scholarly journals Assembly and activation of dynein–dynactin by the cargo adaptor protein Hook3

2016 ◽  
Vol 214 (3) ◽  
pp. 309-318 ◽  
Author(s):  
Courtney M. Schroeder ◽  
Ronald D. Vale
Keyword(s):  
Crystal Structure ◽  
Crucial Role ◽  
Ternary Complex ◽  
Adaptor Protein ◽  
Cytoplasmic Dynein ◽  
Allosteric Activation ◽  
Structural Details ◽  
Dynactin Complex ◽  
Insight Into ◽  
Intermediate Chain

Metazoan cytoplasmic dynein moves processively along microtubules with the aid of dynactin and an adaptor protein that joins dynein and dynactin into a stable ternary complex. Here, we examined how Hook3, a cargo adaptor involved in Golgi and endosome transport, forms a motile dynein–dynactin complex. We show that the conserved Hook domain interacts directly with the dynein light intermediate chain 1 (LIC1). By solving the crystal structure of the Hook domain and using structure-based mutagenesis, we identify two conserved surface residues that are each critical for LIC1 binding. Hook proteins with mutations in these residues fail to form a stable dynein–dynactin complex, revealing a crucial role for LIC1 in this interaction. We also identify a region of Hook3 specifically required for an allosteric activation of processive motility. Our work reveals the structural details of Hook3’s interaction with dynein and offers insight into how cargo adaptors form processive dynein–dynactin motor complexes.

scholarly journals Assembly and Activation of Dynein-Dynactin by the Cargo Adaptor Protein Hook3

2016 ◽  
Author(s):  
Courtney M. Schroeder ◽  
Ronald D. Vale
Keyword(s):  
Crystal Structure ◽  
Crucial Role ◽  
Ternary Complex ◽  
Adaptor Protein ◽  
Cytoplasmic Dynein ◽  
Allosteric Activation ◽  
Structural Details ◽  
Dynactin Complex ◽  
Insight Into ◽  
Intermediate Chain

AbstractMetazoan cytoplasmic dynein moves processively along microtubules with the aid of dynactin and an adaptor protein that joins dynein and dynactin into a stable ternary complex. Here, we have examined how Hook3, a cargo adaptor involved in Golgi and endosome transport, forms a motile dynein-dynactin complex. We show that the conserved Hook domain interacts directly with the dynein light intermediate chain 1 (LIC1). By solving the crystal structure of the Hook domain and using structure-based mutagenesis, we identify two conserved surface residues that are each critical for LIC1 binding. Hook proteins with mutations in these residues fail to form a stable dynein-dynactin complex, revealing a crucial role for LIC1 in this interaction. We also identify a region of Hook3 specifically required for an allosteric activation of processive motility. Our work reveals the structural details of Hook3’s interaction with dynein and offers insight into how cargo adaptors form processive dynein-dynactin motor complexes.

scholarly journals Combinatorial regulation of the balance between dynein microtubule end accumulation and initiation of directed motility

2017 ◽  
Author(s):  
Rupam Jha ◽  
Johanna Roostalu ◽  
Martina Trokter ◽  
Thomas Surrey
Keyword(s):  
Adaptor Protein ◽  
Cytoplasmic Dynein ◽  
Dependent Manner ◽  
Cellular Functions ◽  
Combinatorial Regulation ◽  
Directed Motility ◽  
The Individual ◽  
Dynactin Complex ◽  
Insight Into ◽  
Dynamic Microtubule

