Calmodulin downregulation in conditional knockout HeLa cells inhibits cell migration

The study of calmodulin (CaM) functions in living cells has been tackled up to date using cell-permeant CaM inhibitors or interference-RNA methods. CaM inhibitors may lack specificity and the siRNA interference approach is challenging, as all three CaM genes expressing an identical protein in mammals have to be blocked. Therefore, we recently introduced a novel genetic system using CRISPR/Cas9-mediated gene deletion and conditional CaM expression to study the function of CaM in HeLa cells. Here, we describe the effect of CaM downregulation on the basal and epidermal growth factor (EGF)-dependent 2D- and 3D-migration in HeLa cells. CaM downregulation inhibited cell migration on a 2D-surface in the absence but not in the presence of EGF. In contrast, CaM downregulation led to inhibition of 3D-migration across a porous membrane both in the absence and presence of EGF. CaM downregulation decreased the expression of Racl, Cdc42 and RhoA, all known to play crucial roles in cell migration. These results show that EGF-dependent 2D- and 3D-migration utilize distinct CaM-regulated systems and identify several essential migratory proteins directly or indirectly regulated by CaM.

Automated summary

In this study, a novel genetic system using CRISPR/Cas9-mediated gene deletion and conditional CaM expression was introduced to investigate the function of CaM in HeLa cells. The results showed that CaM downregulation inhibited both 2D and 3D migration in HeLa cells, with an impact on 2D migration in the presence of EGF. Additionally, downregulation of CaM decreased the expression of Racl, Cdc42, and RhoA, known to play crucial roles in cell migration.