TY - JOUR AB - Mosaic analysis with double markers (MADM) technology enables the sparse labeling of genetically defined neurons. We present a protocol for time-lapse imaging of cortical projection neuron migration in mice using MADM. We describe steps for the isolation, culturing, and 4D imaging of neuronal dynamics in MADM-labeled brain tissue. While this protocol is compatible with other single-cell labeling methods, the MADM approach provides a genetic platform for the functional assessment of cell-autonomous candidate gene function and the relative contribution of non-cell-autonomous effects. For complete details on the use and execution of this protocol, please refer to Hansen et al. (2022),1 Contreras et al. (2021),2 and Amberg and Hippenmeyer (2021).3 AU - Hansen, Andi H AU - Hippenmeyer, Simon ID - 14794 IS - 1 JF - STAR Protocols TI - Time-lapse imaging of cortical projection neuron migration in mice using mosaic analysis with double markers VL - 5 ER - TY - JOUR AB - The plant-signaling molecule auxin triggers fast and slow cellular responses across land plants and algae. The nuclear auxin pathway mediates gene expression and controls growth and development in land plants, but this pathway is absent from algal sister groups. Several components of rapid responses have been identified in Arabidopsis, but it is unknown if these are part of a conserved mechanism. We recently identified a fast, proteome-wide phosphorylation response to auxin. Here, we show that this response occurs across 5 land plant and algal species and converges on a core group of shared targets. We found conserved rapid physiological responses to auxin in the same species and identified rapidly accelerated fibrosarcoma (RAF)-like protein kinases as central mediators of auxin-triggered phosphorylation across species. Genetic analysis connects this kinase to both auxin-triggered protein phosphorylation and rapid cellular response, thus identifying an ancient mechanism for fast auxin responses in the green lineage. AU - Kuhn, Andre AU - Roosjen, Mark AU - Mutte, Sumanth AU - Dubey, Shiv Mani AU - Carrillo Carrasco, Vanessa Polet AU - Boeren, Sjef AU - Monzer, Aline AU - Koehorst, Jasper AU - Kohchi, Takayuki AU - Nishihama, Ryuichi AU - Fendrych, Matyas AU - Sprakel, Joris AU - Friml, Jiří AU - Weijers, Dolf ID - 14826 IS - 1 JF - Cell KW - General Biochemistry KW - Genetics and Molecular Biology SN - 0092-8674 TI - RAF-like protein kinases mediate a deeply conserved, rapid auxin response VL - 187 ER - TY - JOUR AB - Production of hydrogen at large scale requires development of non-noble, inexpensive, and high-performing catalysts for constructing water-splitting devices. Herein, we report the synthesis of Zn-doped NiO heterostructure (ZnNiO) catalysts at room temperature via a coprecipitation method followed by drying (at 80 °C, 6 h) and calcination at an elevated temperature of 400 °C for 5 h under three distinct conditions, namely, air, N2, and vacuum. The vacuum-synthesized catalyst demonstrates a low overpotential of 88 mV at −10 mA cm–2 and a small Tafel slope of 73 mV dec–1 suggesting relatively higher charge transfer kinetics for hydrogen evolution reactions (HER) compared with the specimens synthesized under N2 or O2 atmosphere. It also demonstrates an oxygen evolution (OER) overpotential of 260 mV at 10 mA cm–2 with a low Tafel slope of 63 mV dec–1. In a full-cell water-splitting device, the vacuum-synthesized ZnNiO heterostructure demonstrates a cell voltage of 1.94 V at 50 mA cm–2 and shows remarkable stability over 24 h at a high current density of 100 mA cm–2. It is also demonstrated in this study that Zn-doping, surface, and interface engineering in transition-metal oxides play a crucial role in efficient electrocatalytic water splitting. Also, the results obtained from density functional theory (DFT + U = 0–8 eV), where U is the on-site Coulomb repulsion parameter also known as Hubbard U, based electronic structure calculations confirm that Zn doping constructively modifies the electronic structure, in both the valence band and the conduction