bims-nucpor 2022-09-11 papers

Issue of 2022–09–11
four papers selected by
Sara Mingu, Johannes Gutenberg University

Neurotherapeutics. 2022 Sep 07.

Nuclear Pore Dysfunction in Neurodegeneration.

Olivia Spead, Benjamin L Zaepfel, Jeffrey D Rothstein.

The nuclear pore complex (NPC) is a large multimeric structure that is interspersed throughout the membrane of the nucleus and consists of at least 33 protein components. Individual components cooperate within the nuclear pore to facilitate selective passage of materials between the nucleus and cytoplasm while simultaneously performing pore-independent roles throughout the cell. NPC dysfunction is a hallmark of neurodegenerative disorders including Alzheimer's disease, Huntington's disease, and amyotrophic lateral sclerosis (ALS). NPC components can become mislocalized or altered in expression in neurodegeneration. These alterations in NPC structure are often detrimental to the neuronal function and ultimately lead to neuronal loss. This review highlights the importance of nucleocytoplasmic transport and NPC integrity and how dysfunction of such may contribute to neurodegeneration.

Keywords: ALS; Alzheimer’s disease; Huntington’s disease; Neurodegeneration; Nuclear pore complex; Nucleocytoplasmic transport

DOI: https://doi.org/10.1007/s13311-022-01293-w

Curr Protoc. 2022 Sep;2(9): e520

Selective Degradation and Quantification of Nucleoporins in the Nuclear Pore by Auxin-Inducible Degrons and Single-Molecule Microscopy.

Mark Tingey, Yichen Li, Weidong Yang.

There is a significant current question regarding the viable copy numbers of nucleoporins required for the function of the nuclear pore complex (NPC) in eukaryotic cells. The NPC consists of approximately 30 different nucleoporins in an eight-fold symmetry, meaning that there are multiple duplicates of each nucleoporin present within the nuclear pore. We recently developed a method that combines auxin-inducible degrons and single-molecule super-resolution microscopy to evaluate the copy number of nuclear basket nucleoporins required for the successful function of the NPC. Here, we describe the theory behind this auxin-inducible degron and single-molecule super-resolution microscopy method, and we detail a step-by-step process to selectively degrade nucleoporins either completely or in a stepwise manner. © 2022 Wiley Periodicals LLC. Basic Protocol 1: Degradation of target nucleoporins Basic Protocol 2: Quantification of nucleoporin copy number via narrow-field fluorescence microscopy.

Keywords: auxin-inducible degron; nuclear basket; nuclear pore complex; nucleoporin; nucleoporin copy number; single-point edge-excitation subdiffraction (SPEED) microscopy

DOI: https://doi.org/10.1002/cpz1.520

PLoS Pathog. 2022 Sep 06. 18(9): e1010824

Components of the LINC and NPC complexes coordinately target and translocate a virus into the nucleus to promote infection.

Chelsey C Spriggs, Grace Cha, Jiaqian Li, Billy Tsai.

Nuclear entry represents the final and decisive infection step for most DNA viruses, although how this is accomplished by some viruses is unclear. Polyomavirus SV40 transports from the cell surface through the endosome, the endoplasmic reticulum, and the cytosol from where it enters the nucleus to cause infection. Here we elucidate the nuclear entry mechanism of SV40. Our results show that cytosol-localized SV40 is targeted to the nuclear envelope by directly engaging Nesprin-2 of the linker of nucleoskeleton and cytoskeleton (LINC) nuclear membrane complex. Additionally, we identify the NUP188 subunit of the nuclear pore complex (NPC) as a new Nesprin-2-interacting partner. This physical proximity positions the NPC to capture SV40 upon release from Nesprin-2, enabling the channel to facilitate nuclear translocation of the virus. Strikingly, SV40 disassembles during nuclear entry, generating a viral genome-VP1-VP3 subcomplex that efficiently crosses the NPC to enter the nucleus. Our results reveal how two major nuclear membrane protein complexes are exploited to promote targeting and translocation of a virus into the nucleus.

DOI: https://doi.org/10.1371/journal.ppat.1010824

Front Mol Biosci. 2022 ;9 963635

Parallel import mechanisms ensure the robust nuclear localization of actin in Drosophila.

Péter Borkúti, Ildikó Kristó, Anikó Szabó, Csaba Bajusz, Zoltán Kovács, Zsuzsánna Réthi-Nagy, Zoltán Lipinszki, Tamás Lukácsovich, Sven Bogdan, Péter Vilmos.

Actin, as an ancient and fundamental protein, participates in various cytoplasmic as well as nuclear functions in eukaryotic cells. Based on its manifold tasks in the nucleus, it is a reasonable assumption that the nuclear presence of actin is essential for the cell, and consequently, its nuclear localization is ensured by a robust system. However, today only a single nuclear import and a single nuclear export pathway is known which maintain the dynamic balance between cytoplasmic and nuclear actin pools. In our work, we tested the robustness of the nuclear import of actin, and investigated whether the perturbations of nuclear localization affect the viability of the whole organism. For this aim, we generated a genetic system in Drosophila, in which we rescued the lethal phenotype of the null mutation of the Actin5C gene with transgenes that express different derivatives of actin, including a Nuclear Export Signal (NES)-tagged isoform which ensures forced nuclear export of the protein. We also disrupted the SUMOylation site of actin, suggested earlier to be responsible for nuclear retention, and eliminated the activity of the single nuclear import factor dedicated to actin. We found that, individually, none of the above mentioned manipulations led to a notable reduction in nuclear actin levels and thus, fully rescued lethality. However, the NES tagging of actin, together with the knock out of its importin, significantly reduced the amount of nuclear actin and induced lethality, confirming that the presence of actin in the nucleus is essential, and thereby, over-secured. Supporting this, we identified novel nuclear importins specific to actin, which sheds light on the mechanism behind the robustness of nuclear localization of actin, and supports the idea of essentiality of its nuclear functions.

Keywords: Drosophila; actin; importin; nuclear transport; nucleus

DOI: https://doi.org/10.3389/fmolb.2022.963635