Mechanism and Determinants of Amphipathic Helix-Containing Protein Targeting to Lipid Droplets

Prévost, Coline; Sharp, Morris E.; Kory, Nora; Lin, Qingqing; Voth, Gregory A.; Farese, Robert V.; Walther, Tobias C.

doi:10.1016/j.devcel.2017.12.011

Cited by 203 publications

(295 citation statements)

References 97 publications

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“…2D). This arrangement is characteristic of amphipathic peripheral membrane helices; 25 further, none of our analyses predict these sequences to span the bilayer.…”

Section: A Helix-loop-helix Motif Is Conserved Across Srrscontrasting

confidence: 58%

“…This model also suggests an electrostatic interaction between the basic residues H374 and R381 and phospholipid head groups, which may contribute to membrane association. 25 The 2 nd helix of the hSHH SRR contains a HWY motif that is largely conserved among species and Hh isoforms. While this motif has been demonstrated to regulate post-cleavage trafficking of hSHH-C, 35 a contribution to cholesterolysis has not been tested.…”

Section: Conserved Srr Residues Are Required For Hshh Cholesterolysismentioning

confidence: 99%

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Dual roles of the Sterol Recognition Region in Hedgehog protein modification

Purohit

Peng

Vielmas

et al. 2020

Preprint

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Nature provides a number of mechanisms to encode dynamic information in biomolecules. In metazoans, there exist rare chemical modifications that occur through entirely unique mechanistic regimes. One such example occurs in the Hedgehog (Hh) morphogens, proteins singular across all domains of life for the nature of their covalent ligation to cholesterol. The isoform-and context-specific efficiency of the ligation reaction has profound impact on the activity of Hh morphogens and represents an unexplored aspect of Hh ligand-dependent cancers. To elucidate the chemical mechanism of this modification, we have defined roles of the uncharacterized sterol recognition region (SRR) in Hh proteins. We use a combination of sequence conservation, directed mutagenesis, and biochemical assays to specify residues of the SRR that are responsible for cellular and biochemical processes in Hh cholesterolysis. Our investigations offer the first functional template of this region, providing opportunities to identify parallel reactivity in nature and revealing new mechanisms that can be exploited as tools in chemical biology.

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“…2D). This arrangement is characteristic of amphipathic peripheral membrane helices; 25 further, none of our analyses predict these sequences to span the bilayer.…”

Section: A Helix-loop-helix Motif Is Conserved Across Srrscontrasting

confidence: 58%

Section: Conserved Srr Residues Are Required For Hshh Cholesterolysismentioning

confidence: 99%

Dual roles of the Sterol Recognition Region in Hedgehog protein modification

Purohit

Peng

Vielmas

et al. 2020

Preprint

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“…Since viroplasms have been shown to form on lipid droplets (LDs) (32) conformational changes in NSP2, or NSP5, may expose lipophilic residues that allow the proteins to interact with cellular lipids bound for LDs. Several cellular proteins are inserted into LD membranes via amphipathic helices, e.g., perilipin (34). RV NSP5 possesses an amphipathic helix which theoretically could bind to LD membranes.…”

Section: Discussionmentioning

confidence: 99%

Phosphorylation cascade regulates the formation and maturation of rotaviral replication factories

Criglar

Anish

et al. 2018

Proc. Natl. Acad. Sci. U.S.A.

