Spatially regulated β-actin translation in epithelia
Creators: name: Vedula, Pavan
Publisher: name: No Publisher Supplied
Year: 2016

description: The β-actin mRNA contains a 28 nucleotide sequence in the 3′UTR termed the zipcode which, along with a trans acting factor called the Zipcode Binding Protein-1 (ZBP-1/IMP-1), regulates the spatio-temporal expression pattern of the β-actin. This mode of post-transcriptional gene regulation is required for several cellular processes including cell migration, cell-cell adhesion, and cell-matrix adhesion. Adding to the list of signaling cascades such as RhoA and Src, work in this thesis demonstrates E-cadherin endocytosis, and plausibly gene expression pathways regulating cell proliferation, converge on spatially regulated β-actin translation. E-cadherin endocytosis and recycling has been shown to be important during the maintenance of epithelial adherens junctions. However in this thesis, endocytosis of E-cadherin is shown to negatively regulate adherens junction assembly, while spatially regulated β-actin translation balances this by promoting actin filament anchoring of cadherin complexes. Mislocalization of β-actin translation impairs adherens junction maturation in epithelial cells following de novo cell-cell contact. These results have implications in development and disease where the assembly and maintenance of epithelial adherens junctions is essential. In order to quantify the extent of adherens junction assembly defects in epithelial cells following de novo contact, a Fluorescence Covariance Index based on calculating Pearson’s Correlation Coefficients was developed and validated using several components of the adherens junction complex. In addition to quantitating the morphological effects of mislocalizing β-actin translation on adherens junctions, the FCI analysis showed correlation of these morphological defects with loss of epithelial barrier function. Lastly, inhibiting dynamin mediated endocytosis rescued adherens junction structure and epithelial barrier function in cells with partially mislocalized β-actin. Using Matrigel embedded 3D cyst cultures of MDCK cells, spatially regulated β-actin translation was shown to have an effect on cell shape, size and proliferation during epithelial morphogenesis. These defects in assembling a normal 3D epithelium along with disorganized acto-myosin and tight junctions, point to a role for compartmentalized β-actin translation in regulating a mechanosensitive module – plausibly Hippo and/or YAP/TAZ pathways. Finally, collective cell migration, an important paradigm in several biological processes such as wound repair and development, was used to determine the role of spatially regulated β-actin translation in epithelial wound repair. Spatially regulated β-actin translation is shown to regulate actin flux at cell-cell adhesions in leader-follower cell pairs to control tissue migration rates. Although cells with partially mislocalized β-actin translation have a more active leading edge, this increased protrusive activity fails to translate into increased migratory rate. These data taken together suggest that collective cell migration in epithelial sheets is an emergent property of both single cell dynamics and cell-cell interactions.