Illuminating Plant Cell Contact Sites: Using Optogenetics to Explore ER-PM Interactions with LiMETER

In eukaryotic cells, the endoplasmic reticulum (ER) plays a central role in coordinating cellular functions such as lipid synthesis, calcium signaling, and protein trafficking. These functions often rely on the ER's physical and functional interactions with other organelles through membrane contact sites (MCS)—specialized regions where two organelles come into close proximity without fusing. One of the most prominent and functionally significant of these are the ER–plasma membrane contact sites (EPCS). Despite their importance, studying the dynamics and regulation of EPCS in plant cells has remained technically challenging, largely due to the lack of tools capable of labeling these contact sites in a reversible and non-disruptive manner.

As part of a larger interdisciplinary research effort, we applied an optogenetic tool, LiMETER (Light-inducible Membrane-Tethered cortical ER), to overcome these limitations and investigate EPCS behavior in plant cells. LiMETER offers a light-controlled system to reversibly recruit a fluorescently labeled ER marker to the plasma membrane upon blue light stimulation. This level of spatiotemporal control allows for dynamic tracking of EPCS formation and disassembly in live cells, without the prolonged structural changes induced by more static reporters like MAPPER.

Our study showed that LiMETER enables rapid and reversible labeling of EPCS with high specificity and minimal impact on ER morphology or plant development. Importantly, we demonstrated that EPCSs are not uniformly distributed along the ER network. Rather, they preferentially localize at ER tubules and at the edges of ER cisternae—regions likely optimized for rapid signal relay or membrane exchange. Using LiMETER, we further uncovered that EPCS abundance increases under abiotic stress conditions such as cold exposure, and that this increase correlates with the overexpression of ER-shaping proteins like reticulons. This suggests that EPCS formation is both environmentally responsive and developmentally regulated, and may be closely tied to ER structural dynamics.

Moreover, our findings support a model in which the ER network’s shape and function are interdependent with membrane contact site formation. In rapidly growing or highly dynamic cell types such as pollen tubes, where ER streaming is pronounced, fewer stable EPCS are observed—highlighting a possible trade-off between ER mobility and its capacity to establish stable organelle contacts.

By enabling dynamic, non-invasive visualization of contact site formation, LiMETER represents a powerful advancement for plant cell biology. It also exemplifies the utility of optogenetics in resolving long-standing technical barriers in plant systems, where genetic tools and marker systems have historically lagged behind those available in animal models.

Ultimately, this work not only enhances our understanding of how plant cells coordinate membrane interfaces during development and stress response, but also lays the groundwork for future studies into other types of ER-organelle interactions. Tools like LiMETER will be essential for dissecting the roles of MCS in diverse processes ranging from signal transduction to organelle biogenesis, with broad implications for plant productivity and stress resilience.