Biology subjects

Sun, D.-e.

Publications and source records attributed to Sun, D.-e..

4 recordsLinked to original sources

bioRxiv2025

Localizable fluorescent metal ion indicators with tunable colors

Elucidating the role of metal ion homeostasis in physiological and pathological processes requires detection tools with high sensitivity and selectivity, spectral versatility, and precise subcellular localization. Here, we introduce a modular strategy for generating fluorescent metal ion indicators which are comprised of a sulfonamide-functionalized chelator, a rhodamine derivative and a ligand for bioconjugation to self-labeling proteins. With a concise three-step synthesis, the design allows tuning of both the emission color and ligand specificity of the resulting indicators. Specifically, we developed a family of bright, color-tunable potassium indicators which can be selectively coupled to intra- or extracellular HaloTag and SNAP-tag fusion proteins. The increase in fluorescence upon binding to HaloTag or SNAP-tag fluorogenicity enabled wash-free live-cell imaging. The localized potassium indicators enabled the detection of dynamic potassium efflux in rat hippocampal neurons upon glutamate stimulation. Our work thus establishes a versatile platform for the generation of localizable fluorescent metal ion indicators and opens up new avenues for live-cell potassium sensing.

cell biology↗

bioRxiv2025

A high-affinity split-HaloTag for live-cell protein labeling

We introduce a high-affinity split-HaloTag comprised of a short peptide tag (Hpep, 14 residues) and a large, inactive fragment (cpHalo{Delta}3). Hpep binds to cpHalo{Delta}3 spontaneously with nanomolar affinity, enabling subsequent labeling with fluorescent HaloTag ligands. The small size of Hpep facilitates cloning-free endogenous protein tagging using CRISPR/Cas9 and the complementation of Hpep-tagged proteins can be achieved in live cells through co-expression with cpHalo{Delta}3 and in fixed cells through incubation with cpHalo{Delta}3. The approach is compatible with advanced microscopy techniques such as expansion microscopy and live-cell STED imaging. Additionally, variants of Hpep that modulate the spectral properties of labeled fluorophores enable simultaneous imaging of two different Hpep-tagged proteins via fluorescence lifetime microscopy. In summary, our high-affinity split-HaloTag is a robust and versatile tool for live-cell imaging and diverse applications in chemical biology.

biochemistry↗

bioRxiv2024

Molecular recording of cellular protein kinase activity with chemical labeling

Protein kinases control most cellular processes and aberrant kinase activity is involved in numerous diseases. To investigate the link between specific kinase activities and cellular phenotypes in heterogeneous cell populations and in vivo, we introduce molecular recorders of kinase activities for later analysis. Based on split-HaloTag and a phosphorylation-dependent molecular switch, our recorders become rapidly labeled in the presence of a specific kinase activity and a fluorescent HaloTag substrate. The kinase activity in a given cell controls the degree of fluorescent labeling whereas the recording window is set by the presence of the fluorescent substrate. We have designed specific recorders for four protein kinases, including protein kinase A. We apply our protein kinase A recorder for the sorting of heterogeneous cell populations and subsequent transcriptome analysis, in genome-wide CRISPR screens to discover regulators of PKA activity and for the tracking of neuromodulation in freely moving mice.

biochemistry↗

bioRxiv2020

Click-ExM enables expansion microscopy for all biomolecules

Expansion microscopy (ExM) allows super-resolution imaging on conventional fluorescence microscopes, but has been limited to proteins and nucleic acids. Here we develop click-ExM, which integrates click-labeling into ExM to enable a "one-stop-shop" method for nanoscale imaging of various types of biomolecules. Using 18 clickable labels for click-ExM imaging of DNA, RNA, proteins, lipids, glycans and small molecules, we demonstrate its universality, compatibility with signal-amplification techniques, and broad applications in cellular imaging.

bioengineering↗