scholarly journals The P4-ATPase Drs2 regulates homeostasis of Atg9

2020 ◽  
Author(s):  
Irene Pazos ◽  
Marta Puig-Tintó ◽  
Jorge Cordero ◽  
Nereida Jiménez-Menéndez ◽  
Marc Abella ◽  
...  
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Protein Complexes ◽  
Transmembrane Protein ◽  
Transport Pathways ◽  
Vesicle Tethering ◽  
Selective Autophagy ◽  
Lipid Transporters ◽  
Cellular Components ◽  
Lipid Flippase

AbstractAtg9 is a transmembrane protein essential for selective autophagy, a pathway that mediates the targeted degradation of cellular components to sustain the cell fitness. To preserve the functionality of this pathway, the cell adjusts the transport of vesicles loaded with Atg9 through mechanisms that are not understood. Here we used live-cell imaging to investigate the interactome that regulates Multisubunit Tethering Complexes (MTCs), a set of conserved protein complexes that control vesicle tethering. We found that P4-ATPases, a family of lipid transporters involved in the biogenesis of vesicles, interact with MTCs that participate in the transport of Atg9, such as TRAPPIII. Using the lipid flippase Drs2, we demonstrated that the I(S/R)TTK motif nested in the N-terminal tail cavity of P4-ATPases is necessary for the interaction with MTCs and to maintain the homeostasis of Atg9. At low temperature, the cell enhances the assembly of the Drs2-TRAPPIII module and Drs2 is fundamental for the early stages of selective autophagy, a function that is independent from its activity as lipid flippase and its role in other vesicle transport pathways.

scholarly journals Shedding light on biology of bacterial cells

2016 ◽  
Vol 371 (1707) ◽  
pp. 20150499 ◽  
Author(s):  
Johannes P. Schneider ◽  
Marek Basler
Keyword(s):  
Light Microscopy ◽  
Live Cell Imaging ◽  
Spatial Organization ◽  
Cell Imaging ◽  
Live Cell ◽  
Bacterial Cells ◽  
Resolution Limit ◽  
Detection Techniques ◽  
Living Organisms ◽  
Cellular Components

To understand basic principles of living organisms one has to know many different properties of all cellular components, their mutual interactions but also their amounts and spatial organization. Live-cell imaging is one possible approach to obtain such data. To get multiple snapshots of a cellular process, the imaging approach has to be gentle enough to not disrupt basic functions of the cell but also have high temporal and spatial resolution to detect and describe the changes. Light microscopy has become a method of choice and since its early development over 300 years ago revolutionized our understanding of living organisms. As most cellular components are indistinguishable from the rest of the cellular contents, the second revolution came from a discovery of specific labelling techniques, such as fusions to fluorescent proteins that allowed specific tracking of a component of interest. Currently, several different tags can be tracked independently and this allows us to simultaneously monitor the dynamics of several cellular components and from the correlation of their dynamics to infer their respective functions. It is, therefore, not surprising that live-cell fluorescence microscopy significantly advanced our understanding of basic cellular processes. Current cameras are fast enough to detect changes with millisecond time resolution and are sensitive enough to detect even a few photons per pixel. Together with constant improvement of properties of fluorescent tags, it is now possible to track single molecules in living cells over an extended period of time with a great temporal resolution. The parallel development of new illumination and detection techniques allowed breaking the diffraction barrier and thus further pushed the resolution limit of light microscopy. In this review, we would like to cover recent advances in live-cell imaging technology relevant to bacterial cells and provide a few examples of research that has been possible due to imaging. This article is part of the themed issue ‘The new bacteriology’.

scholarly journals Direct observation of aggregate-triggered selective autophagy

2021 ◽  
Author(s):  
Anne FJ Janssen ◽  
Giel Korsten ◽  
Wilco Nijenhuis ◽  
Eugene Katrukha ◽  
Lukas Kapitein
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Relative Timing ◽  
Imaging Studies ◽  
Selective Autophagy ◽  
The Core ◽  
Spatiotemporal Regulation ◽  
Core Components ◽  
Inducible Protein

