Molecular Composition and Ultrastructure of the Caveolar Coat Complex

Ludwig, Alexander; Howard, Gillian; Mendoza-Topaz, Carolina; Deerinck, Thomas J.; Mackey, Mason; Sandin, Sara; Ellisman, Mark H.; Nichols, Benjamin J.

doi:10.1371/journal.pbio.1001640

Cited by 144 publications

(232 citation statements)

References 56 publications

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“…The cavins also have the ability to bind to PtdSer (16,17,20). Furthermore, a recent study demonstrated that there are 50 cavin1 molecules per caveola (15,21). Together these observations suggest that PtdSer may play a critical role in the formation and stabilization of caveolae.…”

Section: Edited By Dennis R Voelkermentioning

confidence: 91%

Phosphatidylserine dictates the assembly and dynamics of caveolae in the plasma membrane

Hirama

Das

Yang

et al. 2017

Journal of Biological Chemistry

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Edited by Dennis R. VoelkerCaveolae are bulb-shaped nanodomains of the plasma membrane that are enriched in cholesterol and sphingolipids. They have many physiological functions, including endocytic transport, mechanosensing, and regulation of membrane and lipid transport. Caveola formation relies on integral membrane proteins termed caveolins (Cavs) and the cavin family of peripheral proteins. Both protein families bind anionic phospholipids, but the precise roles of these lipids are unknown. Here, we studied the effects of phosphatidylserine (PtdSer), phosphatidylinositol 4-phosphate (PtdIns4P), and phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P 2 ) on caveolar formation and dynamics. Using live-cell, single-particle tracking of GFP-labeled Cav1 and ultrastructural analyses, we compared the effect of PtdSer disruption or phosphoinositide depletion with caveola disassembly caused by cavin1 loss. We found that PtdSer plays a crucial role in both caveola formation and stability. Sequestration or depletion of PtdSer decreased the number of detectable Cav1-GFP puncta and the number of caveolae visualized by electron microscopy. Under PtdSer-limiting conditions, the co-localization of Cav1 and cavin1 was diminished, and cavin1 degradation was increased. Using rapamycin-recruitable phosphatases, we also found that the acute depletion of PtdIns4P and PtdIns (4,5)P 2 has minimal impact on caveola assembly but results in decreased lateral confinement. Finally, we show in a model of phospholipid scrambling, a feature of apoptotic cells, that caveola stability is acutely affected by the scrambling. We conclude that the predominant plasmalemmal anionic lipid PtdSer is essential for proper Cav clustering, caveola formation, and caveola dynamics and that membrane scrambling can perturb caveolar stability.

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Section: Edited By Dennis R Voelkermentioning

confidence: 91%

Phosphatidylserine dictates the assembly and dynamics of caveolae in the plasma membrane

Hirama

Das

Yang

et al. 2017

Journal of Biological Chemistry

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“…The caveolae coat can be purified as a detergent-resistant high-molecular-mass complex of around 60-80S composed of caveolins and cavins at a proportional ratio (Hayer et al, 2010;Ludwig et al, 2013). Interestingly, however, the cavins have been shown to form different subcomplexes in different tissues, which might reflect specific modes of action of the cavin coat that are dependent on the precise cavin composition (Hansen et al, 2013).…”

Section: Introductionmentioning

confidence: 99%

“…Interestingly, however, the cavins have been shown to form different subcomplexes in different tissues, which might reflect specific modes of action of the cavin coat that are dependent on the precise cavin composition (Hansen et al, 2013). Cavin1 was proposed to form homo-trimers and has been shown to interact and form complexes with either cavin2 or cavin3 in a fixed stoichiometry (Ludwig et al, 2013). The competition between cavin2 and cavin3 for binding to cavin1 suggests that these proteins play distinct regulatory roles in the caveolae coat.…”

Section: Introductionmentioning

confidence: 99%

Cavin3 interacts with cavin1 and caveolin1 to increase surface dynamics of caveolae

Mohan

Morén

Larsson

et al. 2015

Journal of Cell Science

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Caveolae are invaginations of the cell surface thought to regulate membrane tension, signalling, adhesion and lipid homeostasis owing to their dynamic behaviour ranging from stable surface association to dynamic rounds of fission and fusion with the plasma membrane. The caveolae coat is generated by oligomerisation of the membrane protein caveolin and the family of cavin proteins. Here, we show that cavin3 (also known as PRKCDBP) is targeted to caveolae by cavin1 (also known as PTRF) where it interacts with the scaffolding domain of caveolin1 and promote caveolae dynamics. We found that the N-terminal region of cavin3 binds a trimer of the cavin1 N-terminus in competition with a homologous cavin2 (also known as SDPR) region, showing that the cavins form distinct subcomplexes through their N-terminal regions. Our data shows that cavin3 is enriched at deeply invaginated caveolae and that loss of cavin3 in cells results in an increase of stable caveolae and a decrease of caveolae that are only present at the membrane for a short time. We propose that cavin3 is recruited to the caveolae coat by cavin1 to interact with caveolin1 and regulate the duration time of caveolae at the plasma membrane.

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“…GFP is 27 kDa), but many of them are too small for direct in situ visualization using cryo-ET. Therefore, for EM visualization, small clonable tags are often specifically linked to additional electron-dense probes, such as metal ion binding by metallothionein (7 kDa) (16,18), or tags such as MiniSOG that catalyze the photooxidation of diaminobenzidine into a localized precipitate that can be visualized by OsO 4 treatment (19,20). Despite these recent advances, the techniques used are still limited and/or are not compatible with cryo-EM, because of low contrast, toxicity of metal ion solutions, and/or the requirement to chemically fix and post-stain the cells.…”

mentioning

confidence: 99%

In Situ Localization of N and C Termini of Subunits of the Flagellar Nexin-Dynein Regulatory Complex (N-DRC) Using SNAP Tag and Cryo-electron Tomography

Song

Awata

Tritschler

et al. 2015

Journal of Biological Chemistry

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Background: Techniques to localize proteins in situ at high resolution are important but limited. Results: Combining SNAP tag technology with cryo-electron tomography, we precisely localized proteins within the N-DRC that are important for ciliary motility. Conclusion: The developed method was applied to localize proteins with ϳ3 nm resolution without interfering with the complex function. Significance: The method is a powerful tool for studies of proteins in situ.

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Molecular Composition and Ultrastructure of the Caveolar Coat Complex

Abstract: The single protein caveolar coat complex comprises only cavins and caveolins, coats the caveolar bulb, and is probably responsible for creating caveolae.

Cited by 144 publications

References 56 publications

Phosphatidylserine dictates the assembly and dynamics of caveolae in the plasma membrane

Phosphatidylserine dictates the assembly and dynamics of caveolae in the plasma membrane

Cavin3 interacts with cavin1 and caveolin1 to increase surface dynamics of caveolae

In Situ Localization of N and C Termini of Subunits of the Flagellar Nexin-Dynein Regulatory Complex (N-DRC) Using SNAP Tag and Cryo-electron Tomography

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