ABSTRACTCytoplasmic dynein is involved in a multitude of essential cellular functions. Dynein’s activity is controlled by the combinatorial action of several regulators. The molecular mechanism of this regulation is poorly understood. Using purified proteins, we reconstitute the regulation of the human dynein complex by three prominent regulators on dynamic microtubules in the presence of end binding proteins (EBs). We find that dynein can be in biochemically and functionally distinct pools: either passively tracking dynamic microtubule plus-ends in an EB-dependent manner or moving processively towards minus ends in an adaptor protein-dependent manner. Whereas both dynein pools share the dynactin complex, they have opposite preferences for binding other regulators, either the adaptor protein Bicaudal D2 (BicD2) or the multifunctional regulator Lisencephaly-1 (Lis1). Remarkably, dynactin, but not EBs, strongly biases motility initiation locally from microtubule plus ends by autonomous plus end recognition. BicD2 and Lis1 together control the overall efficiency of motility initiation. Our study provides insight into the mechanism of dynein activity regulation by dissecting the distinct functional contributions of the individual members of a dynein regulatory network.

scholarly journals Systematic dissection of dynein regulators in mitosis

2013 ◽  
Vol 201 (2) ◽  
pp. 201-215 ◽  
Author(s):  
Jonne A. Raaijmakers ◽  
Marvin E. Tanenbaum ◽  
René H. Medema
Keyword(s):  
Adaptor Protein ◽  
Adaptor Proteins ◽  
Cytoplasmic Dynein ◽  
Comprehensive Overview ◽  
Motor Complex ◽  
Essential Components ◽  
Complex Thought ◽  
Specific Activities ◽  
Dynactin Complex

Cytoplasmic dynein is a large minus end–directed motor complex with multiple functions during cell division. The dynein complex interacts with various adaptor proteins, including the dynactin complex, thought to be critical for most dynein functions. Specific activities have been linked to several subunits and adaptors, but the function of the majority of components has remained elusive. Here, we systematically address the function of each dynein–dynactin subunit and adaptor protein in mitosis. We identify the essential components that are required for all mitotic functions of dynein. Moreover, we find specific dynein recruitment factors, and adaptors, like Nde1/L1, required for activation, but largely dispensable for dynein localization. Most surprisingly, our data show that dynactin is not required for dynein-dependent spindle organization, but acts as a dynein recruitment factor. These results provide a comprehensive overview of the role of dynein subunits and adaptors in mitosis and reveal that dynein forms distinct complexes requiring specific recruiters and activators to promote orderly progression through mitosis.

scholarly journals The APC-associated protein EB1 associates with components of the dynactin complex and cytoplasmic dynein intermediate chain

1999 ◽  
Vol 9 (8) ◽  
pp. 425-428 ◽  
Author(s):  
Lisbeth Berrueta ◽  
Jennifer S. Tirnauer ◽  
Scott C. Schuyler ◽  
David Pellman ◽  
Barbara E. Bierer
Keyword(s):  
Cytoplasmic Dynein ◽  
Dynein Intermediate Chain ◽  
Dynactin Complex ◽  
Intermediate Chain

scholarly journals Crystal structure of the cohesin loader Scc2 and insight into cohesinopathy

2016 ◽  
Vol 113 (44) ◽  
pp. 12444-12449 ◽  
Author(s):  
Sotaro Kikuchi ◽  
Dominika M. Borek ◽  
Zbyszek Otwinowski ◽  
Diana R. Tomchick ◽  
Hongtao Yu
Keyword(s):  
Crystal Structure ◽  
Sister Chromatid ◽  
Adaptor Protein ◽  
Sister Chromatid Cohesion ◽  
Conserved Residues ◽  
Chaetomium Thermophilum ◽  
Important Interaction ◽  
Chromatid Cohesion ◽  
Structural Maintenance Of Chromosomes ◽  
Insight Into

