Nearly 10 years ago, the kinase PINK1 and the E3 ubiquitin ligase Parkin, two proteins associated with autosomal-recessive Parkinson’s disease (PD), were identified as key players in the selective degradation of damaged mitochondria (mitophagy). Since then, we have been presented with a central role for both with stabilization of active PINK1 at the mitochondria recruiting Parkin, Parkin redecorating the outer mitochondrial membrane with ubiquitin, ubiquitin recruiting the autophagy and the proteasome machinery, and ultimately inducing mitophagy.
The identification of PINK1 as the first ubiquitin kinase (Kane et al., 2014; Kazlauskaite et al., 2014; Koyano et al., 2014) has considerably improved our understanding of the mitophagy pathway. Now, this thorough study by Richard Youle’s group emphasizes how indispensable PINK1 is because by generating phospho-ubiquitin, the essential ligand for the two primary autophagy receptors (OPTN and NDP52), it recruits the upstream autophagy machinery to mitochondria, and ultimately induces mitophagy.
Parkin, on the other hand, seems to amplify the mitophagy signal by generating more ubiquitin substrate for PINK1 to phosphorylate, as opposed to being indispensable for mitophagy. Whether Parkin is indispensable for mitophagy in HeLa cells or other E3 ubiquitin ligases can play the same role remains to be determined.
Not only does this study clarify the respective roles of PINK1 and Parkin in mitophagy (and show a new PINK1-dependant/Parkin-independent model of autophagy) but it also highlights the probable important role of mitophagy in the pathogenesis of neurodegenerative conditions other than PD. OPTN and DDF52 are associated with primary angle glaucoma, amyotrophic lateral sclerosis, and Crohn’s disease. Why mutations in PINK1, Parkin, OPTN, NDF52, but also other genes such as VCP and MFN2 can all interfere with the mitophagy pathway while leading to different neurodegenerative conditions may be key in understanding selective cellular vulnerability.
References:
Kane LA, Lazarou M, Fogel AI, Li Y, Yamano K, Sarraf SA, Banerjee S, Youle RJ.
PINK1 phosphorylates ubiquitin to activate Parkin E3 ubiquitin ligase activity.
J Cell Biol. 2014 Apr 28;205(2):143-53. Epub 2014 Apr 21
PubMed.
Kazlauskaite A, Kondapalli C, Gourlay R, Campbell DG, Ritorto MS, Hofmann K, Alessi DR, Knebel A, Trost M, Muqit MM.
Parkin is activated by PINK1-dependent phosphorylation of ubiquitin at Ser65.
Biochem J. 2014 May 15;460(1):127-39.
PubMed.
Koyano F, Okatsu K, Kosako H, Tamura Y, Go E, Kimura M, Kimura Y, Tsuchiya H, Yoshihara H, Hirokawa T, Endo T, Fon EA, Trempe JF, Saeki Y, Tanaka K, Matsuda N.
Ubiquitin is phosphorylated by PINK1 to activate parkin.
Nature. 2014 Jun 5;510(7503):162-6. Epub 2014 Jun 4
PubMed.
Commentary: This paper presents an exciting surprise to Parkin/PINK1 and mitophagy community in the form of another important discovery from Richard Youle’s laboratory. The Youle group (among others) had previously discovered the PINK1-dependent Parkin activation and recruitment to damaged mitochondria and the involvement of ubiquitin phosphorylated by PINK1 (pUb) in the process (Narendra et al. 2010; Koyano et al. 2014; Kane et al. 2014; Kazlauskaite et al 2014). These discoveries helped uncover the molecular mechanism of Parkin’s activation (Sauvé et al. 2015; Wauer et al. 2015) from its basal autoinhibited state (Trempe et al. 2013). Here, they show that, while Parkin might be required for efficient mitophagy, the process can still occur independently of Parkin, but relies critically on PINK1’s ubiquitin kinase activity.
The general questions that this study aims to answer are: 1) which autophagy receptors are recruited to mitochondria and are critical for PINK1-dependent mitophagy and 2) does pUb generated by PINK1 have roles in mitophagy besides the allosteric activation of Parkin?
