Ann E. Sutherland
Primary AppointmentAssociate Professor, Cell Biology
Cell Matrix Interactions In Mouse Development
Regulation of trophoblast motility. We discovered that amino acids are signaling molecules that regulate the onset of invasive behavior of the trophoblast cells of the mouse blastocyst that is required for it to implant into the uterus. In particular, arginine and leucine are each necessary, and together they are sufficient, to induce the trophoblast cells to become motile. They regulate motility through activation of the serine/threonine kinase mTOR, which acts through an unknown downstream effector to activate Rac1, thus promoting protrusive activity. It is likely that this signaling system acts during development to coordinate the development of the embryo with the receptivity of the uterus, and we are currently examining this question by assaying concentrations of amino acids and ions in uterine fluid during the preimplantation stages of pregnancy. We are also examining the transport mechanisms responsible for uptake of leucine and arginine, and how the amino acids affect the activity of mTOR.
Extracellular matrix effects on trophoblast motility. We found that the trophoblast cells encounter a new array of extracellular matrix components as they invade into the uterus, and that two of the most prominent proteins are laminin-111 and laminin-521. These two laminins have vastly different effects on cell behavior; laminin-111 is a repulsive substrate leading to cell rounding and boundary formation, while laminin-521 is a very adhesive substrate and promotes spreading. The distribution of laminin-111 vs. laminin-521 suggests that trophoblast cell invasive behavior is limited in the direction of the embryo by laminin-111, and directed into the uterine stroma by laminin-521. We have tested this hypothesis by examining the behavior of trophoblast cells in embryos lacking laminin-111, and in mice where laminin -521 is knocked out in the uterus. The results confirm our hypothesis, and show that the environment of the uterus has a profound effect on the ability of the embryo to implant. Currently we are working on defining the signaling pathways within trophoblast cells that lead to the different responses to the two laminin isoforms.
Cellular mechanisms of gastrulation in the mouse. In collaboration with Drs. Xiaowei Lu (Cell Biology), Ray Keller (Biology), Ammasi Periasamy (Keck Center for Cellular Imaging), and Carol Burdsal (Tulane University), we have undertaken a study of cell behavior during gastrulation in the mid-gestation mouse embryo. We worked out techniques for in vitro culture of the embryos and for long-term (6-8 hours) time-lapse confocal imaging, using transgenic animals that express fluorescent reporter proteins (EGFP and Tomato fluorescent protein). With this we have obtained the first high-resolution imaging of individual cell behavior in the mouse embryo. This has enabled us to describe the cell behaviors that drive elongation of the primary axis of the embryo, and to extend the analysis to embryos lacking proteins of the planar cell polarity (PCP) signaling system, which exhibit both a shortened primary axis and neural tube closure defects. We have found that one of these PCP proteins, PTK7, normally promotes polarization of mesodermal cells and development of intercalation behavior that leads to convergence and extension of the somites. In the absence of this protein, mesodermal cells do not polarize and continue to exhibit directed migration that expands the tissue rather than lengthening it. We are now analyzing the behavior of mesodermal cells in another PCP mutant, Loop tail (Lp), which exhibits a similar phenotype of shortened axis and open neural tube. Interestingly , the mesodermal cells in Lp embryos do not have a defect in polarization or convergent extension, suggesting that they may develop a shortened axis through a different cellular mechanism. We plan to extend this analysis to the neural tube , to determine the normal cellular mechanisms underlying neural tube closure, and to other mutant mouse lines that exhibit shortened axis.