A major focus of current research in my lab is aimed at understanding mechanisms that control axon outgrowth and guidance. Our goal is to understand how guidance signals function to control axon motility and behavior in the complex in vivo environment, where axons must integrate multiple cues. We use zebrafish embryos as a model system, which allows us to combine molecular/genetic manipulations with live imaging of cell behavior and molecular activity in living intact embryos. In the past several years we have identified multiple molecular signals that guide axons in vivo and have shown how these signals function to control axon motile behavior, growth and direction in several regions of the CNS. One of the neuron types we study, the spinal sensory Rohon-Beard (RB) neurons, make an excellent model to investigate guidance mechanisms because they have stereotyped axon arbor morphology and can be readily imaged in vivo.
A main goal of our recent and current research is to further investigate the molecular mechanisms underlying differential guidance of central versus peripheral RB axons. We are exploring the roles of LIM homeodomain transcription factors in regulating RB morphology and axon trajectory. Inhibition of islet family LIM transcription factors by expression of a dominant negative form of a required cofactor (DN-CLIM) causes a strong reduction or elimination of RB and trigeminal peripheral axons. However, the cell bodies and central axons develop normally, indicating that LIM transcription factors function specifically to control peripheral sensory axon outgrowth. Although transcription factors have been shown in several systems to have key functions in defining axon trajectories, the molecular steps between transcriptional regulation and control of axon motility and morphology remain largely unknown. We have taken two general approaches to defining these molecular steps: 1) We are using live imaging of axon behavior and molecular activity to determine which cell motility processes are affected by DN-CLIM; and 2) We have done a microarray gene expression analysis to identify downstream targets of LIM transcription factors. We have recently published a manuscript characterizing motile behaviors of axons and dynamics of F-actin accumulation in DN-CLIM embryos. Other highlights of the past year are promising results analyzing the function of several genes found in our microarray.
Another main research area in the lab is aimed at understanding mechanisms of neural crest cell (NCC) migration, and in particular the epithelial to mesenchymal transition (EMT) that NCCs undergo to delaminate from the neuroepithelium and begin migration. EMT is a dramatic process involving major changes in cell morphology and motility that allow cell migration and formation of new tissues. EMTs are important for many developmental processes, and are also co-opted in several pathological processes, including cancer metastasis. The mechanisms controlling cell changes during EMT remain poorly understood. Because a cellís environment strongly influences its motility and intracellular signaling, it is extremely useful to have a model in which we can study these mechanisms while cells undergo EMT in the natural 3D environment. We are again taking advantage of the zebrafish model to combine in vivo imaging of NCC behavior during EMT with manipulation of potential signaling molecules. This year we received an exploratory R21 grant from NIH to develop this system as a means to examine the function and activity of RhoGTPases in controlling cell behavior in an in vivo EMT model.
Andersen, E.A. and Halloran, M.C. (2012) Centrosome movements in vivo correlate with specific neurite formation downstream of LIM homeodomain transcription factor activity. Development 139:3590-3599.
Andersen, E.A., Asuri, N.S. and Halloran, M.C. (2011) In vivo imaging of cell behaviors and F-actin reveals LIM-HD transcription factor regulation of peripheral versus central sensory axon development. Neural Development 6:27.
Clay, M.R. and Halloran, M.C. (2011) Regulation of cell adhesions and motility during initiation of neural crest migration. Curr Opinion Neurobiol 21(1):17-22
Clay, M.R. and Halloran, M.C. (2010) Control of neural crest cell behavior and migration: insights from live imaging. Cell Adh Migr 4(4):582-590.
Andersen, E., Asuri, N., Clay, M., Halloran, M. (2010). Live Imaging of Cell Motility and Actin Cytoskeleton of Individual Neurons and Neural Crest Cells in Zebrafish Embryos. JoVE 36. http://www.jove.com/index/details.stp?id=1726, doi: 10.3791/1726.
Sittaramane V., Sawant A., Wolman M.A., Maves L., Halloran M.C., Chandrasekhar A. (2009) The cell adhesion molecule Tag1, transmembrane protein Stbm/Vangl2, and Laminin-alpha1 exhibit genetic interactions during migration of facial branchiomotor neurons in zebrafish. Dev Biol 325:363-373.
Berndt, J.D., Clay, M.R., Langenberg, T., and Halloran, M.C. (2008). Rho-kinase and myosin II affect dynamic neural crest cell behaviors during epithelial to mesenchymal transition in vivo. Dev Biol 324:236-244.
Langenberg, T., Kahana, A., Wszalek, J.A., and Halloran, M.C. (2008). The eye organizes neural crest cell migration. Dev Dyn 237:1645-1652.
Wolman, M.A., Sittaramane, V.K., Essner, J.J., Yost, H.J., Chandrasekhar, A. and Halloran, M.C. (2008) Transient axonal glycoprotein-1 (TAG-1) and laminin-?1 regulate dynamic growth cone behaviors and initial axon direction in vivo. Neural Development 3:6.
Wolman, M.A., Regnery, A.M., Becker, T., Becker, C.G., and Halloran, M.C. (2007) Semaphorin3D regulates axon-axon interactions by modulating levels of L1CAM. J Neurosci 27:9653-9663.
Berndt, J.D. and Halloran, M.C. (2006)
Semaphorin3D promotes cell proliferation and neural crest cell
development downstream of TCF in the zebrafish hindbrain. Development 133:3983-3992.
Halloran, M.C. and Wolman, M.A. (2006) Repulsion
or adhesion: receptors make the call. Current Opinion
Cell Biology 18:533-540.
Sakai, J.A. and Halloran, M.C. (2006) Semaphorin3D
guides laterality of retinal ganglion cell projections in zebrafish. Development 133:1035-1044.
Paulus, J.D. and Halloran, M.C. (2006) Zebrafish bashful/Laminin-a1 mutants
exhibit multiple axon guidance defects. Dev Dyn 235:213-224.
Liu, Y. and Halloran, M.C. (2005) Central
and peripheral branches from one neuron are guided differentially
by Sema3D and TAG-1. J Neurosci 25:10556-10563.
Stevens, C.B. and Halloran, M.C. (2005) Developmental
expression of Sema3G, a novel zebrafish semaphorin. Gene
Expr Patterns 5:647-653. Erratum-name change to
Wolman, M.A., Liu, Y., Tawarayama, H., Shoji, W., and Halloran,
M.C. (2004) Repulsion and attraction of axons
by Sema3D are mediated by different neuropilins in vivo. J
Liu, Y., Berndt, J.D., Su, F., Tawarayama, H., Shoji, W., Kuwada,
J.Y., and Halloran, M.C. (2004). Semaphorin3D
guides retinal axons along the dorsoventral axis of the tectum. J