The authors generated gene knockouts of five autophagy cargo receptors to impair mitophagy and show that the reintroduction of only NDP52 or Optineurin (OPTN) rescued the impairment. This rescue occurs even in the absence of Parkin. NDP52 and OPTN (and its phosphorylation by TBK1) have been investigated previously for their roles for xenophagy (Thurston et al. 2009; Wilds et al. 2011) but their involvement in mitophagy is novel. The absence of Parkin helped the authors discriminate better the autophagy receptors that are critical and those that are dispensable. Previous studies (Geisler et al. 2010) had suggested the recruitment of p62 to mitochondria following Parkin activation, but here it is shown not to be essential for mitophagy. The new study demonstrates the value of COXII and mtDNA depletion as markers for mitophagy as MFN1 (a common marker for Parkin’s activity and mitophagy) and TOM20 can be targeted to the proteasome directly by Parkin-mediated ubiquitination in the absence of autophagy.
Youle’s group also discovers another role for pUb: the recruitment of NDP52 and OPTN. They demonstrate that NDP52 and OPTN are preferentially pulled down by phosphomimetic ubiquitin from HeLa cells. The cargo receptor p62 doesn’t have this property. This part of the manuscript will require some additional proof. For example, some experiments were done using phosphomimetic ubiquitin, which doesn’t capture all the functional characteristics of pUb (Ordureau et al. 2015). Also, the use of a Parkin overexpression system in the latter experiments puts the assertion about the Parkin-independence of the process on a difficult footing. Nonetheless, the observations are very significant. Future studies including a paper this week from the Harper group (Heo et al. 2015) will clarify the molecular basis of NDP52 and OPTN recruitment.
Finally, it has been shown that in the absence of NDP52 and OPTN or PINK1 both upstream (ULK1 complex) and downstream components (LC3B) of the autophagy machinery are not effectively recruited, explaining the impairment in mitophagy. The study also shows that mutations in NDP52 and OPTN associated with various diseases including amyotrophic lateral sclerosis (ALS or Lou Gehrig's disease) and Crohn’s disease are impaired in mitophagy and hence indicates that impaired mitophagy might have a contribution towards the observed pathologies. This new study adds significantly to our understanding of mitophagy by assigning roles to different components of the mitochondrial quality control pathway, both upstream and downstream in the process. The findings should also encourage the investigation and development of therapeutics targeting PINK1 activation for the treatment of Parkinson disease.
References:
Geisler S, Holmström KM, Skujat D, Fiesel FC, Rothfuss OC, Kahle PJ, Springer W.
PINK1/Parkin-mediated mitophagy is dependent on VDAC1 and p62/SQSTM1.
Nat Cell Biol. 2010 Feb;12(2):119-31. Epub 2010 Jan 24
PubMed.
Heo JM, Ordureau A, Paulo JA, Rinehart J, Harper JW.
The PINK1-PARKIN Mitochondrial Ubiquitylation Pathway Drives a Program of OPTN/NDP52 Recruitment and TBK1 Activation to Promote Mitophagy.
Mol Cell. 2015 Oct 1;60(1):7-20. Epub 2015 Sep 10
PubMed.
Kane LA, Lazarou M, Fogel AI, Li Y, Yamano K, Sarraf SA, Banerjee S, Youle RJ.
PINK1 phosphorylates ubiquitin to activate Parkin E3 ubiquitin ligase activity.
J Cell Biol. 2014 Apr 28;205(2):143-53. Epub 2014 Apr 21
PubMed.
Kazlauskaite A, Kondapalli C, Gourlay R, Campbell DG, Ritorto MS, Hofmann K, Alessi DR, Knebel A, Trost M, Muqit MM.
Parkin is activated by PINK1-dependent phosphorylation of ubiquitin at Ser65.
Biochem J. 2014 May 15;460(1):127-39.
PubMed.
Koyano F, Okatsu K, Kosako H, Tamura Y, Go E, Kimura M, Kimura Y, Tsuchiya H, Yoshihara H, Hirokawa T, Endo T, Fon EA, Trempe JF, Saeki Y, Tanaka K, Matsuda N.
Ubiquitin is phosphorylated by PINK1 to activate parkin.
Nature. 2014 Jun 5;510(7503):162-6. Epub 2014 Jun 4
PubMed.
Narendra D, Tanaka A, Suen DF, Youle RJ.
Parkin is recruited selectively to impaired mitochondria and promotes their autophagy.
