Curated Optogenetic Publication Database

Search precisely and efficiently by using the advantage of the hand-assigned publication tags that allow you to search for papers involving a specific trait, e.g. a particular optogenetic switch or a host organism.

Showing 1 - 19 of 19 results

Automatic detection of spatio-temporal signalling patterns in cell collectives.

blue CRY2/CIB1 MCF10A Signaling cascade control
J Cell Biol, 27 Jul 2023 DOI: 10.1083/jcb.202207048 Link to full text
Abstract: An increasing experimental evidence points to physiological importance of space-time correlations in signalling of cell collectives. From wound healing to epithelial homeostasis to morphogenesis, coordinated activation of bio-molecules between cells allows the collectives to perform more complex tasks and better tackle environmental challenges. To understand this information exchange and to advance new theories of emergent phenomena, we created ARCOS, a computational method to detect and quantify collective signalling. We demonstrate ARCOS on cell and organism collectives with space-time correlations on different scales in 2D and 3D. We make a new observation that oncogenic mutations in the MAPK/ERK and PIK3CA/Akt pathways of MCF10A epithelial cells induce ERK activity waves with different size, duration, and frequency. The open-source implementations of ARCOS are available as R and Python packages, and as a plugin for napari image viewer to interactively quantify collective phenomena without prior programming experience.

Integrin-based adhesion compartmentalizes ALK3 of the BMPRII to control cell adhesion and migration.

blue iLID C2C12 EpH4 REF52 SYF Control of cytoskeleton / cell motility / cell shape Transgene expression
J Cell Biol, 7 Oct 2022 DOI: 10.1083/jcb.202107110 Link to full text
Abstract: The spatial organization of cell-surface receptors is fundamental for the coordination of biological responses to physical and biochemical cues of the extracellular matrix. How serine/threonine kinase receptors, ALK3-BMPRII, cooperate with integrins upon BMP2 to drive cell migration is unknown. Whether the dynamics between integrins and BMP receptors intertwine in space and time to guide adhesive processes is yet to be elucidated. We found that BMP2 stimulation controls the spatial organization of BMPRs by segregating ALK3 from BMPRII into β3 integrin-containing focal adhesions. The selective recruitment of ALK3 to focal adhesions requires β3 integrin engagement and ALK3 activation. BMP2 controls the partitioning of immobilized ALK3 within and outside focal adhesions according to single-protein tracking and super-resolution imaging. The spatial control of ALK3 in focal adhesions by optogenetics indicates that ALK3 acts as an adhesive receptor by eliciting cell spreading required for cell migration. ALK3 segregation from BMPRII in integrin-based adhesions is a key aspect of the spatio-temporal control of BMPR signaling.

Killing cells using light (activated) sabers.

blue Cryptochromes Review
J Cell Biol, 16 May 2022 DOI: 10.1083/jcb.202205018 Link to full text
Abstract: Many types of regulated cell death exist, however the non-cell autonomous effects of specific forms of cell death remain poorly understood. Addressing this, Shkarina et al. (2022. J. Cell Biol. describe an optogenetic method to activate distinct modes of cell death in select cells.

Optogenetic activators of apoptosis, necroptosis, and pyroptosis.

blue CRY2olig Caco-2 HaCaT HEK293T HeLa MCF7 RAW264.7 zebrafish in vivo Cell death
J Cell Biol, 14 Apr 2022 DOI: 10.1083/jcb.202109038 Link to full text
Abstract: Targeted and specific induction of cell death in an individual or groups of cells hold the potential for new insights into the response of tissues or organisms to different forms of death. Here, we report the development of optogenetically controlled cell death effectors (optoCDEs), a novel class of optogenetic tools that enables light-mediated induction of three types of programmed cell death (PCD)—apoptosis, pyroptosis, and necroptosis—using Arabidopsis thaliana photosensitive protein Cryptochrome-2. OptoCDEs enable a rapid and highly specific induction of PCD in human, mouse, and zebrafish cells and are suitable for a wide range of applications, such as sub-lethal cell death induction or precise elimination of single cells or cell populations in vitro and in vivo. As the proof-of-concept, we utilize optoCDEs to assess the differences in neighboring cell responses to apoptotic or necrotic PCD, revealing a new role for shingosine-1-phosphate signaling in regulating the efferocytosis of the apoptotic cell by epithelia.

