The Proline/Arginine-Rich Domain Is a Major Determinant of Dynamin Self-Activation

Baryłko, Barbara; Wang, Lei; Binns, Derk D.; Ross, Justin A.; Tassin, Tara C.; Collins, Katie A.; Jameson, David M.; Albanesi, Joseph P.

doi:10.1021/bi101343p

Cited by 14 publications

(15 citation statements)

References 23 publications

(42 reference statements)

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“…3e) indicates that dimerization specificity is controlled by a spatial combinatorial code. As predicted from this code, hetero-oligomers consisting of dynamin 1 and dynamin 2 are observed 35 . Several residues contributing to the apparent specificity are localized in the strongly conserved coil of dynamin, Drp1 and Dnm1 that follows α3.…”

Section: The Dynamin–dimer Interfacesupporting

confidence: 59%

The crystal structure of dynamin

Ford

Jenni

Nunnari

2011

Nature

243

313

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Dynamin-related proteins (DRPs) are multi-domain GTPases that function via oligomerization and GTP-dependent conformational changes to play central roles in regulating membrane structure across phylogenetic kingdoms. How DRPs harness self-assembly and GTP-dependent conformational changes to remodel membranes is not understood. Here we present the crystal structure of an assembly-deficient mammalian endocytic DRP, dynamin 1, lacking the proline-rich domain, in its nucleotide-free state. The dynamin 1 monomer is an extended structure with the GTPase domain and bundle signalling element positioned on top of a long helical stalk with the pleckstrin homology domain flexibly attached on its opposing end. Dynamin 1 dimer and higher order dimer multimers form via interfaces located in the stalk. Analysis of these interfaces provides insight into DRP family member specificity and regulation and provides a framework for understanding the biogenesis of higher order DRP structures and the mechanism of DRP-mediated membrane scission events.

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Section: The Dynamin–dimer Interfacesupporting

confidence: 59%

The crystal structure of dynamin

Ford

Jenni

Nunnari

2011

Nature

243

313

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“…The GTPase activities of dynamins are stimulated by their self-assembly into rings and coils (26, 34, 36). Therefore, it seemed likely that the enhancement of Dyn2 polymerization by His 6 -Arc would be accompanied by an increase in Dyn2 GTPase activity.…”

Section: Resultsmentioning

confidence: 99%

“…This finding may simply reflect the fact that Dyn1 has a much lower intrinsic propensity to self-assemble than the other two forms of dynamin (26,36), suggesting that Arc is a less potent scaffold for dynamins than microtubules or liposomes, which have a much higher valency. However, the fact that Dyn1 is predominantly expressed in presynaptic terminals, whereas Arc is absent from presynaptic terminals and axons but, like Dyns 2 and 3, is abundantly expressed in dendrites (9), may point to a more specific reason for this selectivity.…”

Section: Discussionmentioning

confidence: 99%

Enhancement of dynamin polymerization and GTPase activity by Arc/Arg3.1

Byers

Baryłko

Ross

et al. 2015

Biochimica et Biophysica Acta (BBA) - General Subjects

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BACKGROUND The Activity-regulated cytoskeleton-associated protein, Arc, is an immediate-early gene product implicated in various forms of synaptic plasticity. Arc promotes endocytosis of AMPA type glutamate receptors and regulates cytoskeletal assembly in neuronal dendrites. Its role in endocytosis may be mediated by its reported interaction with dynamin 2 (Dyn2), a 100 kDa GTPase that polymerizes around the necks of budding vesicles and catalyzes membrane scission. METHODS Enzymatic and turbidity assays are used in this study to monitor effects of Arc on dynamin activity and polymerization. Arc oligomerization is measured using a combination of approaches, including size exclusion chromatography, sedimentation analysis, dynamic light scattering, fluorescence correlation spectroscopy, and electron microscopy. RESULTS We present evidence that bacterially-expressed His6-Arc facilitates the polymerization of Dyn2 and stimulates its GTPase activity under physiologic conditions (37°C and 100 mM NaCl). At lower ionic strength Arc also stabilizes pre-formed Dyn2 polymers against GTP-dependent disassembly, thereby prolonging assembly-dependent GTP hydrolysis catalyzed by Dyn2. Arc also increases the GTPase activity of Dyn3, an isoform of implicated in dendrite remodeling, but does not affect the activity of Dyn1, a neuron-specific isoform involved in synaptic vesicle recycling. We further show in this study that Arc (either His6-tagged or untagged) has a tendency to form large soluble oligomers, which may function as a scaffold for dynamin assembly and activation. CONCLUSIONS and GENERAL SIGNIFICANCE The ability of Arc to enhance dynamin polymerization and GTPase activation may provide a mechanism to explain Arc-mediated endocytosis of AMPA receptors and the accompanying effects on synaptic plasticity. This study represents the first detailed characterization of the physical properties of Arc.

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“…Previous in vitro studies aimed at defining the size of the minimal dynamin assembly unit have yielded conflicting results. For example, our sedimentation equilibrium measurements in the analytical ultracentrifuge indicated that all three forms of dynamin exist predominantly as monomer-tetramer or monomer-dimer-tetramer equilibria in solutions containing 300 mM NaCl [32-34]. The R369W mutation did not significantly alter sedimentation behavior of Dyn2 [20].…”

Section: Resultsmentioning

confidence: 99%

A mutation associated with centronuclear myopathy enhances the size and stability of dynamin 2 complexes in cells

James

Digman

Ross

et al. 2014

Biochimica et Biophysica Acta (BBA) - General Subjects

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Background Dynamin 2 (Dyn2) is a ~100 kDa GTPase that assembles around the necks of nascent endocytic and Golgi vesicles and catalyzes membrane scission. Mutations in Dyn2 that cause Centronuclear Myopathy (CNM) have been shown to stabilize Dyn2 polymers against GTP-dependent disassembly in vitro. Precisely timed regulation of assembly and disassembly is believed to be critical for Dyn2 function in membrane vesiculation, and the CNM mutations interfere with this regulation by shifting the equilibrium toward the assembled state. Methods In this study we use two fluorescence fluctuation spectroscopy (FFS) approaches to show that a CNM mutant form of Dyn2 also has a greater propensity to self-assemble in the cytosol and on the plasma membrane of living cells. Results Results obtained using brightness analysis indicate that unassembled wild-type Dyn2 is predominantly tetrameric in the cytosol, although different oligomeric species are observed, depending on the concentration of expressed protein. In contrast, an R369W mutant identified in CNM patients forms higher-order oligomers at concentrations above 1 μM. Investigation of Dyn2-R369W by Total Internal Reflection Fluorescence (TIRF) FFS reveals that this mutant forms larger and more stable clathrin-containing structures on the plasma membrane than wild-type Dyn2. Conclusions and General Significance These observations may explain defects in membrane trafficking reported in CNM patient cells and in heterologous systems expressing CNM-associated Dyn2 mutants.

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The Proline/Arginine-Rich Domain Is a Major Determinant of Dynamin Self-Activation

Cited by 14 publications

References 23 publications

The crystal structure of dynamin

The crystal structure of dynamin

Enhancement of dynamin polymerization and GTPase activity by Arc/Arg3.1

A mutation associated with centronuclear myopathy enhances the size and stability of dynamin 2 complexes in cells

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