Kai Zhang

The liver plays a critical role in lipid homeostasis, where lipids are either secreted as very-low-density lipoproteins (VLDL) or stored in lipid droplets (LDs). However, the regulatory mechanisms governing these two interconnected processes remain poorly understood. Here, we demonstrate that SEC16B functions as a lipid-responsive regulator in the liver, promoting VLDL secretion and LD expansion to handle lipid flux and maintain lipid homeostasis. Genome-wide association studies have identified single-nucleotide polymorphisms in SEC16B to be highly associated with serum lipid levels in humans. Hepatic Sec16b deficiency decreases serum lipid levels by impairing VLDL secretion through mechanisms that are at least partially independent of microsomal triglyceride transfer protein (MTP)-mediated ApoB lipidation and COPII-mediated intracellular trafficking. SEC16B partially localizes at ER-LD contact sites and promotes LD expansion by facilitating the targeting of ER proteins to LDs. More importantly, suppression of Sec16b dramatically lowers serum lipid levels and reduces atherosclerotic lesion size in Ldlr null mice. These data reveal a mechanism that coordinates VLDL and LD metabolism and suggest SEC16B as a potential therapeutic target for atherosclerosis treatment.

Mitochondrial membrane potential (MMP) is essential for mitochondrial functions, yet current methods for modulating MMP lack precise spatial and temporal control. Here, we present an optogenetic system that enables reversible formation of inter-mitochondrial contacts (mito-contacts) with high spatiotemporal precision. Blue light stimulation induces rapid formation of mito-contacts, which fully dissipate upon cessation of illumination. These light-induced mito-contacts can enhance MMP, leading to increased ATP production under stress conditions. Moreover, in human retinal cells and C. elegans, high MMP induced by mito-contacts alleviates the deleterious effects of prolonged blue light exposure, restoring energy metabolism and extending organismal lifespan. This optogenetic approach provides a powerful tool for modulating MMP and offers potential therapeutic applications for diseases linked to mitochondrial dysfunction.

Optogenetics and chemogenetics have transformed how physiologists interrogate biological systems by enabling precise control over protein activity and cellular function. Optogenetics uses light-sensitive proteins for rapid and localized control, while chemogenetics employs small molecules to trigger or block specific pathways with systemic and sustained effects. These tools have advanced research in areas such as brain function, heart rhythm, immune response, and gene regulation. They have been applied to disease models that include epilepsy, metabolic and cardiovascular diseases, immunoinflammatory disorders, and cancer. Clinical applications are emerging, such as optogenetic therapies for vision restoration and chemogenetic safety switches in engineered immune cells. In this review, we categorize these tools by their mechanisms of action, compare their advantages and limitations, and discuss strategies to improve their precision, efficiency, and translational capability. As these technologies continue to evolve, they offer powerful approaches to dissect complex physiological processes and drive innovative therapeutic interventions.

Neuronal activity robustly engages the extracellular signal-regulated kinase (ERK) signaling pathway through Calcium-dependent mechanisms; however, whether ERK can acutely and causally modulates ongoing neuronal activity remains unsolved due to complex upstream regulation and diverse subcellular functions. Here, we directly address this question using an optogenetic ERK activator, opto-miniRaf, that enables selective, rapid, graded, and reversible control of ERK signaling. Combining this AAV-compatible system with calcium imaging and electrophysiology, we interrogate ERK functions across biological scales, from cultured neurons, acute brain slices, and the intact brain. Acute optogenetic activation of ERK enhances synchronized network burst activity in cultured rat cortical neurons and increases calcium activity of cortical pyramidal neurons in awake and moving mice following non-invasive light stimulation. Together, these results establish ERK signaling as an acute modulator of neuronal and network activity, positioning opto-miniRaf as a generalizable platform for precise spatiotemporal control of intracellular kinase signaling in complex biological systems.

The soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein syntaxin mediates neuronal exocytosis and self-assembles into large clusters in the plasma membrane. The formation and function of these clusters, and whether they promote or inhibit synaptic-vesicle fusion, remain unclear. Here using optogenetic control of syntaxin clustering in vitro and in vivo, as a light-inducible gain-of-function assay, we show that light-enhanced clustering reduces both spontaneous and triggered vesicle fusion, and this impairs mouse hunting behavior. Cluster formation is induced by liquid–liquid phase separation (LLPS) of the SNARE domain of syntaxin. For the regulatory mechanism, Munc18, which is known to alter syntaxin conformation, acts to reduce LLPS for cluster formation, thereby promoting active syntaxin. These results suggest that exocytosis regulation involves LLPS-induced syntaxin clusters that serve as a syntaxin reservoir from which Munc18 captures syntaxin monomers to form a syntaxin–Munc18 complex, setting the stage for efficient fusion.

Mitochondrial membrane potential (MMP) is essential for mitochondrial functions, yet current methods for modulating MMP lack precise spatial and temporal control. Here, we developed an optogenetic mitochondrial tethering system, termed OptoMitoTether, that enables reversible control of inter-mitochondrial proximity with high spatiotemporal precision. Blue light activation induced CRY2PHR oligomerization between adjacent mitochondrial outer membranes, reduced mitochondrial nearest-neighbor distances, and promoted inter-mitochondrial tethering. OptoMitoTether activation was associated with elevated MMP and was associated with lower reactive oxygen species (ROS) and preserved ATP production during prolonged blue light stress. Moreover, OptoMitoTether expression preserved mitochondrial bioenergetic function in human retinal cells and extended lifespan in C. elegans during chronic blue light exposure. These findings establish OptoMitoTether as a tool for manipulating mitochondrial proximity and reveal a link between inter-mitochondrial tethering and mitochondrial bioenergetic state during blue light stress.

Oligomerization of photoactivatable proteins is widely used in optogenetics to modulate protein activity and regulate biological processes. However, their oligomerization states remain challenging to quantify in living cells. We applied photon counting histogram (PCH) analysis to quantify the oligomerization of two commonly used photoactivatable proteins, Vaucheria frigida Aureochrome light-oxygen-voltage (VfAuLOV) and Arabidopsis Thaliana cryptochrome 2 (AtCRY2), under dark and light conditions, providing a direct measurement of oligomerization states in live cells. Under blue light stimulation, VfAuLOV primarily forms dimers, whereas AtCRY2 forms higher order multimers in HEK293T cells. At intracellular concentrations above 1000 nM, AtCRY2 transitions into tetramers upon light stimulation, consistent with structural data obtained from cryo-electron microscopy. Unexpectedly, AtCRY2 (W374), a constitutively active mutant, still exhibits light-induced oligomerization. Human cryptochrome 2, in contrast, shows light-independent oligomerization at the intracellular concentration above 100 nM, below which the monomeric state dominates. Optogenetic signaling outcomes, demonstrated by light-induced lytic cell death, align with the oligomerization state of tested proteins. This study presents a quantitative framework for elucidating protein oligomerization in live cells, thereby enhancing the understanding of optogenetic mechanisms and their role in cellular signaling. Significance Statement: Photoactivatable proteins enable the development of optical actuators that modulate cell signaling through light-mediated protein-protein interaction. Light-oxygen-voltage (LOV) and cryptochrome 2 (CRY2) are commonly used photoactivable protein modules that undergo blue light-mediated oligomerization, and the latter has been used to create synthetic biomolecular condensates in live cells. Although in vitro structural studies suggested their oligomerization states, their behavior in live cells is unclear. Here, we applied photon counting histogram analysis to determine the dark and light-activated oligomerization states of these photoactivatable proteins in live cells. Insights gained will provide better guidance in designing optical actuators, leading to more quantitative interpretation of the light-mediated signaling outcomes in cells.