Lant B et al. (2014) Cold Spring Harb Protoc "Induction of germline apoptosis in Caenorhabditis elegans."

Derry WB, Lant B

[Cold Spring Harb Protoc, 2014]

RNA interference (RNAi) is an incredibly powerful tool for rapid and efficient knockdown of gene expression. This technology can be used to induce apoptosis in the germline of Caenorhabditis elegans. Genotoxic stressors such as ionizing radiation (IR), ultraviolet light, chemical mutagens (e.g., N-ethyl-N-nitrosourea [ENU]), and DNA cross-linking reagents can also be used to stimulate apoptosis. These approaches, described here, combined with the powers of in vivo imaging methods, should keep C. elegans apoptosis researchers busy for several years, sorting out how various signaling pathways influence life and death decisions in this organism.

Arata Y et al. (2014) Front Physiol "Power law relationship between cell cycle duration and cell volume in the early embryonic development of Caenorhabditis elegans."

Takagi H, Sawa H, Sako Y, Arata Y

[Front Physiol, 2014]

Cell size is a critical factor for cell cycle regulation. In Xenopus embryos after midblastula transition (MBT), the cell cycle duration elongates in a power law relationship with the cell radius squared. This correlation has been explained by the model that cell surface area is a candidate to determine cell cycle duration. However, it remains unknown whether this second power law is conserved in other animal embryos. Here, we found that the relationship between cell cycle duration and cell size in Caenorhabditis elegans embryos exhibited a power law distribution. Interestingly, the powers of the time-size relationship could be grouped into at least three classes: highly size-correlated, moderately size-correlated, and potentially a size-non-correlated class according to C. elegans founder cell lineages (1.2, 0.81, and <0.39 in radius, respectively). Thus, the power law relationship is conserved in Xenopus and C. elegans, while the absolute powers in C. elegans were different from that in Xenopus. Furthermore, we found that the volume ratio between the nucleus and cell exhibited a power law relationship in the size-correlated classes. The power of the volume relationship was closest to that of the time-size relationship in the highly size-correlated class. This correlation raised the possibility that the time-size relationship, at least in the highly size-correlated class, is explained by the volume ratio of nuclear size and cell size. Thus, our quantitative measurements shed a light on the possibility that early embryonic C. elegans cell cycle duration is coordinated with cell size as a result of geometric constraints between intracellular structures.

Garde A et al. (2022) Dev Cell "Localized glucose import, glycolytic processing, and mitochondria generate a focused ATP burst to power basement-membrane invasion."

Garde A, Chi Q, Sherwood DR, Kelley LC, Kenny IW, Mutlu AS, Wang MC

[Dev Cell, 2022]

Invasive cells use transient, energy-consuming protrusions to breach basement membrane (BM) barriers. Using the ATP sensor PercevalHR during anchor cell (AC) invasion in Caenorhabditis elegans, we show that BM invasion is accompanied by an ATP burst from mitochondria at the invasive front. RNAi screening and visualization of a glucose biosensor identified two glucose transporters, FGT-1 and FGT-2, which bathe invasive front mitochondria with glucose and facilitate the ATP burst to form protrusions. FGT-1 localizes at high levels along the invasive membrane, while FGT-2 is adaptive, enriching most strongly during BM breaching and when FGT-1 is absent. Cytosolic glycolytic enzymes that process glucose for mitochondrial ATP production cluster with invasive front mitochondria and promote higher mitochondrial membrane potential and ATP levels. Finally, we show that UNC-6 (netrin), which polarizes invasive protrusions, also orients FGT-1. These studies reveal a robust and integrated energy acquisition, processing, and delivery network that powers BM breaching.

Berry BJ et al. (2020) FASEB J "Neuronal AMPK coordinates mitochondrial energy sensing and hypoxia resistance in C. elegans."

