ALDH1L1 Inhibits Cell Motility Via Dephosphorylation of Cofilin by PP1 and PP2A

Here we report that ALDH1L1 (FDH, a folate enzyme with tumor suppressor-like properties) inhibits cell motility. The underlying mechanism involves F-actin stabilization, re-distribution of cytoplasmic actin towards strong preponderance of filamentous actin, and formation of actin stress fibers. A549 cells expressing FDH demonstrated a much slower recovery of GFP-actin fluorescence in a FRAP assay, as well as an increase in G-actin polymerization and a decrease in F-actin depolymerization rates in pyren-actin fluorescence assays indicating the inhibition of actin dynamics. These effects were associated with robust dephosphorylation of the actin depolymerizing factor cofilin by PP1 and PP2A serine/threonine protein phosphatases but not the cofilin-specific phosphatases slingshot and chronophin. In fact, the PP1/PP2A inhibitor calyculin prevented cofilin dephosphorylation and restored motility. Inhibition of FDH-induced apoptosis by the JNK inhibitor SP600125 or the pan-caspase inhibitor zVAD-fmk did not restore motility or levels of phospho-cofilin, indicating that the observed effects are independent from FDH function in apoptosis. Interestingly, cofilin siRNA or expression of phosphorylation-deficient S3A cofilin mutant resulted in a decrease of G-actin and the actin stress fiber formation, the effects seen upon FDH expression. In contrast, the expression of S3D mutant, mimicking constitutive phosphorylation, prevented these effects further supporting the cofilin-dependent mechanism. Dephosphorylation of cofilin and inhibition of motility in response to FDH can be also prevented by the increased folate in media. Furthermore, folate depletion itself, in the absence of FDH, resulted in cofilin dephosphorylation and inhibition of motility in several cell lines. Our experiments showed that these effects were folate-specific and not a general response to nutrient starvation. Overall, this study demonstrates the presence of distinct intracellular signaling pathways regulating motility in response to folate status and points toward mechanisms involving folates in promoting a malignant phenotype.