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The rotavirus (RV) genome is replicated and packaged into virus progeny in cytoplasmic inclusions called viroplasms, which require interactions between RV nonstructural proteins NSP2 and NSP5. How viroplasms form remains unknown. We previously found two forms of NSP2 in RV-infected cells: a cytoplasmically dispersed dNSP2, which interacts with hypophosphorylated NSP5; and a viroplasm-specific vNSP2, which interacts with hyperphosphorylated NSP5. Other studies report that CK1α, a ubiquitous cellular kinase, hyperphosphorylates NSP5, but requires NSP2 for reasons that are unclear. Here we show that silencing CK1α in cells before RV infection resulted in (i) >90% decrease in RV replication, (ii) disrupted vNSP2 and NSP5 interaction, (iii) dispersion of vNSP2 throughout the cytoplasm, and (iv) reduced vNSP2 protein levels. Together, these data indicate that CK1α directly affects NSP2. Accordingly, an in vitro kinase assay showed that CK1α phosphorylates serine 313 of NSP2 and triggers NSP2 octamers to form a lattice structure as demonstrated by crystallographic analysis. Additionally, a dual-specificity autokinase activity for NSP2 was identified and confirmed by mass spectrometry. Together, our studies show that phosphorylation of NSP2 involving CK1α controls viroplasm assembly. Considering that CK1α plays a role in the replication of other RNA viruses, similar phosphorylation-dependent mechanisms may exist for other virus pathogens that require cytoplasmic virus factories for replication.M any viral pathogens, including both DNA viruses (poxviruses) and RNA viruses [rotavirus (RV), orthoreovirus, hepatitis C virus, dengue virus, Marburg virus] replicate in cytoplasmic compartments that are physical scaffolds composed of both viral and cellular proteins (1-3). These compartments, excluded from the rest of the cytoplasm, are sites where the virus genome is replicated and progeny particles are assembled. Such replication compartments are alternately known as virus factories, virus replication centers or complexes, and viroplasms. Exactly how these virus factories form is largely unknown, so it is intriguing to consider that there may be common mechanisms that initiate their formation and regulate their assembly.RV-induced severe, dehydrating gastroenteritis remains a leading cause of morbidity and mortality in infants and children less than 5 y of age worldwide and accounts for greater than 200,000 deaths annually (4). To begin to understand the mechanism of replication factory assembly, we studied the assembly of RV replication complexes known as viroplasms. Viroplasm formation requires the interaction of two nonstructural proteins, NSP2 and NSP5, and expression of NSP2 with NSP5 in cells lacking all other viral proteins results in the formation of viroplasm-like structures (VLS) (5). Loss of either NSP2 or NSP5 abolishes viroplasm formation and rotavirus replication (6-9). While information about the individual characteristics of NSP2 and NSP5 and some knowledge of how they physically interact are available, exactly...

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“…A second possibility is that the distinct lipid composition of LDs is key to protein targeting. With its neutral lipid core, monolayer membrane (possible bare neutral lipid interacts with protein directly due to the phospholipid packing defect), and distinct polar lipid composition, the LD may be a favorable binding site for some proteins. A third possible mechanism is the translation of the proteins at the LD surface.…”

Section: The Subcellular Sorting Mechanism Of Proteinsmentioning

confidence: 99%

The New Face of the Lipid Droplet: Lipid Droplet Proteins

Zhang

Liu

2018

Proteomics

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The lipid droplet (LD) is an organelle with vital functions found in nearly all organisms. LD proteomic research has provided fundamentally important insights into this organelle's functions. The review provides a summary of LD proteomic studies conducted across diverse organisms and cell and tissue types. The accumulated proteomic data are reviewed for evidence of a protein targeting mechanism for the organelle. The hypotheses for several specific localization mechanisms based on what is known about targeting mechanisms for other organelles and vesicles are provided. Although the nature of the mechanism is not known, the functional data demonstrate that the targeting mechanism and, indeed, the organelle itself, is conserved from prokaryotes to eukaryotes. It is hoped that the review will help inspire further research leading to novel discoveries in the field.

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Mechanism and Determinants of Amphipathic Helix-Containing Protein Targeting to Lipid Droplets

Cited by 203 publications

References 97 publications

Dual roles of the Sterol Recognition Region in Hedgehog protein modification

Dual roles of the Sterol Recognition Region in Hedgehog protein modification

Phosphorylation cascade regulates the formation and maturation of rotaviral replication factories

The New Face of the Lipid Droplet: Lipid Droplet Proteins

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