Global ubiquitylation analysis of mitochondria in primary neurons identifies endogenous Parkin targets following activation of PINK1

TY - JOUR

T1 - Global ubiquitylation analysis of mitochondria in primary neurons identifies endogenous Parkin targets following activation of PINK1

AU - Antico, Odetta

AU - Ordureau, Alban

AU - Stevens, Michael

AU - Singh, Francois

AU - Nirujogi, Raja S.

AU - Gierlinski, Marek

AU - Barini, Erica

AU - Rickwood, Mollie L.

AU - Prescott, Alan

AU - Toth, Rachel

AU - Ganley, Ian G.

AU - Harper, J. Wade

AU - Muqit, Miratul M. K.

N1 - Funding: This work was supported by a Wellcome Trust Senior Research Fellowship in Clinical Science (210753/Z/18/Z to M.M.K.M.), the Rosetrees Trust (PhD Studentship to O.A.), EMBO YIP Award and EMBO Small Grant (M.M.K.M.), NIH (RO1 NS083524 to J.W.H.), the Michael J. Fox Foundation (J.W.H. and M.M.K.M.), Medical Research Council (MC_UU_00018/2 to I.G.G.), a gift from N. Goodnow (J.W.H.), and Aligning Science Across Parkinson’s (ASAP) initiative. Michael J. Fox Foundation administers the grant (ASAP-000463; 000282) on behalf of ASAP and itself (J.W.H. and M.M.K.M). M.L.R. was a recipient of a Dundee Research Interest Group (DRIG) studentship, Dundee School of Life Sciences Scholarship, and Bath University Alumni Placement Grant. This research was funded, in part, by Aligning Science Across Parkinson’s. Michael J Fox Foundation administers the grants, ASAP-000463 and ASAP-000282 on behalf of ASAP and itself.

PY - 2021/11/12

Y1 - 2021/11/12

N2 - How activation of PINK1 and Parkin leads to elimination of damaged mitochondria by mitophagy is largely based on cell lines with few studies in neurons. Here, we have undertaken proteomic analysis of mitochondria from mouse neurons to identify ubiquitylated substrates of endogenous Parkin. Comparative analysis with human iNeuron datasets revealed a subset of 49 PINK1 activation–dependent diGLY sites in 22 proteins conserved across mouse and human systems. We use reconstitution assays to demonstrate direct ubiquitylation by Parkin in vitro. We also identified a subset of cytoplasmic proteins recruited to mitochondria that undergo PINK1 and Parkin independent ubiquitylation, indicating the presence of alternate ubiquitin E3 ligase pathways that are activated by mitochondrial depolarization in neurons. Last, we have developed an online resource to search for ubiquitin sites and enzymes in mitochondria of neurons, MitoNUb. These findings will aid future studies to understand Parkin activation in neuronal subtypes.

AB - How activation of PINK1 and Parkin leads to elimination of damaged mitochondria by mitophagy is largely based on cell lines with few studies in neurons. Here, we have undertaken proteomic analysis of mitochondria from mouse neurons to identify ubiquitylated substrates of endogenous Parkin. Comparative analysis with human iNeuron datasets revealed a subset of 49 PINK1 activation–dependent diGLY sites in 22 proteins conserved across mouse and human systems. We use reconstitution assays to demonstrate direct ubiquitylation by Parkin in vitro. We also identified a subset of cytoplasmic proteins recruited to mitochondria that undergo PINK1 and Parkin independent ubiquitylation, indicating the presence of alternate ubiquitin E3 ligase pathways that are activated by mitochondrial depolarization in neurons. Last, we have developed an online resource to search for ubiquitin sites and enzymes in mitochondria of neurons, MitoNUb. These findings will aid future studies to understand Parkin activation in neuronal subtypes.

UR - https://www.scopus.com/pages/publications/85119273551

U2 - 10.1126/sciadv.abj0722

DO - 10.1126/sciadv.abj0722

M3 - Article

C2 - 34767452

SN - 2375-2548

VL - 7

SP - 1

EP - 21

JO - Science Advances

JF - Science Advances

IS - 46

M1 - eabj0722

ER -