Localization of Calmodulin 1a by In Situ Hybridization During Morphogenesis at the Midbrain-hindbrain Boundary in Zebrafish.

Proper development of vertebrate brain shape is required for normal brain function, obtained through the process of morphogenesis while cell shape changes are the core mediators of tissue shaping. Our lab uses the zebrafish system as a model to determine the molecular mechanisms that regulate cell shape during brain morphogenesis. Zebrafish make for a good model organism for brain development observations because of their external fertilization, rapid development, and transparent nature. The midbrain-hindbrain boundary (MHB) is a highly conserved fold that is one of the first to form after neurulation. We have found that the MHB constriction is mediated by non-muscle myosin (NMII) motor proteins to shorten and narrow cells at the MHB followed by basal constriction and tissue folds. However, the upstream mediators of NMII proteins in this context are unknown. Based on the known role for calcium in morphogenesis, we hypothesize that calcium may signal to NMII to regulate cell shape. It is known that calcium signals via calmodulins to regulate downstream kinases; therefore, we further hypothesize that calmodulin mediates the calcium signal. In zebrafish there are 6 calmodulin genes, all encoding for the same protein; however, calmodulin 1a has been shown to be expressed specifically in the MHB. We performed in situ hybridizations for calm1a throughout brain morphogenesis to determine if calm1a is a potential target for calcium signaling. Our results indicate that calm1a is expressed at 12 somite stage (ss) and disappears by 20ss at the MHB. This suggests that calm1a may have an active role in regulating calcium signaling at the MHB during morphogenesis. Additional experiments in the lab further suggest that calm1a is required to regulate cell length