Early Vertebrate Brain Formation Requires Wnt5b and Microtubule Mediated Tissue Folding
Mentor 1
Jennifer Gutzman
Location
Union Wisconsin Room
Start Date
27-4-2018 1:00 PM
Description
Congenital brain defects can lead to physical or mental disabilities and are among the most common birth defects, occurring in 1 to 2 out of every 1000 live births. Determining how the brain acquires its shape during development is critical for elucidating the etiology of these defects. To understand formation of brain structure, we use the zebrafish model to study the mechanisms that regulate the first fold in the neuroepithelium, the highly conserved midbrain-hindbrain boundary (MHB). We have identified several cell shape changes that are required to form the deep constriction in the neuroepithelium, the MHB constriction (MHBC). One mediator of these cell shape changes is Wnt5b, which is specifically expressed in this region during the onset of morphogenesis. Using wnt5b knockdown, cell shape was analyzed by live confocal imaging of membrane GFP-injected embryos. We developed a new morphometric technique to analyze these data using digital sectioning to reveal 3D cell shape. With this analysis, we have uncovered a role for Wnt5b in mediating anisotropic, or polarized, cell shape to fold the neuroepithelium. Data from 2D gel analysis indicated that Wnt5b regulates α-tubulin levels at the MHB. α-tubulin is one of the monomers of microtubules, and its regulation modulates overall microtubule dynamics. To test the role of microtubules during tissue folding, colchicine was used to destabilize microtubules and MHBC cell shape was analyzed. We found that destabilization of microtubules disrupts cell shape and MHB folding. To test whether Wnt5b-mediated cell shape was microtubule-dependent, paclitaxel was used to stabilize microtubules and was found to rescue the cell shape defects that result from wnt5b knockdown. Overall, our data suggests that Wnt5b is crucial in regulating microtubule dynamics to control cell shape. This mechanism may be important for shaping other Wnt5b expressing cells and tissues throughout development.
Early Vertebrate Brain Formation Requires Wnt5b and Microtubule Mediated Tissue Folding
Union Wisconsin Room
Congenital brain defects can lead to physical or mental disabilities and are among the most common birth defects, occurring in 1 to 2 out of every 1000 live births. Determining how the brain acquires its shape during development is critical for elucidating the etiology of these defects. To understand formation of brain structure, we use the zebrafish model to study the mechanisms that regulate the first fold in the neuroepithelium, the highly conserved midbrain-hindbrain boundary (MHB). We have identified several cell shape changes that are required to form the deep constriction in the neuroepithelium, the MHB constriction (MHBC). One mediator of these cell shape changes is Wnt5b, which is specifically expressed in this region during the onset of morphogenesis. Using wnt5b knockdown, cell shape was analyzed by live confocal imaging of membrane GFP-injected embryos. We developed a new morphometric technique to analyze these data using digital sectioning to reveal 3D cell shape. With this analysis, we have uncovered a role for Wnt5b in mediating anisotropic, or polarized, cell shape to fold the neuroepithelium. Data from 2D gel analysis indicated that Wnt5b regulates α-tubulin levels at the MHB. α-tubulin is one of the monomers of microtubules, and its regulation modulates overall microtubule dynamics. To test the role of microtubules during tissue folding, colchicine was used to destabilize microtubules and MHBC cell shape was analyzed. We found that destabilization of microtubules disrupts cell shape and MHB folding. To test whether Wnt5b-mediated cell shape was microtubule-dependent, paclitaxel was used to stabilize microtubules and was found to rescue the cell shape defects that result from wnt5b knockdown. Overall, our data suggests that Wnt5b is crucial in regulating microtubule dynamics to control cell shape. This mechanism may be important for shaping other Wnt5b expressing cells and tissues throughout development.
