, Rainero I, Chio A, Rogaeva E, Galimberti D, Fenoglio P, Grinberg Y, Isaia G, Calvo A, Gentile S (2012) SQSTM1 mutations in frontotemporal lobar degeneration and amyotrophic lateral sclerosis . Neurology 79:1556–1562

53 Rue L, Lopez-Soop G, Gelpi E, Martinez-Vicente M, Alberch J, Perez-Navarro E (2013) Brain region-and age-dependent dysregulation of p62 and NBR1 in a mouse model of Huntington’s disease . Neurobiol Dis 52:219–228

54 Sarraf SA, Raman M, Guarani-Pereira V, Sowa ME, Huttlin EL, Gygi SP, Harper JW (2013) Landscape of the PARKIN-dependent ubiquitylome in response to mitochondrial depolarization . Nature496 : 372–376

55 Seibenhener ML, Babu JR, Geetha T, Wong HC, Krishna NR, Wooten MW (2004) Sequestosome 1/p62 is a polyubiquitin chain binding protein involved in ubiquitin proteasome degradation . Mol Cell Biol 24:8055–8068

56 Shin JH, Ko HS, Kang H, Lee Y, Lee YI, Pletinkova O, Troconso JC, Dawson VL, Dawson TM (2011) PARIS (ZNF746) repression of PGC-1alpha contributes to neurodegeneration in Parkinson’s disease . Cell 144:689–702

57 Singleton AB, Farrer M, Johnson J, Singleton A, Hague S, Kachergus J, Hulihan M, Peuralinna T, Dutra A, Nussbaum R (2003) alpha-Synuclein locus triplication causes Parkinson’s disease. Science 302:841

58 Spillantini MG, Schmidt ML, Lee VM, Trojanowski JQ, Jakes R, Goedert M (1997) Alpha-synuclein in Lewy bodies . Nature 388:839–840

59 Sriram SR, Li X, Ko HS, Chung KK, Wong E, Lim KL, Dawson VL, Dawson TM (2005) Familial-associated mutations differentially disrupt the solubility, localization, binding and ubiquitination properties of parkin . Hum Mol Genet 14:2571–2586

60 Sterky FH, Lee S, Wibom R, Olson L, Larsson NG (2011) Impaired mitochondrial transport and Parkin-independent degeneration of respiratory chain-deficient dopamine neurons in vivo . Proc Natl Acad Sci USA 108:12937–12942

61 Tanaka A, Cleland MM, Xu S, Narendra DP, Suen DF, Karbowski M, Youle RJ (2010) Proteasome and p97 mediate mitophagy and degradation of mitofusins induced by Parkin . J Cell Biol 191:1367–1380

62 Thomas, B., and Beal, M.F. (2007). Parkinson’s disease . Human molecular genetics 16 Spec No. 2, R183–194

63 Van Laar VS, Arnold B, Cassady SJ, Chu CT, Burton EA, Berman SB (2011) Bioenergetics of neurons inhibit the translocation response of Parkin following rapid mitochondrial depolarization . Hum Mol Genet 20:927–940

64 Wang H, Song P, Du L, Tian W, Yue W, Liu M, Li D, Wang B, Zhu Y, Cao C (2011a) Parkin ubiquitinates Drp1 for proteasomedependent degradation:implication of dysregulated mitochondrial dynamics in Parkinson disease . J Biol Chem 286:11649–11658

65 Wang XN, Winter D, Ashrafi G, Schlehe J, Wong YL, Selkoe D, Rice S, Steen J, LaVoie MJ, Schwarz TL (2011b) PINK1 and Parkin target miro for phosphorylation and degradation to arrest mitochondrial motility . Cell 147:893–906

66 Wooten MW, Geetha T, Babu JR, Seibenhener ML, Peng J, Cox N, Diaz-Meco MT, Moscat J (2008) Essential role of sequestosome 1/p62 in regulating accumulation ofLys63-ubiquitinated proteins . J Biol Chem 283:6783–6789

67 Zatloukal K, Stumptner C, Fuchsbichler A, Heid H, Schnoelzer M, Kenner L, Kleinert R, Prinz M, Aguzzi A, Denk H(2002) p62Isa common component of cytoplasmic inclusions in protein aggregation diseases . Am J Pathol 160:255–263