Cellular Organisation: Putting organelles in their place

Experiments in C. elegans reveal new insights into how the ANC-1 protein helps to anchor the nucleus and other organelles in place.
  1. Patricia Ulm
  2. Verena Jantsch  Is a corresponding author
  1. Department of Chromosome Biology, Max Perutz Laboratories, University of Vienna, Vienna Biocenter, Austria

Cells contain an assortment of organelles which each have their own specialized role. To work correctly, most organelles need to be properly positioned within the cell. For example, mis-localization of the cell’s largest organelle, the nucleus, has been observed in neuromuscular diseases, such as Emery-Dreyfuss muscular dystrophy (Luxton and Starr, 2014).

Current models suggest that positioning of the nucleus relies on a complex called LINC (short for Linker of Nucleoskeleton and Cytoskeleton), which is made up of proteins that contain either a SUN or KASH domain. The SUN proteins (SAD1, UNC-84) sit across the inner nuclear membrane and connect to structures in the nucleus, such as chromatin and the nuclear lamina, and the KASH proteins (Klarsicht, ANC-1, Syne Homology) span across the outer nuclear membrane and interact with proteins in the cytoskeleton. The SUN and KASH domains of these proteins join together to form a bridge that mechanically couples the nucleus and cytoskeleton, which helps to anchor the nucleus in the right place (Kim et al., 2015).

This model of how nuclear positioning works is primarily based on experiments in Caenorhabditis elegans worms with mutations in the genes for either the UNC-84 or ANC-1 protein (Starr and Fridolfsson, 2010). The hypodermis of adult wild-type worms is made up of several huge hyp-7 cells (or syncytia) which each contain 139 evenly spaced nuclei (Shemer and Podbilewicz, 2000), making them a useful system for investigating nuclear anchorage. According to the model, if the nuclei in hyp-7 cells are exclusively anchored via the SUN–KASH bridge, then loss of the genes for UNC-84 or ANC-1 should have an identical effect and result in the same amount of nuclear clustering. However, in 2018, a group of researchers made a puzzling discovery: they found that deleting the gene for ANC-1 resulted in more severe nuclear clustering than removing the gene for UNC-84 (Cain et al., 2018). Now, in eLife, Daniel A Starr and co-workers from University California, Davis – including Hongyan Hao as first author – report that the model for how the nucleus is positioned may need re-defining (Hao et al., 2021).

The team (which includes some of the authors involved in the 2018 study) found, as expected, that removing the gene for ANC-1 led to nuclear clustering in hyp-7 cells, indicating that nuclear anchorage had been lost (Figure 1). The untethered nuclei also disrupted the network of microtubules in the cytoskeleton, and appeared much smaller and less rounded, suggesting that the cells lacked mechanical stability. In C. elegans, the ANC-1 protein contains multiple domains: an actin-binding domain at its N-terminus, several cytoplasmic domains that likely bind to other proteins, a transmembrane domain, and a KASH domain at its C-terminus (Gundersen and Worman, 2013; Starr and Han, 2002). To gain a better understanding of how ANC-1 positions the nucleus, Hao et al. deleted these different domains, either separately or in combination, to see how this affected the protein’s role in the cell.

Loss of ANC-1 leads to unanchored and misshaped organelles and a smaller body size in worms.

The hypodermis of C. elegans worms (top schematic) is made up of hyp-7 cells which contain over a hundred nuclei (represented as black dots). In wild-type worms (left), the KASH protein ANC-1 (depicted as spikes) localizes to the membrane of the nucleus and endoplasmic reticulum (ER). As a result, the nuclei (purple) are spherical and evenly spaced, and the ER (blue), mitochondria (orange) and lipid droplets (yellow) are well anchored. The microtubule network (black lines) is also evenly distributed throughout the cytoplasm. Meanwhile, in mutant worms lacking the gene for ANC-1 (right), the ER and mitochondria are fragmented, and the nuclei are unanchored and clustered together. Lipid droplets are also clustered and the microtubule network is disrupted by the movement of the untethered organelles. This causes the mutant worm to have a smaller body size and the nuclei in its hyp-7 cells to be mispositioned.

Image credit: Patricia Ulm.

Deletion of the KASH domain only caused moderate nuclear clustering, and loss of the actin-binding domain did not generate any nuclear anchorage defects. In contrast, deleting parts of ANC-1 protein that sit within the cytoplasm and likely bind to other cytoskeleton proteins led to more severe positioning defects. Notably, nuclear mispositioning was greatly increased in double mutants lacking both a functional SUN domain in UNC-84 and a cytoplasmic domain of ANC-1. This synergistic effect suggests that nuclear positioning is likely controlled by cooperation between two distinct domains of ANC-1: the cytoplasmic domain and the domain that binds to UNC-84 in the SUN-KASH bridge.