J Cell Biol. 2008 Dec 1;183(5):795-803. Epub 2008 Nov 24
PubMed.
Ordureau A, Heo JM, Duda DM, Paulo JA, Olszewski JL, Yanishevski D, Rinehart J, Schulman BA, Harper JW.
Defining roles of PARKIN and ubiquitin phosphorylation by PINK1 in mitochondrial quality control using a ubiquitin replacement strategy.
Proc Natl Acad Sci U S A. 2015 May 26;112(21):6637-42. Epub 2015 May 12
PubMed.
Sauvé V, Lilov A, Seirafi M, Vranas M, Rasool S, Kozlov G, Sprules T, Wang J, Trempe JF, Gehring K.
A Ubl/ubiquitin switch in the activation of Parkin.
EMBO J. 2015 Aug 7;
PubMed.
Thurston TL, Ryzhakov G, Bloor S, von Muhlinen N, Randow F.
The TBK1 adaptor and autophagy receptor NDP52 restricts the proliferation of ubiquitin-coated bacteria.
Nat Immunol. 2009 Nov;10(11):1215-21. Epub 2009 Oct 11
PubMed.
Trempe JF, Sauvé V, Grenier K, Seirafi M, Tang MY, Ménade M, Al-Abdul-Wahid S, Krett J, Wong K, Kozlov G, Nagar B, Fon EA, Gehring K.
Structure of parkin reveals mechanisms for ubiquitin ligase activation.
Science. 2013 Jun 21;340(6139):1451-5. Epub 2013 May 9
PubMed.
Wauer T, Simicek M, Schubert A, Komander D.
Mechanism of phospho-ubiquitin-induced PARKIN activation.
Nature. 2015 Aug 20;524(7565):370-4. Epub 2015 Jul 10
PubMed.
Wild P, Farhan H, McEwan DG, Wagner S, Rogov VV, Brady NR, Richter B, Korac J, Waidmann O, Choudhary C, Dötsch V, Bumann D, Dikic I.
Phosphorylation of the autophagy receptor optineurin restricts Salmonella growth.
Science. 2011 Jul 8;333(6039):228-33. Epub 2011 May 26
PubMed.
Comments
University College London
Nearly 10 years ago, the kinase PINK1 and the E3 ubiquitin ligase Parkin, two proteins associated with autosomal-recessive Parkinson’s disease (PD), were identified as key players in the selective degradation of damaged mitochondria (mitophagy). Since then, we have been presented with a central role for both with stabilization of active PINK1 at the mitochondria recruiting Parkin, Parkin redecorating the outer mitochondrial membrane with ubiquitin, ubiquitin recruiting the autophagy and the proteasome machinery, and ultimately inducing mitophagy.
The identification of PINK1 as the first ubiquitin kinase (Kane et al., 2014; Kazlauskaite et al., 2014; Koyano et al., 2014) has considerably improved our understanding of the mitophagy pathway. Now, this thorough study by Richard Youle’s group emphasizes how indispensable PINK1 is because by generating phospho-ubiquitin, the essential ligand for the two primary autophagy receptors (OPTN and NDP52), it recruits the upstream autophagy machinery to mitochondria, and ultimately induces mitophagy.
Parkin, on the other hand, seems to amplify the mitophagy signal by generating more ubiquitin substrate for PINK1 to phosphorylate, as opposed to being indispensable for mitophagy. Whether Parkin is indispensable for mitophagy in HeLa cells or other E3 ubiquitin ligases can play the same role remains to be determined.
Not only does this study clarify the respective roles of PINK1 and Parkin in mitophagy (and show a new PINK1-dependant/Parkin-independent model of autophagy) but it also highlights the probable important role of mitophagy in the pathogenesis of neurodegenerative conditions other than PD. OPTN and DDF52 are associated with primary angle glaucoma, amyotrophic lateral sclerosis, and Crohn’s disease. Why mutations in PINK1, Parkin, OPTN, NDF52, but also other genes such as VCP and MFN2 can all interfere with the mitophagy pathway while leading to different neurodegenerative conditions may be key in understanding selective cellular vulnerability.
References:
Kane LA, Lazarou M, Fogel AI, Li Y, Yamano K, Sarraf SA, Banerjee S, Youle RJ. PINK1 phosphorylates ubiquitin to activate Parkin E3 ubiquitin ligase activity. J Cell Biol. 2014 Apr 28;205(2):143-53. Epub 2014 Apr 21 PubMed.
