Panoply™ Human CTSS Knockdown Stable Cell Line

Cathepsin S (CTSS) is a lysosomal cysteine protease reportedly involved in extracellular matrix (ECM) degradation, thereby promoting cell migration and invasion. Here, researchers knocked down CTSS expression using siRNA transfection and inhibited its enzymatic activity using the highly selective CTSS inhibitor RJW-58. In vitro functional experiments, Western blot analysis, and in vivo colonization models showed a positive correlation between CTSS and cell adhesion. Furthermore, both knockdown and inhibition of CTSS significantly reduced calcium influx via the storage-operated calcium ion influx (SOCE) pathway without altering the expression levels of STIM1 and Orai1. RJW-58 dose-dependently reduced the activation of calcium-dependent downstream effector molecules NFAT1 and Rac1. Immunoprecipitation experiments indicated that CTSS can bind to STIM1, and this binding can be reversed by the CTSS inhibitor. Furthermore, confocal microscopy and super-resolution imaging revealed that CTSS inhibition led to the accumulation of STIM1 spots in the endoplasmic reticulum and reduced the interaction between active STIM1 and EB1. In summary, these results demonstrate for the first time that the lysosomal cysteine protease CTSS plays a crucial role in mediating Ca2+ homeostasis by regulating STIM1 transport, thereby inhibiting cell migration and invasion.

To select a suitable cell model, the researchers first examined the expression of CTSS in six human cancer cell lines (Figure 1a). Since OEC-M1 and MDA-MB-231 showed similar expression patterns, these two cancer cell lines were selected to construct CTSS knockdown cells to investigate the role of CTSS in cell migration and invasion. The results of scratch assays and Transwell invasion assays showed that the migration (Figure 1b, c) and invasion (Figure 1d, e) abilities of cells were significantly reduced in CTSS knockdown cells.

Figure 1. CTSS knockdown significantly decreases cell migration and invasion. (Lin H H, et al., 2019)