TTL-Expression Modulates Epithelial Morphogenesis.

Epithelial monolayer formation depends on the architecture and composition of the microtubule cytoskeleton. Microtubules control bidirectional trafficking and determine the positioning of structural cellular proteins. We studied the role of tubulin tyrosination in epithelial cell shape and motility. Tubulin tyrosine ligase (TTL), the enzyme that adds tyrosine to the carboxy terminus of detyrosinated α-tubulin, was depleted or overexpressed in 2D epithelial monolayers as well as in 3D intestinal organoids. We demonstrate qualitatively and quantitatively that in the absence of TTL the cells comprise high levels of detyrosinated tubulin, change their shape into an initial flat morphology and retardedly acquire a differentiated columnar epithelial cell shape. Enhanced adhesion and accelerated migration patterns of TTL-knockout cells combined with reverse effects in TTL-overexpressing cells indicate that the loss of TTL affects the organization of cell adhesion foci. Precipitation of detyrosinated tubulin with focal adhesion scaffold components coincides with increased quantities and persistence of focal adhesion plaques. Our results indicate that the equilibrium between microtubules enriched in detyrosinated or tyrosinated tubulin modulates epithelial tissue formation, cell morphology, and adhesion.

Sorafenib induces paradoxical phosphorylation of the extracellular signal-regulated kinase pathway in acute myeloid leukemia cells lacking FLT3-ITD mutation.

Gain-of-function mutations in the RAS and FLT3 genes are frequently found in cells of acute myeloid leukemia (AML), leading to constitutive activation of signaling pathways that regulate fundamental cellular processes, and are therefore attractive targets for AML therapy. The multi-targeted kinase inhibitor sorafenib is efficacious in AML with FLT3-internal tandem duplication (ITD), but resistance to therapy is an important clinical problem. It is unclear whether AML lacking FLT3-ITD responds to sorafenib. Using AML cell lines, we have shown that a low concentration of sorafenib induces opposing effects depending on the oncogenic background. In FLT3-ITD positive cells sorafenib blocks Erk activity and cell proliferation, and triggers apoptosis. However, in cells lacking FLT3-ITD, sorafenib paradoxically activates Erk2, and stimulates cellular proliferation and metabolic activity. Thus, depending on the genetic context, sorafenib is a beneficial inhibitor or paradoxical activator of mitogenic signaling pathways in AML. These results harbor important consequences in planning clinical trials in AML.