Mechanical competition alters the cellular interpretation of an endogenous genetic programme.

blue CRY2/CIB1 D. melanogaster in vivo Developmental processes
J Cell Biol, 27 Aug 2021 DOI: 10.1083/jcb.202104107 Link to full text
Abstract: The intrinsic genetic programme of a cell is not sufficient to explain all of the cell’s activities. External mechanical stimuli are increasingly recognized as determinants of cell behaviour. In the epithelial folding event that constitutes the beginning of gastrulation in Drosophila, the genetic programme of the future mesoderm leads to the establishment of a contractile actomyosin network that triggers apical constriction of cells, and thereby, furrow formation. However, some cells do not constrict but instead stretch, even though they share the same genetic programme as their constricting neighbours. We show here that tissue-wide interactions force these cells to expand even when an otherwise sufficient amount of apical, active actomyosin is present. Models based on contractile forces and linear stress-strain responses are not sufficient to reproduce experimental observations, but simulations in which cells behave as ductile materials with non-linear mechanical properties do. Our models show that this behaviour is a general emergent property of supracellular actomyosin networks, in accordance with our experimental observations of actin reorganisation within stretching cells.

O-GlcNAc modification of nuclear pore complexes accelerates bidirectional transport.

blue AsLOV2 U-2 OS
J Cell Biol, 5 Jul 2021 DOI: 10.1083/jcb.202010141 Link to full text
Abstract: Macromolecular transport across the nuclear envelope depends on facilitated diffusion through nuclear pore complexes (NPCs). The interior of NPCs contains a permeability barrier made of phenylalanine-glycine (FG) repeat domains that selectively facilitates the permeation of cargoes bound to nuclear transport receptors (NTRs). FG-repeat domains in NPCs are a major site of O-linked N-acetylglucosamine (O-GlcNAc) modification, but the functional role of this modification in nucleocytoplasmic transport is unclear. We developed high-throughput assays based on optogenetic probes to quantify the kinetics of nuclear import and export in living human cells. We found that increasing O-GlcNAc modification of the NPC accelerated NTR-facilitated transport of proteins in both directions, and decreasing modification slowed transport. Superresolution imaging revealed strong enrichment of O-GlcNAc at the FG-repeat barrier. O-GlcNAc modification also accelerated passive permeation of a small, inert protein through NPCs. We conclude that O-GlcNAc modification accelerates nucleocytoplasmic transport by enhancing the nonspecific permeability of the FG-repeat barrier, perhaps by steric inhibition of interactions between FG repeats.

What sugar does to your pores.

blue LOV domains Review
J Cell Biol, 16 Jun 2021 DOI: 10.1083/jcb.202105163 Link to full text
Abstract: FG-repeat nucleoporins at the center of the nuclear pore complex (NPC) are highly modified with O-GlcNAc. In this issue, Yoo and Mitchison (2021. J. Cell Biol. use optogenetic probes to show that O-GlcNAc enhances permeability of the NPC, accelerating transport in both directions.

The mitotic protein NuMA plays a spindle-independent role in nuclear formation and mechanics.

J Cell Biol, 7 Dec 2020 DOI: 10.1083/jcb.202004202 Link to full text
Abstract: Eukaryotic cells typically form a single, round nucleus after mitosis, and failures to do so can compromise genomic integrity. How mammalian cells form such a nucleus remains incompletely understood. NuMA is a spindle protein whose disruption results in nuclear fragmentation. What role NuMA plays in nuclear integrity, and whether its perceived role stems from its spindle function, are unclear. Here, we use live imaging to demonstrate that NuMA plays a spindle-independent role in forming a single, round nucleus. NuMA keeps the decondensing chromosome mass compact at mitotic exit and promotes a mechanically robust nucleus. NuMA's C terminus binds DNA in vitro and chromosomes in interphase, while its coiled-coil acts as a central regulatory and structural element: it prevents NuMA from binding chromosomes at mitosis, regulates its nuclear mobility, and is essential for nuclear formation. Thus, NuMA plays a structural role over the cell cycle, building and maintaining the spindle and nucleus, two of the cell's largest structures.