Previous studies have shown that ANC-1 also controls the distribution of mitochondria (Starr and Han, 2002). Therefore, Hao et al. investigated whether ANC-1 is also essential for positioning the mitochondria and another organelle called the endoplasmic reticulum (or ER for short). In mutant worms lacking the gene for ANC-1, they found unanchored fragments of mitochondria and the ER along with clusters of lipid droplets (Figure 1). As seen for the nucleus, deleting the KASH domain only resulted in mild defects in ER positioning, suggesting that localization of the ER also mainly relies on parts of the ANC-1 protein outside the KASH domain.

These findings suggest that ANC-1 positions the nucleus and ER largely independently from the KASH domain. Further experiments revealed that the ANC-1 protein is also located on the membrane of the ER. This led Hao et al. to propose a new cytoplasmic integrity model in which ANC-1 localizes to both the ER and nuclear membranes, and reaches out to correctly position organelles via an interconnecting network that permeates the entire cytoplasm.

This study raises the question of how ANC-1 can position and hold organelles in place with little or no help from its KASH and actin-binding domains. Notably, studies in mice have also shown that the actin-binding domains of Nesprin1 (mouse ANC-1) are not essential to anchor the nucleus within the cytoplasm (Stroud et al., 2017). Therefore, some important questions remain: which cytoskeletal component(s) work with ANC-1 to correctly position organelles and confer mechanical stability to the cell? Are the neuromuscular diseases associated with mutations in the LINC complex the result of alterations in cytoplasmic integrity rather than nuclear anchorage defects? Further experiments using C. elegans as a model system may help to answer these questions and shed further light on how ANC-1 anchors organelles in place.

References

Article and author information

Author details

  1. Patricia Ulm

    Patricia Ulm is in the Department of Chromosome Biology, Max Perutz Laboratories, University of Vienna, Vienna Biocenter, Vienna, Austria

    Competing interests
    No competing interests declared
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8876-2576
  2. Verena Jantsch

    Verena Jantsch is in the Department of Chromosome Biology, Max Perutz Laboratories, University of Vienna, Vienna Biocenter, Vienna, Austria

    For correspondence
    verena.jantsch@univie.ac.at
    Competing interests
    No competing interests declared
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-1978-682X

Publication history

  1. Version of Record published:

Copyright

© 2021, Ulm and Jantsch

This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 1,479
    views
  • 116
    downloads
  • 1
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Patricia Ulm
  2. Verena Jantsch
(2021)
Cellular Organisation: Putting organelles in their place
eLife 10:e69422.
https://doi.org/10.7554/eLife.69422

Further reading

    1. Cell Biology
    Tamás Visnovitz, Dorina Lenzinger ... Edit I Buzas
    Short Report

    Recent studies showed an unexpected complexity of extracellular vesicle (EV) biogenesis pathways. We previously found evidence that human colorectal cancer cells in vivo release large multivesicular body-like structures en bloc. Here, we tested whether this large EV type is unique to colorectal cancer cells. We found that all cell types we studied (including different cell lines and cells in their original tissue environment) released multivesicular large EVs (MV-lEVs). We also demonstrated that upon spontaneous rupture of the limiting membrane of the MV-lEVs, their intraluminal vesicles (ILVs) escaped to the extracellular environment by a ‘torn bag mechanism’. We proved that the MV-lEVs were released by ectocytosis of amphisomes (hence, we termed them amphiectosomes). Both ILVs of amphiectosomes and small EVs separated from conditioned media were either exclusively CD63 or LC3B positive. According to our model, upon fusion of multivesicular bodies with autophagosomes, fragments of the autophagosomal inner membrane curl up to form LC3B positive ILVs of amphisomes, while CD63 positive small EVs are of multivesicular body origin. Our data suggest a novel common release mechanism for small EVs, distinct from the exocytosis of multivesicular bodies or amphisomes, as well as the small ectosome release pathway.

    1. Cell Biology
    Yajun Zhai, Peiyi Liu ... Gongzheng Hu
    Research Article

    Discovering new strategies to combat the multidrug-resistant bacteria constitutes a major medical challenge of our time. Previously, artesunate (AS) has been reported to exert antibacterial enhancement activity in combination with β-lactam antibiotics via inhibition of the efflux pump AcrB. However, combination of AS and colistin (COL) revealed a weak synergistic effect against a limited number of strains, and few studies have further explored its possible mechanism of synergistic action. In this article, we found that AS and EDTA could strikingly enhance the antibacterial effects of COL against mcr-1- and mcr-1+ Salmonella strains either in vitro or in vivo, when used in triple combination. The excellent bacteriostatic effect was primarily related to the increased cell membrane damage, accumulation of toxic compounds and inhibition of MCR-1. The potential binding sites of AS to MCR-1 (THR283, SER284, and TYR287) were critical for its inhibition of MCR-1 activity. Additionally, we also demonstrated that the CheA of chemosensory system and virulence-related protein SpvD were critical for the bacteriostatic synergistic effects of the triple combination. Selectively targeting CheA, SpvD, or MCR using the natural compound AS could be further investigated as an attractive strategy for the treatment of Salmonella infection. Collectively, our work opens new avenues toward the potentiation of COL and reveals an alternative drug combination strategy to overcome COL-resistant bacterial infections.