Biology subjects

Lemon, W. C.

Publications and source records attributed to Lemon, W. C..

5 recordsLinked to original sources

bioRxiv2025

Multimodal cell lineage reconstruction in the hindbrain reveals a link between progenitor origin and activity patterning

Does the stem cell origin impact how daughter neurons acquire functional characteristics and assemble into circuits? Here, by multimodal cell lineage reconstruction in the zebrafish hindbrain we related a neurons embryonic origin to its future terminal differentiation features, such as neurotransmitter identity, and neuronal activity pattern. Intersectional lineage tracing, new developed computational tools, and genetic knockouts revealed that different progenitors formed functionally distinct neuron subtypes and could not compensate for the loss of adjacent progenitor pools, indicating developmental hardwiring. Dynamics of neuronal production suggest that progenitor competence changes over time. Whereas neurog1-expressing progenitors contributed to both glutamatergic and GABAergic lineages at early embryonic stages, later, other progenitor pools also assumed this role. Whole-hindbrain 3D atlases combining calcium imaging to monitor spontaneous neuronal activity, with genetic perturbations and progenitor origin information, unveiled that the emergence of neuronal activity patterns was presaged by their progenitor origins. This reveals a link between cell ontogeny and neuronal activity in the zebrafish hindbrain.

developmental biology↗

bioRxiv2023

A modular chemigenetic calcium indicator enables in vivo functional imaging with near-infrared light

Genetically encoded fluorescent calcium indicators have revolutionized neuroscience and other biological fields by allowing cellular-resolution recording of physiology during behavior. However, we currently lack bright, genetically targetable indicators in the near infrared that can be used in animals. Here, we describe WHaloCaMP, a modular chemigenetic calcium indicator built from bright dye-ligands and protein sensor domains that can be genetically targeted to specific cell populations. Fluorescence change in WHaloCaMP results from reversible quenching of the bound dye via a strategically placed tryptophan. WHaloCaMP is compatible with rhodamine dye-ligands that fluoresce from green to near-infrared, including several dye-ligands that efficiently label the central nervous system in animals. When bound to a near-infrared dye-ligand, WHaloCaMP1a is more than twice as bright as jGCaMP8s, and shows a 7x increase in fluorescence intensity and a 2.1 ns increase in fluorescence lifetime upon calcium binding. We use WHaloCaMP1a with near-infrared fluorescence emission to image Ca2+ responses in flies and mice, to perform three-color multiplexed functional imaging of hundreds of neurons and astrocytes in zebrafish larvae, and to quantitate calcium concentration using fluorescence lifetime imaging microscopy (FLIM).

neuroscience↗

bioRxiv2021

Automated Reconstruction of Whole-Embryo Cell Lineages by Learning from Sparse Annotations

We present a method for automated nucleus identification and tracking in time-lapse microscopy recordings of entire developing embryos. Our method combines deep learning and global optimization to enable complete lineage reconstruction from sparse point annotations, and uses parallelization to process multi-terabyte light-sheet recordings, which we demonstrate on three common model organisms: mouse, zebrafish, Drosophila. On the most difficult dataset (mouse), our method correctly reconstructs 75.8% of cell lineages spanning 1 hour, compared to 31.8% for the previous state of the art, thus enabling biologists to determine where and when cell fate decisions are made in developing embryos, tissues, and organs.

bioinformatics↗

bioRxiv2019

In vivo glucose imaging in multiple model organisms with an engineered single-wavelength sensor

Glucose is arguably the most important molecule in metabolism, and its mismanagement underlies diseases of vast societal import, most notably diabetes. Although glucose-related metabolism has been the subject of intense study for over a century, tools to track glucose in living organisms with high spatio-temporal resolution are lacking. We describe the engineering of a family of genetically encoded glucose sensors with high signal-to-noise ratio, fast kinetics and affinities varying over four orders of magnitude (1 {micro}M to 10 mM). The sensors allow rigorous mechanistic characterization of glucose transporters expressed in cultured cells with high spatial and temporal resolution. Imaging of neuron/glia co-cultures revealed [~]3-fold higher glucose changes in astrocytes versus neurons. In larval Drosophila central nervous system explants, imaging of intracellular neuronal glucose suggested a novel rostro-caudal transport pathway in the ventral nerve cord neuropil, with paradoxically slower uptake into the peripheral cell bodies and brain lobes. In living zebrafish, expected glucose-related physiological sequelae of insulin and epinephrine treatments were directly visualized in real time. Additionally, spontaneous muscle twitches induced glucose uptake in muscle, and sensory- and pharmacological perturbations gave rise to large but enigmatic changes in the brain. These sensors will enable myriad experiments, most notably rapid, high-resolution imaging of glucose influx, efflux, and metabolism in behaving animals.

bioengineering↗

bioRxiv2017

A general method to fine-tune fluorophores for live-cell and in vivo imaging

AO_SCPCAPBSTRACTC_SCPCAPPushing the frontier of fluorescence microscopy requires the design of enhanced fluorophores with finely tuned properties. We recently discovered that incorporation of four-membered azetidine rings into classic fluorophore structures elicits substantial increases in brightness and photostability, resulting in the Janelia Fluor (JF) series of dyes. Here, we refine and extend this strategy, showing that incorporation of 3-substituted azetidine groups allows rational tuning of the spectral and chemical properties with unprecedented precision. This strategy yields a palette of new fluorescent and fluorogenic labels with excitation ranging from blue to the far-red with utility in live cells, tissue, and animals.

cell biology↗