The ring-shaped cohesin complex topologically entraps chromosomes and regulates chromosome segregation, transcription, and DNA repair. The cohesin core consists of the structural maintenance of chromosomes 1 and 3 (Smc1–Smc3) heterodimeric ATPase, the kleisin subunit sister chromatid cohesion 1 (Scc1) that links the two ATPase heads, and the Scc1-bound adaptor protein Scc3. The sister chromatid cohesion 2 and 4 (Scc2–Scc4) complex loads cohesin onto chromosomes. Mutations of cohesin and its regulators, including Scc2, cause human developmental diseases termed cohesinopathy. Here, we report the crystal structure of Chaetomium thermophilum (Ct) Scc2 and examine its interaction with cohesin. Similar to Scc3 and another Scc1-interacting cohesin regulator, precocious dissociation of sisters 5 (Pds5), Scc2 consists mostly of helical repeats that fold into a hook-shaped structure. Scc2 binds to Scc1 through an N-terminal region of Scc1 that overlaps with its Pds5-binding region. Many cohesinopathy mutations target conserved residues in Scc2 and diminish Ct Scc2 binding to Ct Scc1. Pds5 binding to Scc1 weakens the Scc2–Scc1 interaction. Our study defines a functionally important interaction between the kleisin subunit of cohesin and the hook of Scc2. Through competing with Scc2 for Scc1 binding, Pds5 might contribute to the release of Scc2 from loaded cohesin, freeing Scc2 for additional rounds of loading.

scholarly journals A Ras-like domain in the light intermediate chain bridges the dynein motor to a cargo-binding region

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Courtney M Schroeder ◽  
Jonathan ML Ostrem ◽  
Nicholas T Hertz ◽  
Ronald D Vale
Keyword(s):  
G Proteins ◽  
Heavy Chain ◽  
Structural Information ◽  
Cytoplasmic Dynein ◽  
Protein Fold ◽  
Binding Region ◽  
Aromatic Residues ◽  
Dynein Motor ◽  
Insight Into ◽  
Intermediate Chain

Cytoplasmic dynein, a microtubule-based motor protein, transports many intracellular cargos by means of its light intermediate chain (LIC). In this study, we have determined the crystal structure of the conserved LIC domain, which binds the motor heavy chain, from a thermophilic fungus. We show that the LIC has a Ras-like fold with insertions that distinguish it from Ras and other previously described G proteins. Despite having a G protein fold, the fungal LIC has lost its ability to bind nucleotide, while the human LIC1 binds GDP preferentially over GTP. We show that the LIC G domain binds the dynein heavy chain using a conserved patch of aromatic residues, whereas the less conserved C-terminal domain binds several Rab effectors involved in membrane transport. These studies provide the first structural information and insight into the evolutionary origin of the LIC as well as revealing how this critical subunit connects the dynein motor to cargo.

scholarly journals Allosteric activation of CwlD amidase activity by the GerS lipoprotein during Clostridioides difficile spore formation

2021 ◽  
Author(s):  
Carolina Alves Feliciano ◽  
Brian E Eckenroth ◽  
Oscar R Diaz ◽  
Syvlie Doublie ◽  
Aimee Shen
Keyword(s):  
Crystal Structure ◽  
Spore Germination ◽  
Active Sites ◽  
Catalytic Mechanism ◽  
Allosteric Activation ◽  
Amidase Activity ◽  
Clostridioides Difficile ◽  
Binding Partners ◽  
Insight Into ◽  
Peptidoglycan Layer

Spore-forming pathogens like Clostridioides difficile depend on germination to initiate infection. Spore germination depends on the degradation of the protective spore peptidoglycan layer known as the spore cortex. Cortex degradation is mediated by enzymes that recognize the spore-specific peptidoglycan modification, muramic-∂-lactam (MAL). In C. difficile, MAL synthesis depends on the activity of the CwlD amidase and the GerS lipoprotein, which directly binds CwlD. To gain insight into how GerS regulates CwlD activity, we solved the crystal structure of the CwlD:GerS complex. In this structure, a GerS homodimer is bound to two CwlD monomers such that the CwlD active sites are exposed. Although CwlD structurally resembles amidase_3 family members, we found that CwlD does not bind zinc stably on its own, unlike previously characterized amidase_3 enzymes. Instead, GerS binding to CwlD promotes CwlD binding to zinc, which is required for its catalytic mechanism. Thus, in determining the first structure of an amidase bound to its regulator, we reveal stabilization of zinc co-factor binding as a novel mechanism for regulating bacterial amidase activity. Our results further suggest that allosteric regulation by binding partners may be a more widespread mode for regulating bacterial amidase activity than previously thought.

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