Ascidians belong to the Tunicate phylum, which groups the closest living relatives to vertebrates and display a simplified but typical chordate body plan during embryonic and larval stages. Ascidian embryos offer several advantages for experimental studies of developmental gene activities and cell behavior. First, they develop with a reduced number of cells, the lineages of which are invariant and have been determined up to the pre-gastrula 110-cell stage. This allows for visualization of developmental events with a sub-cellular resolution using live microscopy. Second, extensive genomic and expression resources are readily available through user-friendly web interfaces. This facilitates functional studies utilizing microinjection of antisense morpholino oligonucleotides and an experimental tool unique to ascidians among metazoans: the simultaneous electroporation of hundreds of fertilized eggs with synthetic plasmid DNA constructs, which allows for cis-regulatory analyses using reporter genes and targeted expression of recombinant and/or fluorescent proteins using tissue-specific enhancers for functional studies and fluorescent microscopy. In ascidians, the so-called trunk ventral cells (TVCs) migrate from the anterior part of the tail to the ventral part of the trunk in tailbud embryos and constitute the precardiac mesoderm. They originate from a single pair of blastomeres in the early embryo, called B7.5 cells. The B7.5 blastomeres give birth to the TVCs and their sister cells, the anterior tail muscles (ATMs), which differentiate into skeletal muscle and do not migrate. Previous studies showed that TVC-speific gene expression and migration require transcriptional inputs from the bHLH transcription factor Mesp, the FGF signaling pathway and the forkhead transcription factor FoxF. Because of these functional evidences, TVC migration constitutes a suitable model system to investigate the relationship between transcription regulation and directed cell migration. To this aim, I developed a method utilizing fluorescence activated cell sorting (FACS) and microarray analysis to obtain TVC-specific genome transcription profiles. These data, together with an analysis of the function and regulation of the Rho GTPase RhoDF, indicated that 1) the gene regulatory network impinges on most cellular processes underlying cell migration, including actin dynamics, cell-matrix adhesion, polarity and vesicle trafficking 2) for each cellular process, only a subset of the effector genes are subjected to transcription regulation and 30 specific cell behaviors result from the modular association of individual cellular processes that can be experimentally uncoupled from each other. This study led to the identification of additional candidate regulators and effectors of TVC migration and to the definition of a conceptual and experimental framework for future studies. 1) A systems-level analysis is required to understand the structure and function of the interface between the precardiac GRN and TVC migration. Towards this goal, functional analysis of additional candidate transcription factors and signaling molecules will be conducted and complemented by a reciprocal focus on the sub-cellular processes at work during TVC migration and subjected to transcription regulation. 2) Transcription profiles of migrating TVC determine their ability to interpret the extracellular signals that influence their behavior. These extrinsic cues will be sought by a candidate gene approach and unbiased screens to assess the function of surrounding tissues during TVC migration. 3) The hypothesized modularity of cell behavior implies that similar basic processes function in other cell-types with distinct behaviors, thus motivating comparisons with other cell-types in the ascidian embryo and with other migrating cells in other species. Areas of Research/Interest Tissue-specific transcription control of basic processes underlying cell behavior.
Publications
|
|
|
|
