Dissecting the Structural Dynamics of the Nuclear Pore Complex

Hakhverdyan, Zhanna; Molloy, Kelly R.; Keegan, Sarah; Herricks, Thurston; Lepore, Dante; Munson, Mary; Subbotin, Roman I.; Fenyö, David; Aitchison, John D.; Fernández-Martı́nez, Javier; Chait, Brian T.; Rout, Michael P.

doi:10.1016/j.molcel.2020.11.032

Cited by 35 publications

(66 citation statements)

References 84 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“… Timing: [1 week] Prepare synthetic defined agar plates lacking Histidine (SD-His) from SC-His (synthetic complete lacking Histidine) amino acid mixture following manufacturer's instructions (Sunrize Sciences). PCR amplify the N-terminal tagging cassette from Ttc3GFP-His5 plasmid with 45–60 bp overhangs homologous to the insertion locus (i.e., right at the ATG start codon), using standard approaches and as described ( Hakhverdyan et al., 2021 ). …”

Section: Step-by-step Methods Detailsmentioning

confidence: 99%

“…PCR amplify the N-terminal tagging cassette from Ttc3GFP-His5 plasmid with 45–60 bp overhangs homologous to the insertion locus (i.e., right at the ATG start codon), using standard approaches and as described ( Hakhverdyan et al., 2021 ).…”

Section: Step-by-step Methods Detailsmentioning

confidence: 99%

“…Ttc3GFP-His5 contains the pTDH3 yeast promoter, 3 tetracycline-binding aptamers in the 5′-UTR and GFP - all parts necessary for conditional expression of the tagged gene of interest in the absence of tetracycline/ chlortetracycline (ClTc) and repression in the presence of the antibiotic ( Hakhverdyan et al., 2021 ; Kotter et al., 2009 ; Suess et al., 2012 ). Use the standard S. cerevisiae lithium acetate transformation procedure with the diploid strain of interest.…”

Section: Step-by-step Methods Detailsmentioning

confidence: 99%

“…This method can determine and distinguish both subunit turnover and exchange specifically in an assembly to provide a comprehensive picture of assembly dynamics. For complete details on the use and execution of this protocol, please refer to Hakhverdyan et al. (2021) .…”

mentioning

confidence: 99%

“…For complete details on the use and execution of this protocol, please refer to Hakhverdyan et al. (2021) .…”

mentioning

confidence: 99%

See 4 more Smart Citations

Measuring in vivo protein turnover and exchange in yeast macromolecular assemblies

et al. 2021

Self Cite

View full text Add to dashboard Cite

Summary We present a comprehensive and robust protocol to track the dynamics of all proteins in a complex in yeast cells. A single member of the protein assembly is tagged and conditionally expressed, minimizing the perturbations to the protein complex. Then, SILAC labeling and affinity purification are used for the assessment of the whole protein complex dynamics. This method can determine and distinguish both subunit turnover and exchange specifically in an assembly to provide a comprehensive picture of assembly dynamics. For complete details on the use and execution of this protocol, please refer to Hakhverdyan et al. (2021) .

show abstract

Section: Step-by-step Methods Detailsmentioning

confidence: 99%

Section: Step-by-step Methods Detailsmentioning

confidence: 99%

Section: Step-by-step Methods Detailsmentioning

confidence: 99%

mentioning

confidence: 99%

“…For complete details on the use and execution of this protocol, please refer to Hakhverdyan et al. (2021) .…”

mentioning

confidence: 99%

See 3 more Smart Citations

Measuring in vivo protein turnover and exchange in yeast macromolecular assemblies

et al. 2021

Self Cite

View full text Add to dashboard Cite

show abstract

Puzzling out nuclear pore complex assembly

Penzo,

Palancade

2023

FEBS Letters

View full text Add to dashboard Cite

Nuclear pore complexes (NPCs) are sophisticated multiprotein assemblies embedded within the nuclear envelope and controlling the exchanges of molecules between the cytoplasm and the nucleus. In this review, we summarize the mechanisms by which these elaborate complexes are built from their subunits, the nucleoporins, based on our ever‐growing knowledge of NPC structural organization and on the recent identification of additional features of this process. We present the constraints faced during the production of nucleoporins, their gathering into oligomeric complexes, and the formation of NPCs within nuclear envelopes, and review the cellular strategies at play, from co‐translational assembly to the enrolment of a panel of cofactors. Remarkably, the study of NPCs can inform our perception of the biogenesis of multiprotein complexes in general – and vice versa.

show abstract

Coordinating nucleoporin condensation and nuclear pore complex assembly

Kuiper,

Prophet,

Schlieker

2023

FEBS Letters

View full text Add to dashboard Cite

The nuclear pore complex (NPC) is among the most elaborate protein complexes in eukaryotes. While ribosomes and proteasomes are known to require dedicated assembly machinery, our understanding of NPC assembly is at a relatively early stage. Defects in NPC assembly or homeostasis are tied to movement disorders, including dystonia and amyotrophic lateral sclerosis (ALS), as well as aging, requiring a better understanding of these processes to enable therapeutic intervention. Here, we discuss recent progress in the understanding of NPC assembly and highlight how related defects in human disorders can shed light on NPC biogenesis. We propose that the condensation of phenylalanine‐glycine repeat nucleoporins needs to be carefully controlled during NPC assembly to prevent aberrant condensation, aggregation, or amyloid formation.

show abstract

Dissecting the Structural Dynamics of the Nuclear Pore Complex

Cited by 35 publications

References 84 publications

Measuring in vivo protein turnover and exchange in yeast macromolecular assemblies

Measuring in vivo protein turnover and exchange in yeast macromolecular assemblies

Puzzling out nuclear pore complex assembly

Coordinating nucleoporin condensation and nuclear pore complex assembly

Contact Info

Product

Resources

About