The proline-rich domain promotes Tau liquid-liquid phase separation in cells.

blue CRY2olig SH-SY5Y Control of cytoskeleton / cell motility / cell shape Organelle manipulation
J Cell Biol, 2 Nov 2020 DOI: 10.1083/jcb.202006054 Link to full text
Abstract: Tau protein in vitro can undergo liquid-liquid phase separation (LLPS); however, observations of this phase transition in living cells are limited. To investigate protein state transitions in living cells, we attached Cry2 to Tau and studied the contribution of each domain that drives the Tau cluster in living cells. Surprisingly, the proline-rich domain (PRD), not the microtubule binding domain (MTBD), drives LLPS and does so under the control of its phosphorylation state. Readily observable, PRD-derived cytoplasmic condensates underwent fusion and fluorescence recovery after photobleaching consistent with the PRD LLPS in vitro. Simulations demonstrated that the charge properties of the PRD predicted phase separation. Tau PRD formed heterotypic condensates with EB1, a regulator of plus-end microtubule dynamic instability. The specific domain properties of the MTBD and PRD serve distinct but mutually complementary roles that use LLPS in a cellular context to implement emergent functionalities that scale their relationship from binding α-beta tubulin heterodimers to the larger proportions of microtubules.

An optimized toolbox for the optogenetic control of intracellular transport.

blue iLID VVD Cos-7 HeLa U-2 OS Control of cytoskeleton / cell motility / cell shape
J Cell Biol, 6 Apr 2020 DOI: 10.1083/jcb.201907149 Link to full text
Abstract: Cellular functioning relies on active transport of organelles by molecular motors. To explore how intracellular organelle distributions affect cellular functions, several optogenetic approaches enable organelle repositioning through light-inducible recruitment of motors to specific organelles. Nonetheless, robust application of these methods in cellular populations without side effects has remained challenging. Here, we introduce an improved toolbox for optogenetic control of intracellular transport that optimizes cellular responsiveness and limits adverse effects. To improve dynamic range, we employed improved optogenetic heterodimerization modules and engineered a photosensitive kinesin-3, which is activated upon blue light-sensitive homodimerization. This opto-kinesin prevented motor activation before experimental onset, limited dark-state activation, and improved responsiveness. In addition, we adopted moss kinesin-14 for efficient retrograde transport with minimal adverse effects on endogenous transport. Using this optimized toolbox, we demonstrate robust reversible repositioning of (endogenously tagged) organelles within cellular populations. More robust control over organelle motility will aid in dissecting spatial cell biology and transport-related diseases.

Spatiotemporal control of phosphatidic acid signaling with optogenetic, engineered phospholipase Ds.

blue CRY2/CIB1 HEK293T Signaling cascade control
J Cell Biol, 2 Mar 2020 DOI: 10.1083/jcb.201907013 Link to full text
Abstract: Phosphatidic acid (PA) is both a central phospholipid biosynthetic intermediate and a multifunctional lipid second messenger produced at several discrete subcellular locations. Organelle-specific PA pools are believed to play distinct physiological roles, but tools with high spatiotemporal control are lacking for unraveling these pleiotropic functions. Here, we present an approach to precisely generate PA on demand on specific organelle membranes. We exploited a microbial phospholipase D (PLD), which produces PA by phosphatidylcholine hydrolysis, and the CRY2-CIBN light-mediated heterodimerization system to create an optogenetic PLD (optoPLD). Directed evolution of PLD using yeast membrane display and IMPACT, a chemoenzymatic method for visualizing cellular PLD activity, yielded a panel of optoPLDs whose range of catalytic activities enables mimicry of endogenous, physiological PLD signaling. Finally, we applied optoPLD to elucidate that plasma membrane, but not intracellular, pools of PA can attenuate the oncogenic Hippo signaling pathway. OptoPLD represents a powerful and precise approach for revealing spatiotemporally defined physiological functions of PA.

Mps1-mediated release of Mad1 from nuclear pores ensures the fidelity of chromosome segregation.

