Scalable recorders of cellular calcium dynamics and molecular interactions across time

Cellular processes are mediated by dynamic biochemical events in their native context, including changes in intracellular ion concentrations and transient interactions between molecules. Studying these processes across large cell populations requires recording their dynamics across time in single cells, simultaneously and at scale. Here, we introduce split-1POK, a split protein tape monomer whose two halves reconstitute when the domains fused to them interact. The reconstituted monomer is incorporated into the protein tape as it elongates across time in live cells, enabling scalable, spatiotemporally resolved endpoint readout under conventional light microscopy. Split-1POK provides a programmable scaffold for new recorders in the CytoSense family, our previously introduced framework (ref. 1) in which the same cytosol-localized monomer senses cellular activity and writes it along protein tape recorders. Unlike transcription-based activity recorders, which encode the signal through new gene expression, CytoSense recorders act on monomers post-synthesis, so the activity signal is sensed and written directly in the cytosol without passing through transcription- or translation-associated cascades. Fusing calmodulin and its binding peptide to split-1POK, we built TCaMP (Tape monomer-CalModulin-Peptide), a genetically encoded calcium recorder (GECR) that measures calcium dynamics on a single, analog intensity scale along a continuous time axis, providing quantitative comparison of calcium levels over time within the same cell. In cultured human cells, TCaMP records transient calcium dynamics from transmembrane entry, receptor activation, and intracellular store release, detecting stimulations as brief as ten minutes. With hour-scale kinetics over days, TCaMP complements fluorescent calcium indicators by trading sub-second temporal resolution for scalable, long-term recording. Fusing conditional dimerization pairs to split-1POK yields recorders of molecular interaction histories in response to external stimuli. Split-1POK thus brings scalable, spatiotemporally resolved recording to intracellular calcium dynamics and molecular interactions across time.