Berry BJ, Nieves TO, Baldzizhar A, Wojtovich AP

[FASEB J, 2020]

Organisms adapt to their environment through coordinated changes in mitochondrial function and metabolism. The mitochondrial protonmotive force (PMF) is an electrochemical gradient that powers ATP synthesis and adjusts metabolism to energetic demands via cellular signaling. It is unknown how or where transient PMF changes are sensed and signaled due to the lack of precise spatiotemporal control in vivo. We addressed this by expressing a light-activated proton pump in mitochondria to spatiotemporally "turn off" mitochondrial function through PMF dissipation in tissues with light. We applied our construct-mitochondria-OFF (mtOFF)-to understand how metabolic status impacts hypoxia resistance, a response that relies on mitochondrial function. Activation of mtOFF induced starvation-like behavior mediated by AMP-activated protein kinase (AMPK). We found prophylactic mtOFF activation increased survival following hypoxia, and that protection relied on neuronal AMPK. Our study links spatiotemporal control of mitochondrial PMF to cellular metabolic changes that mediate behavior and stress resistance.

Yang F et al. (2023) Nat Methods "Pulsed stimulated Brillouin microscopy enables high-sensitivity mechanical imaging of live and fragile biological specimens."

Koehler S, Prevedel R, Ellenberg J, Watanabe K, Gopalan A, Bernabeu M, Govendir M, Neves A, Bevilacqua C, Yang F, Hambura S, Diz-Munoz A, Jechlinger M, Rapti G

[Nat Methods, 2023]

Brillouin microscopy is an emerging optical elastography technique capable of assessing mechanical properties of biological samples in a three-dimensional, all-optical and noncontact fashion. The typically weak Brillouin scattering signal can be substantially enhanced via a stimulated Brillouin scattering (SBS) process; however, current implementations require high pump powers, which prohibit applications to photosensitive or live imaging of biological samples. Here we present a pulsed SBS scheme that takes advantage of the nonlinearity of the pump-probe interaction. In particular, we show that the required pump laser power can be decreased ~20-fold without affecting the signal levels or spectral precision. We demonstrate the low phototoxicity and high specificity of our pulsed SBS approach by imaging, with subcellular detail, sensitive single cells, zebrafish larvae, mouse embryos and adult Caenorhabditis elegans. Furthermore, our method permits observing the mechanics of organoids and C. elegans embryos over time, opening up further possibilities for the field of mechanobiology.

Zhang F et al. (2007) Nature "Multimodal fast optical interrogation of neural circuitry."

Gottschalk A, Liewald JF, Deisseroth K, Zhang F, Wood PG, Bamberg E, Wang LP, Watzke N, Brauner M, Nagel G, Kay K

[Nature, 2007]

Our understanding of the cellular implementation of systems-level neural processes like action, thought and emotion has been limited by the availability of tools to interrogate specific classes of neural cells within intact, living brain tissue. Here we identify and develop an archaeal light-driven chloride pump (NpHR) from Natronomonas pharaonis for temporally precise optical inhibition of neural activity. NpHR allows either knockout of single action potentials, or sustained blockade of spiking. NpHR is compatible with ChR2, the previous optical excitation technology we have described, in that the two opposing probes operate at similar light powers but with well-separated action spectra. NpHR, like ChR2, functions in mammals without exogenous cofactors, and the two probes can be integrated with calcium imaging in mammalian brain tissue for bidirectional optical modulation and readout of neural activity. Likewise, NpHR and ChR2 can be targeted together to Caenorhabditis elegans muscle and cholinergic motor neurons to control locomotion bidirectionally. NpHR and ChR2 form a complete system for multimodal, high-speed, genetically targeted, all-optical interrogation of living neural circuits.

Tian D et al. (2015) Curr Biol "Anillin Regulates Neuronal Migration and Neurite Growth by Linking RhoG to the Actin Cytoskeleton."

Chen Z, Zhang Y, Sun L, Diao M, Ou G, Jiang Y, Huang S, Tian D

[Curr Biol, 2015]

Neuronal migration and neurite growth are essential events in neural development, but it remains unclear how guidance cues are transduced through receptors to the actin cytoskeleton, which powers these processes. We report that a cytokinetic scaffold protein, Anillin, is redistributed to the leading edge of the C.elegans Q neuroblast during cell migration and neurite growth. To bypass the requirement for Anillin in cytokinesis, we used the somatic CRISPR-Cas9 technique to generate conditional mutations in Anillin. We demonstrate that Anillin regulates cell migration and growth cone extension by stabilizing the F-actin network at the leading edge. Our biochemical analysis shows that the actin-binding domain of Anillin is sufficient to stabilize F-actin by antagonizing the F-actin severing activity of Cofilin. We further uncover that the active form of RhoG/MIG-2 directly binds to Anillin and recruits it to the leading edge. Our results reveal a novel pathway in which Anillin transduces the RhoG signal to the actin cytoskeleton during neuronal migration and neurite growth.