blue CRY2/CIB1 Schneider 2 Cell cycle control
J Cell Biol, 2 Mar 2020 DOI: 10.1083/jcb.201906039 Link to full text
Abstract: The spindle assembly checkpoint (SAC) relies on the recruitment of Mad1-C-Mad2 to unattached kinetochores but also on its binding to Megator/Tpr at nuclear pore complexes (NPCs) during interphase. However, the molecular underpinnings controlling the spatiotemporal redistribution of Mad1-C-Mad2 as cells progress into mitosis remain elusive. Here, we show that activation of Mps1 during prophase triggers Mad1 release from NPCs and that this is required for kinetochore localization of Mad1-C-Mad2 and robust SAC signaling. We find that Mps1 phosphorylates Megator/Tpr to reduce its interaction with Mad1 in vitro and in Drosophila cells. Importantly, preventing Mad1 from binding to Megator/Tpr restores Mad1 accumulation at kinetochores, the fidelity of chromosome segregation, and genome stability in larval neuroblasts of mps1-null mutants. Our findings demonstrate that the subcellular localization of Mad1 is tightly coordinated with cell cycle progression by kinetochore-extrinsic activity of Mps1. This ensures that both NPCs in interphase and kinetochores in mitosis can generate anaphase inhibitors to efficiently preserve genomic stability.

Cross-linker-mediated regulation of actin network organization controls tissue morphogenesis.

blue CRY2/CIB1 D. melanogaster in vivo Control of cytoskeleton / cell motility / cell shape
J Cell Biol, 28 Jun 2019 DOI: 10.1083/jcb.201811127 Link to full text
Abstract: Contraction of cortical actomyosin networks driven by myosin activation controls cell shape changes and tissue morphogenesis during animal development. In vitro studies suggest that contractility also depends on the geometrical organization of actin filaments. Here we analyze the function of actomyosin network topology in vivo using optogenetic stimulation of myosin-II in Drosophila embryos. We show that early during cellularization, hexagonally arrayed actomyosin fibers are resilient to myosin-II activation. Actomyosin fibers then acquire a ring-like conformation and become contractile and sensitive to myosin-II. This transition is controlled by Bottleneck, a Drosophila unique protein expressed for only a short time during early cellularization, which we show regulates actin bundling. In addition, it requires two opposing actin cross-linkers, Filamin and Fimbrin. Filamin acts synergistically with Bottleneck to facilitate hexagonal patterning, while Fimbrin controls remodeling of the hexagonal network into contractile rings. Thus, actin cross-linking regulates the spatio-temporal organization of actomyosin contraction in vivo, which is critical for tissue morphogenesis.

Control of microtubule dynamics using an optogenetic microtubule plus end-F-actin cross-linker.

blue iLID Schneider 2 Control of cytoskeleton / cell motility / cell shape
J Cell Biol, 19 Dec 2017 DOI: 10.1083/jcb.201705190 Link to full text
Abstract: We developed a novel optogenetic tool, SxIP-improved light-inducible dimer (iLID), to facilitate the reversible recruitment of factors to microtubule (MT) plus ends in an end-binding protein-dependent manner using blue light. We show that SxIP-iLID can track MT plus ends and recruit tgRFP-SspB upon blue light activation. We used this system to investigate the effects of cross-linking MT plus ends and F-actin in Drosophila melanogaster S2 cells to gain insight into spectraplakin function and mechanism. We show that SxIP-iLID can be used to temporally recruit an F-actin binding domain to MT plus ends and cross-link the MT and F-actin networks. Cross-linking decreases MT growth velocities and generates a peripheral MT exclusion zone. SxIP-iLID facilitates the general recruitment of specific factors to MT plus ends with temporal control enabling researchers to systematically regulate MT plus end dynamics and probe MT plus end function in many biological processes.

A module for Rac temporal signal integration revealed with optogenetics.

red PhyB/PIF6 HL-60 Signaling cascade control Control of cytoskeleton / cell motility / cell shape
J Cell Biol, 7 Jul 2017 DOI: 10.1083/jcb.201604113 Link to full text
Abstract: Sensory systems use adaptation to measure changes in signaling inputs rather than absolute levels of signaling inputs. Adaptation enables eukaryotic cells to directionally migrate over a large dynamic range of chemoattractant. Because of complex feedback interactions and redundancy, it has been difficult to define the portion or portions of eukaryotic chemotactic signaling networks that generate adaptation and identify the regulators of this process. In this study, we use a combination of optogenetic intracellular inputs, CRISPR-based knockouts, and pharmacological perturbations to probe the basis of neutrophil adaptation. We find that persistent, optogenetically driven phosphatidylinositol (3,4,5)-trisphosphate (PIP3) production results in only transient activation of Rac, a hallmark feature of adaptive circuits. We further identify the guanine nucleotide exchange factor P-Rex1 as the primary PIP3-stimulated Rac activator, whereas actin polymerization and the GTPase-activating protein ArhGAP15 are essential for proper Rac turnoff. This circuit is masked by feedback and redundancy when chemoattractant is used as the input, highlighting the value of probing signaling networks at intermediate nodes to deconvolve complex signaling cascades.