Degradation of aggregates by selective autophagy is important as damaged proteins may impose a threat to cellular homeostasis. Although the core components of the autophagy machinery are well-characterized, the spatiotemporal regulation of many selective autophagy processes, including aggrephagy, remains largely unexplored. Furthermore, because most live-cell imaging studies have so far focused on starvation-induced autophagy, little is known about the dynamics of aggrephagy. Here, we describe the development and application of the mKeima-PIM assay, which enables live-cell observation of autophagic turnover and degradation of inducible protein aggregates in conjunction with key autophagy players. This allowed us to quantify the relative timing and duration of different steps of aggrephagy and revealed the short-lived nature of the autophagosome. The assay furthermore showed the spatial distribution of omegasome formation, highlighting that autophagy initiation is directly instructed by the cargo. Moreover, we found that nascent autophagosomes mostly remain immobile until acidification occurs. Thus, our assay provides new insights into the spatiotemporal regulation and dynamics of aggrephagy.

scholarly journals Caught in the Act: Live-Cell Imaging of Plant Meiosis

2021 ◽  
Vol 12 ◽  
Author(s):  
Maria Ada Prusicki ◽  
Martina Balboni ◽  
Kostika Sofroni ◽  
Yuki Hamamura ◽  
Arp Schnittger
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Ex Vivo ◽  
Live Cell ◽  
Cellular Processes ◽  
Short Period ◽  
Molecular Forces ◽  
Cellular Components ◽  
Plant Meiosis ◽  
Imaging Conditions

Live-cell imaging is a powerful method to obtain insights into cellular processes, particularly with respect to their dynamics. This is especially true for meiosis, where chromosomes and other cellular components such as the cytoskeleton follow an elaborate choreography over a relatively short period of time. Making these dynamics visible expands understanding of the regulation of meiosis and its underlying molecular forces. However, the analysis of meiosis by live-cell imaging is challenging; specifically in plants, a temporally resolved understanding of chromosome segregation and recombination events is lacking. Recent advances in live-cell imaging now allow the analysis of meiotic events in plants in real time. These new microscopy methods rely on the generation of reporter lines for meiotic regulators and on the establishment of ex vivo culture and imaging conditions, which stabilize the specimen and keep it alive for several hours or even days. In this review, we combine an overview of the technical aspects of live-cell imaging in plants with a summary of outstanding questions that can now be addressed to promote live-cell imaging in Arabidopsis and other plant species and stimulate ideas on the topics that can be addressed in the context of plant meiotic recombination.

scholarly journals Archaeal imaging: leading the hunt for new discoveries

2018 ◽  
Vol 29 (14) ◽  
pp. 1675-1681 ◽  
Author(s):  
Alexandre W. Bisson-Filho ◽  
Jenny Zheng ◽  
Ethan Garner
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Structural Information ◽  
Eukaryotic Cell ◽  
Live Cell ◽  
Biological Processes ◽  
Cellular Processes ◽  
Archaeal Species ◽  
Eukaryotic Organisms ◽  
Cellular Components

Since the identification of the archaeal domain in the mid-1970s, we have collected a great deal of metagenomic, biochemical, and structural information from archaeal species. However, there is still little known about how archaeal cells organize their internal cellular components in space and time. In contrast, live-cell imaging has allowed bacterial and eukaryotic cell biologists to learn a lot about biological processes by observing the motions of cells, the dynamics of their internal organelles, and even the motions of single molecules. The explosion of knowledge gained via live-cell imaging in prokaryotes and eukaryotes has motivated an ever-improving set of imaging technologies that could allow analogous explorations into archaeal biology. Furthermore, previous studies of essential biological processes in prokaryotic and eukaryotic organisms give methodological roadmaps for the investigation of similar processes in archaea. In this perspective, we highlight a few fundamental cellular processes in archaea, reviewing our current state of understanding about each, and compare how imaging approaches helped to advance the study of similar processes in bacteria and eukaryotes.

scholarly journals Plant immune and growth receptors share common signalling components but localise to distinct plasma membrane nanodomains

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Christoph A Bücherl ◽  
Iris K Jarsch ◽  
Christian Schudoma ◽  
Cécile Segonzac ◽  
Malick Mbengue ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Protein Complexes ◽  
Spatial Separation ◽  
Single Particle Tracking ◽  
Surface Receptors ◽  
Receptor Complexes ◽  
Receptor Kinases ◽  
Multicellular Organisms

