Fig. 1: The structure, localization, and expression of RIM-BP2 molecular biosensors.
From: Visualization of RIM-BP2’s crane-like function in neuronal vesicle transport using FRET
A Connection modes of BKTS and RKTS biosensors to RIM-BP2 and presynaptic membrane. B The mechanosensing principle of the stress detection unit of biosensors. The flexible linker is selected from spider silk spring protein, and the stress detection unit is equipped with FRET fluorescent protein pair. Mechanical stretching/compression alters the distance between fluorescent proteins, enabling reflection of molecular tension through FRET efficiency changes. Fluorescence images of (C) BKTS and (F) RKTS biosensor in the 488 nm emission (ECFP) and 530 nm emission (YPet) under 405 nm excitation in SH-SY5Y cells. D, E, G, H FRET ratio images obtained by K-means clustering analysis after YPet/ECFP. D, G Depict regions with significant FRET ratio changes for the BKTS and RKTS biosensors, respectively. E, H Illustrate non-responsive regions for the BKTS and RKTS biosensors, respectively. I–K Expression of (I) BTS, J RTS, and K KTS control biosensors in SH-SY5Y cells, and YPet/ECFP FRET ratio images of the cells during 5 min of 10 Hz electrical stimulation, respectively. L YPet/ECFP FRET ratio images of BKTS and RKTS in SH-SY5Y cells under electrical stimulation after knockdown of RIM-BP2 using siRIMBP2-1. M Normalized FRET ratio curves and N statistical analysis of BKTS and RKTS before and after knocking down RIM-BP2 over time. BKTS, n = 6; RKTS, n = 6. Student’s t test, ***P < 0.001. Data are mean ± SEM.