Positioning the cleavage furrow: All you need is Rho.

blue LOV domains Review
J Cell Biol, 20 Jun 2016 DOI: 10.1083/jcb.201606010 Link to full text
Abstract: RhoA controls cleavage furrow formation during cell division, but whether RhoA suffices to orchestrate spatiotemporal dynamics of furrow formation is unknown. In this issue, Wagner and Goltzer (2016. J. Cell Biol show that RhoA activity can induce furrow formation in all cell cortex positions and cell cycle phases.

Local RhoA activation induces cytokinetic furrows independent of spindle position and cell cycle stage.

blue TULIP HeLa NIH/3T3 Control of cytoskeleton / cell motility / cell shape Cell cycle control
J Cell Biol, 13 Jun 2016 DOI: 10.1083/jcb.201603025 Link to full text
Abstract: The GTPase RhoA promotes contractile ring assembly and furrow ingression during cytokinesis. Although many factors that regulate RhoA during cytokinesis have been characterized, the spatiotemporal regulatory logic remains undefined. We have developed an optogenetic probe to gain tight spatial and temporal control of RhoA activity in mammalian cells and demonstrate that cytokinetic furrowing is primarily regulated at the level of RhoA activation. Light-mediated recruitment of a RhoGEF domain to the plasma membrane leads to rapid induction of RhoA activity, leading to assembly of cytokinetic furrows that partially ingress. Furthermore, furrow formation in response to RhoA activation is not temporally or spatially restricted. RhoA activation is sufficient to generate furrows at both the cell equator and cell poles, in both metaphase and anaphase. Remarkably, furrow formation can be initiated in rounded interphase cells, but not adherent cells. These results indicate that RhoA activation is sufficient to induce assembly of functional contractile rings and that cell rounding facilitates furrow formation.

Junctional actin assembly is mediated by Formin-like 2 downstream of Rac1.

blue AsLOV2 MCF10A Control of cytoskeleton / cell motility / cell shape Control of cell-cell / cell-material interactions
J Cell Biol, 11 May 2015 DOI: 10.1083/jcb.201412015 Link to full text
Abstract: Epithelial integrity is vitally important, and its deregulation causes early stage cancer. De novo formation of an adherens junction (AJ) between single epithelial cells requires coordinated, spatial actin dynamics, but the mechanisms steering nascent actin polymerization for cell-cell adhesion initiation are not well understood. Here we investigated real-time actin assembly during daughter cell-cell adhesion formation in human breast epithelial cells in 3D environments. We identify formin-like 2 (FMNL2) as being specifically required for actin assembly and turnover at newly formed cell-cell contacts as well as for human epithelial lumen formation. FMNL2 associates with components of the AJ complex involving Rac1 activity and the FMNL2 C terminus. Optogenetic control of Rac1 in living cells rapidly drove FMNL2 to epithelial cell-cell contact zones. Furthermore, Rac1-induced actin assembly and subsequent AJ formation critically depends on FMNL2. These data uncover FMNL2 as a driver for human epithelial AJ formation downstream of Rac1.

A light-triggered protein secretion system.

UV UVR8/UVR8 Cos-7 HEK293T rat hippocampal neurons Control of vesicular transport
J Cell Biol, 13 May 2013 DOI: 10.1083/jcb.201210119 Link to full text
Abstract: Optical control of protein interactions has emerged as a powerful experimental paradigm for manipulating and studying various cellular processes. Tools are now available for controlling a number of cellular functions, but some fundamental processes, such as protein secretion, have been difficult to engineer using current optical tools. Here we use UVR8, a plant photoreceptor protein that forms photolabile homodimers, to engineer the first light-triggered protein secretion system. UVR8 fusion proteins were conditionally sequestered in the endoplasmic reticulum, and a brief pulse of light triggered robust forward trafficking through the secretory pathway to the plasma membrane. UVR8 was not responsive to excitation light used to image cyan, green, or red fluorescent protein variants, allowing multicolor visualization of cellular markers and secreted protein cargo as it traverses the cellular secretory pathway. We implemented this novel tool in neurons to demonstrate restricted, local trafficking of secretory cargo near dendritic branch points.
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