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4 Janelia Publications

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    09/28/21 | A serotonergic axon-cilium synapse drives nuclear signaling to maintain chromatin accessibility
    Shu-Hsien Sheu , Srigokul Upadhyayula , Vincent Dupuy , Song Pang , Andrew L. Lemire , Deepika Walpita , H. Amalia Pasolli , Fei Deng , Jinxia Wan , Lihua Wang , Justin Houser , Silvia Sanchez-Martinez , Sebastian E. Brauchi , Sambashiva Banala , Melanie Freeman , C. Shan Xu , Tom Kirchhausen , Harald F. Hess , Luke Lavis , Yu-Long Li , Séverine Chaumont-Dubel , David E. Clapham
    bioRxiv. 2021 Sep 28:

    Chemical synapses between axons and dendrites mediate much of the brain’s intercellular communication. Here we describe a new kind of synapse – the axo-ciliary synapse - between axons and primary cilia. By employing enhanced focused ion beam – scanning electron microscopy on samples with optimally preserved ultrastructure, we discovered synapses between the serotonergic axons arising from the brainstem, and the primary cilia of hippocampal CA1 pyramidal neurons. Functionally, these cilia are enriched in a ciliary-restricted serotonin receptor, 5-hydroxytryptamine receptor 6 (HTR6), whose mutation is associated with learning and memory defects. Using a newly developed cilia-targeted serotonin sensor, we show that optogenetic stimulation of serotonergic axons results in serotonin release onto cilia. Ciliary HTR6 stimulation activates a non-canonical Gαq/11-RhoA pathway. Ablation of this pathway results in nuclear actin and chromatin accessibility changes in CA1 pyramidal neurons. Axo-ciliary synapses serve as a distinct mechanism for neuromodulators to program neuron transcription through privileged access to the nuclear compartment.

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    04/01/21 | The HaloTag as a general scaffold for far-red tunable chemigenetic indicators.
    Deo C, Abdelfattah AS, Bhargava HK, Berro AJ, Falco N, Farrants H, Moeyaert B, Chupanova M, Lavis LD, Schreiter ER
    Nature Chemical Biology. 2021 Apr 01:. doi: 10.1038/s41589-021-00775-w

    Functional imaging using fluorescent indicators has revolutionized biology, but additional sensor scaffolds are needed to access properties such as bright, far-red emission. Here, we introduce a new platform for 'chemigenetic' fluorescent indicators, utilizing the self-labeling HaloTag protein conjugated to environmentally sensitive synthetic fluorophores. We solve a crystal structure of HaloTag bound to a rhodamine dye ligand to guide engineering efforts to modulate the dye environment. We show that fusion of HaloTag with protein sensor domains that undergo conformational changes near the bound dye results in large and rapid changes in fluorescence output. This generalizable approach affords bright, far-red calcium and voltage sensors with highly tunable photophysical and chemical properties, which can reliably detect single action potentials in cultured neurons.

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    08/20/20 | Rational design of bioavailable photosensitizers for manipulation and imaging of biological systems.
    Binns TC, Ayala AX, Grimm JB, Tkachuk AN, Castillon GA, Phan S, Zhang L, Brown TA, Liu Z, Adams SR, Ellisman MH, Koyama M, Lavis LD
    Cell Chemical Biology. 2020 Aug 20;27(8):1063-72. doi: 10.1016/j.chembiol.2020.07.001

    Light-mediated chemical reactions are powerful methods for manipulating and interrogating biological systems. Photosensitizers, compounds that generate reactive oxygen species upon excitation with light, can be utilized for numerous biological experiments, but the repertoire of bioavailable photosensitizers is limited. Here, we describe the synthesis, characterization, and utility of two photosensitizers based upon the widely used rhodamine scaffold and demonstrate their efficacy for chromophore-assisted light inactivation, cell ablation in culture and in vivo, and photopolymerization of diaminobenzidine for electron microscopy. These chemical tools will facilitate a broad range of applications spanning from targeted destruction of proteins to high-resolution imaging.

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    01/09/20 | Bright and tunable far-red chemigenetic indicators.
    Deo C, Abdelfattah AS, Bhargava HK, Berro AJ, Falco N, Moeyaert B, Chupanova M, Lavis LD, Schreiter ER
    bioRxiv. 2020 Jan 9: