A vinculin tail hotspot mutation in cancer patients affects mouse embryonic fibroblast motility

Vinculin, an essential adhesion scaffolding protein, physically links membrane bound integrin and cadherin receptors to filamentous actin. Cells that fail to express vinculin or possess dysfunctional vinculin exhibit rounded morphology, enhanced motility, and resistance to apoptosis and anoikis. Therefore, vinculin is classified as a tumor suppressor protein in which mutations have serious consequences in cancer. A possible ‘hotspot’ region—an encoded region of a protein that is highly inclined to mutate and phenotypically manifest—has been identified at in the tail-domain of vinculin at residue R925. At this residue, a specific histidine mutation has been linked to several types of cancer. The purpose of this study was to examine the possibility of residue R925 as a hotspot region and test whether the R925H mutation in vinculin enhances motility properties in a model cell line. To examine the status of R925 as a hotspot region, we mined several databases to link certain prevalent mutations in vinculin to different types of cancer. R925H was the most prominent mutation in vinculin that was present in 5 types of cancer, confirming R925 as a hotspot region. In order to assess whether R925H affected the motility of mouse embryonic fibroblasts (MEFs), actin binding and bundling activity of mutant vinculin was compared to that of wild-type vinculin. We found that R925H mutant vinculin had lower actin bundling ability. To evaluate the changes in motility patterns caused by the introduction of R925H, we conducted random motility assays to measure the two-dimensional migration of MEFs on micro-fabricated and microfluidic devices. Mutant MEFs on hydrogels displayed significant differences in motility compared to wild-type cells—particularly in total accumulated distance, persistence and velocity. However, further investigation is required to interpret these findings. The insight on the effects of the R925H mutation on cell motility will serve as one of the first steps to unravel the complexities of vinculin’s potential role in several human cancer settings.Bachelor of Scienc