band, and found to be suitable in tailoring the carrier’s effective masses of electrons and holes. The decrease in electron’s effective masses together with large differences between the effective masses of electrons and holes is noticed, which is found to be mainly responsible for achieving the best water-splitting performance from a 9% Zn-doped NiO sample prepared under vacuum. AU - Kiran, Gundegowda Kalligowdanadoddi AU - Singh, Saurabh AU - Mahato, Neelima AU - Sreekanth, Thupakula Venkata Madhukar AU - Dillip, Gowra Raghupathy AU - Yoo, Kisoo AU - Kim, Jonghoon ID - 14828 IS - 1 JF - ACS Applied Energy Materials KW - Electrical and Electronic Engineering KW - Materials Chemistry KW - Electrochemistry KW - Energy Engineering and Power Technology KW - Chemical Engineering (miscellaneous) SN - 2574-0962 TI - Interface engineering modulation combined with electronic structure modification of Zn-doped NiO heterostructure for efficient water-splitting activity VL - 7 ER - TY - JOUR AB - Bacteria divide by binary fission. The protein machine responsible for this process is the divisome, a transient assembly of more than 30 proteins in and on the surface of the cytoplasmic membrane. Together, they constrict the cell envelope and remodel the peptidoglycan layer to eventually split the cell into two. For Escherichia coli, most molecular players involved in this process have probably been identified, but obtaining the quantitative information needed for a mechanistic understanding can often not be achieved from experiments in vivo alone. Since the discovery of the Z-ring more than 30 years ago, in vitro reconstitution experiments have been crucial to shed light on molecular processes normally hidden in the complex environment of the living cell. In this review, we summarize how rebuilding the divisome from purified components – or at least parts of it - have been instrumental to obtain the detailed mechanistic understanding of the bacterial cell division machinery that we have today. AU - Radler, Philipp AU - Loose, Martin ID - 14834 IS - 1 JF - European Journal of Cell Biology KW - Cell Biology KW - General Medicine KW - Histology KW - Pathology and Forensic Medicine SN - 0171-9335 TI - A dynamic duo: Understanding the roles of FtsZ and FtsA for Escherichia coli cell division through in vitro approaches VL - 103 ER - TY - JOUR AB - De novo heterozygous variants in KCNC2 encoding the voltage-gated potassium (K+) channel subunit Kv3.2 are a recently described cause of developmental and epileptic encephalopathy (DEE). A de novo variant in KCNC2 c.374G > A (p.Cys125Tyr) was identified via exome sequencing in a patient with DEE. Relative to wild-type Kv3.2, Kv3.2-p.Cys125Tyr induces K+ currents exhibiting a large hyperpolarizing shift in the voltage dependence of activation, accelerated activation, and delayed deactivation consistent with a relative stabilization of the open conformation, along with increased current density. Leveraging the cryogenic electron microscopy (cryo-EM) structure of Kv3.1, molecular dynamic simulations suggest that a strong π-π stacking interaction between the variant Tyr125 and Tyr156 in the α-6 helix of the T1 domain promotes a relative stabilization of the open conformation of the channel, which underlies the observed gain of function. A multicompartment computational model of a Kv3-expressing parvalbumin-positive cerebral cortex fast-spiking γ-aminobutyric acidergic (GABAergic) interneuron (PV-IN) demonstrates how the Kv3.2-Cys125Tyr variant impairs neuronal excitability and dysregulates inhibition in cerebral cortex circuits to explain the resulting epilepsy. AU - Clatot, Jerome AU - Currin, Christopher AU - Liang, Qiansheng AU - Pipatpolkai, Tanadet AU - Massey, Shavonne L. AU - Helbig, Ingo AU - Delemotte, Lucie AU - Vogels, Tim P AU - Covarrubias, Manuel AU - Goldberg, Ethan M. ID - 14841 IS - 3 JF - Proceedings of the National Academy of Sciences of the United States of America TI - A structurally precise mechanism links an epilepsy-associated KCNC2 potassium channel mutation to interneuron dysfunction VL - 121 ER -