Phase separation organizes the site of autophagosome formation

Fujioka, Yūko; Alam, Jahangir Md.; Noshiro, Daisuke; Mouri, Kazunari; Ando, Toshio; Okada, Yasushi; May, Alexander I.; Knorr, Roland L.; Suzuki, Kuninori; Ohsumi, Yoshinori; Noda, Nobuo N.

doi:10.1038/s41586-020-1977-6

Cited by 312 publications

(275 citation statements)

References 42 publications

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“…Our results suggest that the interaction of nucleoids with membranes is due to the emergent wetting behavior of the condensate on the membrane (Snead and Gladfelter, 2019). Similar observations have been made for contact sites between tethering proteins from various membrane bound organelles, including the mitochondrial mitofusin 1 (Mfn1) tethering protein and Sec61b of the ER membrane (King et al, 2020), and for the condensation of Atg1-complex droplets, as part of the pre-autophagosomal structure (PAS), along vacuolar membranes (Fujioka et al, 2020). It is tempting to speculate that the wetting behavior between the mitochondrial inner membrane and the nucleoid may not only play a role in regulating the size and diffusion of nucleoids, but also in functional processes such as replication (Lewis et al, 2016).…”

Section: Discussionsupporting

confidence: 80%

Self-assembly of multi-component mitochondrial nucleoids via phase separation

Feric

Demarest

Tian

et al. 2019

Preprint

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Mitochondria contain an autonomous and spatially segregated genome. The organizational unit of mitochondrial genomes is the nucleoid which consists of mitochondrial DNA (mtDNA) and associated architectural proteins. The physical properties and mechanisms of assembly of mitochondrial nucleoids remain largely unclear. Here, we show that mitochondrial nucleoids form by phase separation. Mitochondrial nucleoids exhibit morphological features, coarsening behavior, and dynamics of phase-separated structures in vitro and in vivo. The major mtDNA-binding protein TFAM spontaneously phase separates in vitro into viscoelastic droplets with slow internal dynamics, and the combination of TFAM and mtDNA is sufficient to promote the in vitro formation of multiphase structures with gel-like properties, which recapitulate the in vivo behavior of mitochondrial nucleoids in live human cells. Enlarged phase-separated mitochondrial nucleoids are present in the premature aging disease Hutchinson-Gilford Progeria Syndrome (HGPS) and altered mitochondrial nucleoid structure is accompanied by mitochondrial dysfunction. These results identify phase separation as the physical mechanism driving the organization of mitochondrial nucleoids, and they have ramifications for mitochondrial function.

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Section: Discussionsupporting

confidence: 80%

Self-assembly of multi-component mitochondrial nucleoids via phase separation

Feric

Demarest

Tian

et al. 2019

Preprint

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“…Like many other proteins associated with neurodegenerative diseases [13][14][15][16][17][18] , UBQLN2 spontaneously phase separates to form condensates, or liquid droplets, in which proteins are concentrated yet remain mobile 13,19,20 . Liquid-like condensates are tightly regulated by cells and can provide sites for specific cellular functions [21][22][23][24] . Evidence for this includes a recent finding that UBQLN2 regulates the fluidity of protein-RNA complexes in FUS-related ALS/FTD 25 .…”

Section: Introductionmentioning

confidence: 99%

Shared and divergent phase separation and aggregation properties of brain-expressed ubiquilins

Gerson

Linton

Xing

et al. 2020

Preprint

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The brain-expressed ubiquilins, UBQLNs 1, 2 and 4, are highly homologous proteins that participate in multiple aspects of protein homeostasis and are implicated in neurodegenerative diseases. Studies have established that UBQLN2 forms liquid-like condensates and accumulates in pathogenic aggregates, much like other proteins linked to neurodegenerative diseases. However, the relative condensate and aggregate formation of the three brain-expressed ubiquilins is unknown. We report that the three ubiquilins differ in aggregation propensity, revealed by in-vitro experiments, cellular models, and analysis of human brain tissue. UBQLN4 displays heightened aggregation propensity over the other ubiquilins and, like amyloids, UBQLN4 forms ThioflavinT-positive fibrils in vitro. Measuring fluorescence recovery after photobleaching (FRAP) of puncta in cells, we report that all three ubiquilins undergo liquid-liquid phase transition. UBQLN2 and 4 exhibit slower recovery than UBQLN1, suggesting the condensates formed by the brain-expressed ubiquilins have different compositions and undergo distinct internal rearrangements. We conclude that while all brain-expressed ubiquilins exhibit self-association behavior manifesting as condensates, they follow distinct courses of phase-separation and levels of aggregation. This variability among ubiquilins along the continuum from liquid-like to solid likely informs both the normal ubiquitin-linked functions of ubiquilins and their accumulation and potential contribution to toxicity in various neurodegenerative diseases.

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“…High-speed AFM provides real-time video of the protein in solution at nm-resolution on the second or sub-second timescale, and has been used to investigate the polymers of LC proteins (Babinchak et al 2019) and gels of protein fibers (Cui et al 2019;Wang et al 2019). More recently, high-speed AFM has been utilized in the observation of the protein in the LLPS droplet (Fujioka et al 2020).…”

Section: Other Biochemical and Biophysical Methodsmentioning

confidence: 99%

Biological phase separation: cell biology meets biophysics

et al. 2020

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Progress in development of biophysical analytic approaches has recently crossed paths with macromolecule condensates in cells. These cell condensates, typically termed liquid-like droplets, are formed by liquid-liquid phase separation (LLPS). More and more cell biologists now recognize that many of the membrane-less organelles observed in cells are formed by LLPS caused by interactions between proteins and nucleic acids. However, the detailed biophysical processes within the cell that lead to these assemblies remain largely unexplored. In this review, we evaluate recent discoveries related to biological phase separation including stress granule formation, chromatin regulation, and processes in the origin and evolution of life. We also discuss the potential issues and technical advancements required to properly study biological phase separation.

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Phase separation organizes the site of autophagosome formation

Cited by 312 publications

References 42 publications

Self-assembly of multi-component mitochondrial nucleoids via phase separation

Self-assembly of multi-component mitochondrial nucleoids via phase separation

Shared and divergent phase separation and aggregation properties of brain-expressed ubiquilins

Biological phase separation: cell biology meets biophysics

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