Justin R. Fallon






Ph.D., University of Pennsylvania
Associate Professor
Department of Neuroscience
312 Medical Research Lab
Tel. (401) 863-9308
E-mail: [email protected]










Lab Personnel

Postdoctoral Fellows

Graduate Students
Mark A. Bowe, Ph.D. [email protected] Mary-Alice Abbott [email protected]
Katherine Deyst, Ph.D. [email protected] Laura Megeath [email protected]
Duane Mendis, Ph.D. [email protected]
David Wells, Ph.D. [email protected]

Senior Research Assistants

Beth A. McKechnie [email protected]

Pamela J. Zabel [email protected]

Research Summary

Mechanisms of Synapse Formation and Plasticity

The synapse is the fundamental unit of cell-cell communication in the nervous system. As such it plays a central role in all aspects of neural function - ranging from the simplest reflex arc to learning and memory. Once thought of as static, synapses are now known to be dynamic structures capable of adaptations throughout life. The structure and physiology of mature synapses are understood in considerable depth, but much less is known about the molecular cues that underlie synaptic development, maintenance and plasticity. A cardinal event in synapse formation is the clustering of neurotransmitter receptors at postsynaptic sites on the cell surface. This aggregation is essential: if the density of neurotransmitter receptors drops below a critical threshold the synapse ceases to function. Synaptic strength can also be modulated by altering the receptor density in the postsynaptic membrane.

The overall goal of research in our laboratory is to understand the molecular mechanisms regulating synapse formation, plasticity, and regeneration. Our work centers on agrin, an extracellular matrix protein that directs the formation of pre- and post- synaptic specializations at nerve-muscle synapses. Several lines of evidence also suggest that agrin plays a related role at synapse in the CNS. We are investigating two fundamental questions. First, what are the signal transduction pathways that parlay the binding of agrin to the cell surface into the differentiation of the postsynaptic apparatus? Our studies here are concentrated at the neuromuscular junction, since this is by far the best understood synapse in the nervous system. The second major area of inquiry is to determine what role agrin plays in the differentiation of synapses between neurons. In both cases we are taking a combined cell biological and molecular approach.

I. Signaling in synaptic differentiation

Agrin binding to cells results in the rapid phosphorylation of the receptor tyrosine kinase MuSK, and subsequently the phosphorylation of AChRs on tyrosine (for a review see Wells and Fallon, 1996). However, the existence of downstream, or parallel signaling pathways is unknown. We asked whether intracellular calcium fluxes might be involved in agrin's pathway (Megeath and Fallon, 1996). We found that loading cells with an intracellular calcium chelator inhibits agrin-induced AChR clustering, but not the phosphorylation of AChRs. Since treatment with these chelators did not change agrin binding levels, and altered neither the number nor mobility of AChRs, we conclude that intracellular calcium fluxes are required for agrin-induced AChR clustering. Moreover, such fluxes function downstream of or parallel to AChR phosphorylation. This finding raises the intriguing possibility that this intracellular calcium flux could be a site of intersection between activity- and agrin- driven postsynaptic differentiation. Current work is aimed at identifying the targets of these calcium fluxes. A current model agrin's signaling pathway is shown in Figure 1.

Agrin isoforms and their receptors

Alternative mRNA splicing yields agrin isoforms that differ greatly in their AChR clustering potency. However, the biochemical basis of this difference was unknown. Splicing at one site ('y') introduces a four amino acid insert . If this insert is missing, agrin is 100-fold less active. We have recently shown that this insert confers heparin binding on agrin, and regulates its association with the cell surface (O'Toole et al., 1996). Although this insert is not required for the association of agrin with its major binding protein, dystroglycan, or to the cell surface, its presence confers heparin sensitivity to both these events. We hypothesize that the four amino acid insert could modulate the binding of agrin to cell surface GAGs.

It seems likely that each of the alternatively-spliced agrin isoforms will have a distinct function. For example, at least four different agrin isoforms are expressed at the developing neuromuscular junction. Some of these forms appear very early in development, while others are not expressed until the final stages of synaptic maturation. To understand the role of these isoforms it is essential to characterize their receptor(s). In ongoing work we have found that each of the agrin isoforms binds to a distinct site on the cell surface, suggesting that each may have a unique cognate receptor.

II. Agrin in the CNS

Agrin binding sites at neuronal synapses

Despite provocative circumstantial evidence, there is no direct support for the proposal that agrin plays a role in synaptogenesis in the CNS. In an attempt to produce cellular and molecular evidence for such a role, we are investigating the localization and dynamics of agrin binding sites on cultured hippocampal neurons. We find that agrin binds selectively at synapses on these cells. Importantly, agrin binds to a majority, but not to all synapses on these cells (Fig. 2). We are currently determining the neurochemical identity of these synapses (e.g. GluR, NMDAR, GABAR). We are also working to determine the identity of these agrin binding sites and their function in synaptic differentiation on neurons.




Recent Publications

Bowe, M.A., K. A. Deyst, J.D. Leszyk, and J R. Fallon. (1994) Identification and purification of an agrin receptor from Torpedo postsynaptic membranes: a heteromeric complex related to the dystroglycans Neuron, 12:1173-1180.

Fallon, J.R., and Hall, Z.W. (1994) Building synapses: Agrin and dystroglycan stick together Trends in Neurosciences 17: 469-473.

Deyst, K.A., M.A. Bowe, J. D. Leszyk, and J.R. Fallon. (1995) The alpha- /beta- dystroglycan complex: Membrane organization and relationship to an agrin receptor J. Biol. Chem. 270: 25956-25959.

Bowe, M. A., and J.R. Fallon. (1995) The Role of Agrin in Synapse formation. Ann. Rev. Neurosci. 18: 443-462. O'Toole, J. J., Deyst, K.A., Nastuk, M.A., and J.R. Fallon. (1996) Alternative splicing of agrin regulates its binding to heparin, a-dystroglycan, and the cell surface. Proc. Natl. Acad. Sci. 93:7369-7374.

Megeath, L.J., and J.F. Fallon (1996) Intracellular calcium fluxes are required for agrin-induced AChR clustering. Soc. Neurosci. Abstr. 22 (1) 535.

Wells, D.G., and J.R. Fallon. (1996) Neuromuscular Junction: State of the Union. Current Biology 6: 1073-1075.

Megeath, L.J., and J.F. Fallon. (1998) Intracellular calcium fluxes regulate agrin-induced AChR clustering. J. Neurosci. 18: (In Press).

Nastuk, M.A., Davis, S., Yancopolous, G.D. and J. R. Fallon. Expression cloning and characterization of NSIST, a novel sulfotransferase expressed by a subset of neurons and postsynaptic targets. (In Review)