Prion Aggregates Are Recruited to the Insoluble Protein Deposit (IPOD) via Myosin 2-Based Vesicular Transport.
Aggregation of amyloidogenic proteins is associated with several neurodegenerative diseases. Sequestration of misfolded and aggregated proteins into specialized deposition sites may reduce their potentially detrimental properties. Yeast exhibits a distinct deposition site for amyloid aggregates termed "Insoluble PrOtein Deposit (IPOD)", but nothing is known about the mechanism of substrate recruitment to this site. The IPOD is located directly adjacent to the Phagophore Assembly Site (PAS) where the cell initiates autophagy and the Cytoplasm-to-Vacuole Targeting (CVT) pathway destined for delivery of precursor peptidases to the vacuole. Recruitment of CVT substrates to the PAS was proposed to occur via vesicular transport on Atg9 vesicles and requires an intact actin cytoskeleton and "SNAP (Soluble NSF Attachment Protein) Receptor Proteins (SNARE)" protein function. It is, however, unknown how this vesicular transport machinery is linked to the actin cytoskeleton. We demonstrate that recruitment of model amyloid PrD-GFP and the CVT substrate precursor-aminopeptidase 1 (preApe1) to the IPOD or PAS, respectively, is disturbed after genetic impairment of Myo2-based actin cable transport and SNARE protein function. Rather than accumulating at the respective deposition sites, both substrates reversibly accumulated often together in the same punctate structures. Components of the CVT vesicular transport machinery including Atg8 and Atg9 as well as Myo2 partially co-localized with the joint accumulations. Thus we propose a model where vesicles, loaded with preApe1 or PrD-GFP, are recruited to tropomyosin coated actin cables via the Myo2 motor protein for delivery to the PAS and IPOD, respectively. We discuss that deposition at the IPOD is not an integrated mandatory part of the degradation pathway for amyloid aggregates, but more likely stores excess aggregates until downstream degradation pathways have the capacity to turn them over after liberation by the Hsp104 disaggregation machinery.
Department of Medicine I and Clinical Chemistry, Heidelberg University Hospital, Heidelberg, Germany.
Rajesh
http://orcid.org/0000-0001-9346-8806
R
Kumar
Department of Medicine I and Clinical Chemistry, Heidelberg University Hospital, Heidelberg, Germany.|Joint Heidelberg-IDC Translational Diabetes Program, Dept. Inner Medicine I, Heidelberg University Hospital, Germany.|German Center for Diabetes Research (DZD), München-Neuherberg, Germany.
Peter P
PP
Nawroth
Department of Medicine I and Clinical Chemistry, Heidelberg University Hospital, Heidelberg, Germany.
Jens
J
Tyedmers
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