Cell surface receptors govern a multitude of signalling pathways in multicellular organisms. In plants, prominent examples are the receptor kinases FLS2 and BRI1, which activate immunity and steroid-mediated growth, respectively. Intriguingly, despite inducing distinct signalling outputs, both receptors employ common downstream signalling components, which exist in plasma membrane (PM)-localised protein complexes. An important question is thus how these receptor complexes maintain signalling specificity. Live-cell imaging revealed that FLS2 and BRI1 form PM nanoclusters. Using single-particle tracking we could discriminate both cluster populations and we observed spatiotemporal separation between immune and growth signalling platforms. This finding was confirmed by visualising FLS2 and BRI1 within distinct PM nanodomains marked by specific remorin proteins and differential co-localisation with the cytoskeleton. Our results thus suggest that signalling specificity between these pathways may be explained by the spatial separation of FLS2 and BRI1 with their associated signalling components within dedicated PM nanodomains.

scholarly journals Direct observation of aggregate-triggered selective autophagy

2021 ◽  
Author(s):  
Anne F.J. Janssen ◽  
Giel Korsten ◽  
Wilco Nijenhuis ◽  
Eugene A. Katrukha ◽  
Lukas C. Kapitein
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Relative Timing ◽  
Imaging Studies ◽  
Selective Autophagy ◽  
The Core ◽  
Spatiotemporal Regulation ◽  
Core Components ◽  
Inducible Protein

Degradation of aggregates by selective autophagy is important as damaged proteins may impose a threat to cellular homeostasis. Although the core components of the autophagy machinery are well-characterized, the spatiotemporal regulation of many selective autophagy processes, including aggrephagy, remains largely unexplored. Furthermore, because most live-cell imaging studies have so far focused on starvation-induced autophagy, little is known about the dynamics of aggrephagy. Here, we describe the development and application of the mKeima-PIM assay, which enables live-cell observation of autophagic turnover and degradation of inducible protein aggregates in conjunction with key autophagy players. This allowed us to quantify the relative timing and duration of different steps of aggrephagy and revealed the short-lived nature of the autophagosome. The assay furthermore showed the spatial distribution of omegasome formation, highlighting that autophagy initiation is directly instructed by the cargo. Moreover, we found that nascent autophagosomes mostly remain immobile until acidification occurs. Thus, our assay provides new insights into the spatiotemporal regulation and dynamics of aggrephagy.

scholarly journals Arabidopsis cyclophilins direct plasmodesmata-targeting of mobile mRNA via organelle hitchhiking

2022 ◽  
Author(s):  
Kai-Ren Luo ◽  
Nien-Chen Huang ◽  
Yu-Hsin Chang ◽  
Tien-Shin Yu
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Rna Binding ◽  
Spatial Information ◽  
Rna Binding Proteins ◽  
Protein Complexes ◽  
Intercellular Transport ◽  
Cytoskeleton Organization ◽  
Organelle Trafficking ◽  
Rna Protein Complexes

Abstract Plants selectively transport mobile mRNAs through intercellular pores, plasmodesmata (PD), to distribute spatial information for synchronizing meristematic differentiation with environmental dynamics. However, how plants recognize and deliver mobile mRNAs to PD remains unknown. Here, by using RNA-live cell imaging, we show that mobile mRNAs hitchhike on organelle trafficking to transport to PD. Perturbed cytoskeleton organization or organelle trafficking severely disrupts the subcellular distribution of mobile mRNAs. We further show that Arabidopsis rotamase cyclophilins (ROCs), which are organelle-localized RNA-binding proteins (RBPs), specifically bind mobile mRNAs on the surface of organelles to direct PD-targeting. Arabidopsis roc quadruple mutants showed reduced in PD-targeting of mobile mRNAs, along with phenotype alterations. ROCs can move intercellularly and form RNA-protein complexes in phloem, suggesting the roles of ROCs in delivery of mobile mRNAs through PD. Our results highlight that an RBP-mediated hitchhiking system is purposely recruited to orient plant-mobile mRNAs to PD for intercellular transport.

Sign in / Sign up

Export Citation Format

Share Document