Kazlauskaite A, Kondapalli C, Gourlay R, Campbell DG, Ritorto MS, Hofmann K, Alessi DR, Knebel A, Trost M, Muqit MM. Parkin is activated by PINK1-dependent phosphorylation of ubiquitin at Ser65. Biochem J. 2014 May 15;460(1):127-39. PubMed.
Koyano F, Okatsu K, Kosako H, Tamura Y, Go E, Kimura M, Kimura Y, Tsuchiya H, Yoshihara H, Hirokawa T, Endo T, Fon EA, Trempe JF, Saeki Y, Tanaka K, Matsuda N. Ubiquitin is phosphorylated by PINK1 to activate parkin. Nature. 2014 Jun 5;510(7503):162-6. Epub 2014 Jun 4 PubMed.
View all comments by Helene Plun-FavreauMcGill University
McGill University
Commentary: This paper presents an exciting surprise to Parkin/PINK1 and mitophagy community in the form of another important discovery from Richard Youle’s laboratory. The Youle group (among others) had previously discovered the PINK1-dependent Parkin activation and recruitment to damaged mitochondria and the involvement of ubiquitin phosphorylated by PINK1 (pUb) in the process (Narendra et al. 2010; Koyano et al. 2014; Kane et al. 2014; Kazlauskaite et al 2014). These discoveries helped uncover the molecular mechanism of Parkin’s activation (Sauvé et al. 2015; Wauer et al. 2015) from its basal autoinhibited state (Trempe et al. 2013). Here, they show that, while Parkin might be required for efficient mitophagy, the process can still occur independently of Parkin, but relies critically on PINK1’s ubiquitin kinase activity.
The general questions that this study aims to answer are: 1) which autophagy receptors are recruited to mitochondria and are critical for PINK1-dependent mitophagy and 2) does pUb generated by PINK1 have roles in mitophagy besides the allosteric activation of Parkin?
The authors generated gene knockouts of five autophagy cargo receptors to impair mitophagy and show that the reintroduction of only NDP52 or Optineurin (OPTN) rescued the impairment. This rescue occurs even in the absence of Parkin. NDP52 and OPTN (and its phosphorylation by TBK1) have been investigated previously for their roles for xenophagy (Thurston et al. 2009; Wilds et al. 2011) but their involvement in mitophagy is novel. The absence of Parkin helped the authors discriminate better the autophagy receptors that are critical and those that are dispensable. Previous studies (Geisler et al. 2010) had suggested the recruitment of p62 to mitochondria following Parkin activation, but here it is shown not to be essential for mitophagy. The new study demonstrates the value of COXII and mtDNA depletion as markers for mitophagy as MFN1 (a common marker for Parkin’s activity and mitophagy) and TOM20 can be targeted to the proteasome directly by Parkin-mediated ubiquitination in the absence of autophagy.
Youle’s group also discovers another role for pUb: the recruitment of NDP52 and OPTN. They demonstrate that NDP52 and OPTN are preferentially pulled down by phosphomimetic ubiquitin from HeLa cells. The cargo receptor p62 doesn’t have this property. This part of the manuscript will require some additional proof. For example, some experiments were done using phosphomimetic ubiquitin, which doesn’t capture all the functional characteristics of pUb (Ordureau et al. 2015). Also, the use of a Parkin overexpression system in the latter experiments puts the assertion about the Parkin-independence of the process on a difficult footing. Nonetheless, the observations are very significant. Future studies including a paper this week from the Harper group (Heo et al. 2015) will clarify the molecular basis of NDP52 and OPTN recruitment.
Finally, it has been shown that in the absence of NDP52 and OPTN or PINK1 both upstream (ULK1 complex) and downstream components (LC3B) of the autophagy machinery are not effectively recruited, explaining the impairment in mitophagy. The study also shows that mutations in NDP52 and OPTN associated with various diseases including amyotrophic lateral sclerosis (ALS or Lou Gehrig's disease) and Crohn’s disease are impaired in mitophagy and hence indicates that impaired mitophagy might have a contribution towards the observed pathologies. This new study adds significantly to our understanding of mitophagy by assigning roles to different components of the mitochondrial quality control pathway, both upstream and downstream in the process. The findings should also encourage the investigation and development of therapeutics targeting PINK1 activation for the treatment of Parkinson disease.
