We have discovered a requirement for a novel connexin gene (frm-cx) in specification of the anterior neural plate in the invertebrate chordate Ciona. (The connexin proteins make gap junctions.) In embryos from the frimousse (frm) mutant line, the anterior neural plate is transfated to epidermis. The anterior neural plate of Ciona gives rise both to brain and to two other non-neural tissues, the mouth and adhesive palp. The initial FGF-dependent induction of the anterior neural plate appears to be normal in the mutant, as judged by expression of early response genes. However, the expression of these genes is quickly lost, and the anterior neural plate derivatives become incorporated into the epidermis and express epidermal genes. We hypothesize that the frm-cx gene product is required for maintaining neural plate identity by allowing for intercellular communication between the newly induced anterior neural plate cells. We also hypothesize that that frm-cx expression is an early response to anterior neural induction. Experiments in the first two Specific Aims of this proposal will test these hypotheses. Because some connexins have biological activities distinct from their channel forming role, in the first Specific Aim we will investigate the necessity of gap junction communication within the newly-induced neural plate. In further experiments to determine whether the anterior neural plate makes a distinct communication compartment within the embryo, we will assess the cellular connectivity of cells using a photoactivatable gap junction-permeable fluorescent dye. Additional experiments will investigate whether over expression of frm-cx is sufficient to cause expansion of the neural plate. In the second Specific Aim we will place frm-cx within the regulatory context of anterior neural induction by determining its epigenetic relationship to other known anterior neural transcription factors. To test for the necessity of these factors we will isolate a minimal regulatory element from the frm-cx gene. In this element we will identify and then mutate potential transcription factor binding sites. The final Specific Aim will characterize a new mutant line, bugeye (bug) that disrupts the same set of cells affected in the frm mutant. In the bug mutant neural tube folding and closure (neurulation), but not specification, are disrupted. Specific experiments will fully characterize the mechanical defects in neurulation in bug mutants by confocal imaging and 3-dimensional modeling. The final goal of this Specific Aim is to postionally clone the bug mutant using established methods.

Public Health Relevance

The formation of the central nervous system is a critical component of human development. This proposal will examine the cellular and molecular mechanisms of early events in brain development using an animal that models human brain development.

National Institute of Health (NIH)
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Research Project (R01)
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Special Emphasis Panel (ZRG1-CB-N (02))
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Mukhopadhyay, Mahua
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University of California Santa Barbara
Schools of Arts and Sciences
Santa Barbara
United States
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Abdul-Wajid, Sarah; Veeman, Michael T; Chiba, Shota et al. (2014) Exploiting the extraordinary genetic polymorphism of ciona for developmental genetics with whole genome sequencing. Genetics 197:49-59
Hackley, Christopher; Mulholland, Erin; Kim, Gil Jung et al. (2013) A transiently expressed connexin is essential for anterior neural plate development in Ciona intestinalis. Development 140:147-55
Veeman, Michael T; Chiba, Shota; Smith, William C (2011) Ciona genetics. Methods Mol Biol 770:401-22
Tresser, Jason; Chiba, Shota; Veeman, Michael et al. (2010) doublesex/mab3 related-1 (dmrt1) is essential for development of anterior neural plate derivatives in Ciona. Development 137:2197-203
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Veeman, Michael T; Nakatani, Yuki; Hendrickson, Carolyn et al. (2008) Chongmague reveals an essential role for laminin-mediated boundary formation in chordate convergence and extension movements. Development 135:33-41

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