Berry, Brandon J. et al. (2021) MicroPubl Biol

Wojtovich, Andrew P., Berry, Brandon J., Nieves, Tyrone O.

[MicroPubl Biol, 2021]

Mitochondria are organelles that make ATP by generating a proton gradient across the inner membrane through the action of the electron transport chain (ETC). The gradient is composed of a membrane potential (m) and a concentration gradient (pH). In addition to energy production, mitochondria are recognized as signaling organelles (Chandel 2015), and m powers diverse signaling outputs that mitochondria coordinate. One such signaling response is the mitochondrial unfolded protein response (UPRmt). Some ETC complex components are encoded by mitochondrial or nuclear genes and require coordination between the genomes for proper stoichiometry (Shpilka and Haynes 2018). The UPRmt is a protein homeostasis response that is activated when nuclear and mitochondrial gene expression are not correctly coordinated (Melber and Haynes 2018). When the expression of mitochondrial and nuclear encoded proteins is mismatched, the UPRmt is activated. In C. elegans, the UPRmt is controlled by the transcription factor ATFS-1, which is normally trafficked to mitochondria and degraded. When nuclear:mitochondrial gene expression is perturbed, ATFS-1 is blocked from entering mitochondria and is instead trafficked to the nucleus where a host of chaperones and stress-response genes are activated and transcribed.

Watts DJ et al. (1998) Nature "Collective dynamics of 'small-world' networks."

Strogatz SH, Watts DJ

[Nature, 1998]

Networks of coupled dynamical systems have been used to model biological oscillators, Josephson junction arrays, excitable media, neural networks, spatial games, genetic control networks and many other self-organizing systems. Ordinarily, the connection topology is assumed to be either completely regular or completely random. But many biological, technological and social networks lie somewhere between these two extremes. Here we explore simple models of networks that can be tuned through this middle ground: regular networks 'rewired' to introduce increasing amounts of disorder. We find that these systems can be highly clustered, like regular lattices, yet have small characteristic path lengths, like random graphs. We call the 'small-world' networks, by analogy with the small-world phenomenon (popularly known as six degrees of separation). The neural network of the worm Caenorhabditis elegans, the power grid of the western United States, and the collaboration graph of film actors are shown to be small-world networks. Models of dynamical systems with small-world coupling display enhanced signal-propagation speed, computational powers and synchronizability. In particular, infectious diseases spread more easily in small-world networks than in regular lattices.

Leitz G et al. (2002) Biophys J "Stress response in Caenorhabditis elegans caused by optical tweezers: wavelength, power, and time dependence."

Leitz G, Tuck S, Fallman E, Axner O

[Biophys J, 2002]

Optical tweezers have emerged as a powerful technique for micromanipulation of living cells. Although the technique often has been claimed to be nonintrusive, evidence has appeared that this is not always the case. This work presents evidence that near-infrared continuous-wave laser light from optical tweezers can produce stress in Caenorhabditis elegans. A transgenic strain of C. elegans, carrying an integrated heat-shock-responsive reporter gene, has been exposed to laser light under a variety of illumination conditions. It was found that gene expression was most often induced by light of 760 nm, and least by 810 nm. The stress response increased with laser power and irradiation time. At 810 nm, significant gene expression could be observed at 360 mW of illumination, which is more than one order of magnitude above that normally used in optical tweezers. In the 700-760-nm range, the results show that the stress response is caused by photochemical processes, whereas at 810 nm, it mainly has a photothermal origin. These results give further evidence that the 700-760-nm wavelength region is unsuitable for optical tweezers and suggest that work at 810 nm at normal laser powers does not cause stress at the cellular level.