References:
Geisler S, Holmström KM, Skujat D, Fiesel FC, Rothfuss OC, Kahle PJ, Springer W. PINK1/Parkin-mediated mitophagy is dependent on VDAC1 and p62/SQSTM1. Nat Cell Biol. 2010 Feb;12(2):119-31. Epub 2010 Jan 24 PubMed.
Heo JM, Ordureau A, Paulo JA, Rinehart J, Harper JW. The PINK1-PARKIN Mitochondrial Ubiquitylation Pathway Drives a Program of OPTN/NDP52 Recruitment and TBK1 Activation to Promote Mitophagy. Mol Cell. 2015 Oct 1;60(1):7-20. Epub 2015 Sep 10 PubMed.
Kane LA, Lazarou M, Fogel AI, Li Y, Yamano K, Sarraf SA, Banerjee S, Youle RJ. PINK1 phosphorylates ubiquitin to activate Parkin E3 ubiquitin ligase activity. J Cell Biol. 2014 Apr 28;205(2):143-53. Epub 2014 Apr 21 PubMed.
Kazlauskaite A, Kondapalli C, Gourlay R, Campbell DG, Ritorto MS, Hofmann K, Alessi DR, Knebel A, Trost M, Muqit MM. Parkin is activated by PINK1-dependent phosphorylation of ubiquitin at Ser65. Biochem J. 2014 May 15;460(1):127-39. PubMed.
Koyano F, Okatsu K, Kosako H, Tamura Y, Go E, Kimura M, Kimura Y, Tsuchiya H, Yoshihara H, Hirokawa T, Endo T, Fon EA, Trempe JF, Saeki Y, Tanaka K, Matsuda N. Ubiquitin is phosphorylated by PINK1 to activate parkin. Nature. 2014 Jun 5;510(7503):162-6. Epub 2014 Jun 4 PubMed.
Narendra D, Tanaka A, Suen DF, Youle RJ. Parkin is recruited selectively to impaired mitochondria and promotes their autophagy. J Cell Biol. 2008 Dec 1;183(5):795-803. Epub 2008 Nov 24 PubMed.
Ordureau A, Heo JM, Duda DM, Paulo JA, Olszewski JL, Yanishevski D, Rinehart J, Schulman BA, Harper JW. Defining roles of PARKIN and ubiquitin phosphorylation by PINK1 in mitochondrial quality control using a ubiquitin replacement strategy. Proc Natl Acad Sci U S A. 2015 May 26;112(21):6637-42. Epub 2015 May 12 PubMed.
Sauvé V, Lilov A, Seirafi M, Vranas M, Rasool S, Kozlov G, Sprules T, Wang J, Trempe JF, Gehring K. A Ubl/ubiquitin switch in the activation of Parkin. EMBO J. 2015 Aug 7; PubMed.
Thurston TL, Ryzhakov G, Bloor S, von Muhlinen N, Randow F. The TBK1 adaptor and autophagy receptor NDP52 restricts the proliferation of ubiquitin-coated bacteria. Nat Immunol. 2009 Nov;10(11):1215-21. Epub 2009 Oct 11 PubMed.
Trempe JF, Sauvé V, Grenier K, Seirafi M, Tang MY, Ménade M, Al-Abdul-Wahid S, Krett J, Wong K, Kozlov G, Nagar B, Fon EA, Gehring K. Structure of parkin reveals mechanisms for ubiquitin ligase activation. Science. 2013 Jun 21;340(6139):1451-5. Epub 2013 May 9 PubMed.
Wauer T, Simicek M, Schubert A, Komander D. Mechanism of phospho-ubiquitin-induced PARKIN activation. Nature. 2015 Aug 20;524(7565):370-4. Epub 2015 Jul 10 PubMed.
Wild P, Farhan H, McEwan DG, Wagner S, Rogov VV, Brady NR, Richter B, Korac J, Waidmann O, Choudhary C, Dötsch V, Bumann D, Dikic I. Phosphorylation of the autophagy receptor optineurin restricts Salmonella growth. Science. 2011 Jul 8;333(6039):228-33. Epub 2011 May 26 PubMed.
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