[Frontiers in Bioscience 2, d3438-448, September 15, 1997]

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Kimberly M. Rieger-Christ, Karina L. Brierley and Michael A. Reale

Department of Internal Medicine/Oncology, Yale School of Medicine/West Haven Veterans Administration Medical Center, 333 Cedar St., P.O. Box 208032, New Haven, CT 06520-8032

Received 8/25/97 Accepted 9/5/97


1. Abstract
2. Introduction
3. DCC and Development
3.1. DCC guides axonal migrations
3.2 DCC guides cell migrations
3.3. Summary
4. DCC and Cancer
4.1. 18q Allelic loss and DCC expression studies
4.2. Experimental approaches
4.3. Summary
5. Perspective
6. Acknowledgments
7. References


Studies in the developing chicken have implicated DCC in the epithelial-mesenchymal interactions of feather bud formation and have begun to characterize its expression in developing epithelia (14). However, the majority of studies have focused on DCC in neural differentiation and development. DCC antisense and overexpression studies in the rat pheochromocytoma (PC12) model system have suggested a role for DCC in the induction of neuronal differentiation (15-16). Work in the mouse (11) and Xenopus laevis (12) have demonstrated that DCC expression is highest in the neural tube and developing forebrain, midbrain and hindbrain, and this expression is developmentally regulated. Moreover, DCC was expressed as a consequence of neural induction in the Xenopus system. These studies suggested an important functional role for DCC in developing neural tissue, a role that would be characterized in the studies described below.

3.1. DCC guides axonal migrations

The guided migration of axons to connect with a specific target cell is a fundamental process in developmental neurobiology. It has become increasingly clear that this process involves both attractive and repulsive environmental cues that act via signal transduction mechanisms to assist the growth cone in finding its proper pathway (17-18). The netrins are one such family of guidance molecules. Netrins were originally identified as the activity in rat floor plate cells that can promote the outgrowth of commissural axons from rat dorsal spinal cord explants (19-20). These proteins constitute a family of highly phylogenetically conserved molecules related to the extracellular matrix protein, laminin (20-21).

Figure 2. Summary of mutational studies of commissural axon migration in nematodes (C. Elegans). Thick horizontal bars indicate a major inhibitory effect, while the thin horizontal bar indicates a lesser effect of DCC/unc-40 on dorsal axonal migrations [adapted from Drescher (26) and Goodman ].

The netrin response pathway has begun to be clarified with mutational studies primarily in C. Elegans (8,22). These studies have demonstrated that the products of the unc-5, unc-6 and unc-40 ("uncoordinated") genes are required for proper dorsoventral migrations of commissural axons (Figure 2). unc-40 is now known to be a C. elegans homolog of DCC (8), while unc-6 is a netrin homolog (20,23). unc-5 is a transmembrane protein characterized by extracellular immunoglobulin and thrombospondin-1 domains and a cytoplasmic domain that is similar to the ZO-1 tight junction protein (24,25).

Netrin/unc-6 function requires either unc-5 or DCC /unc-40, and mutations in netrin/unc-6 affect both ventral and dorsal migrations of commissural axons. unc-5 mutations affect only dorsal axonal migrations, and there has been no demonstration as yet that unc-5 can function independently of netrin/unc-6. DCC/unc-40 acts cell autonomously to predominantly affect ventral migrations, though it is also required along with unc-5 for dorsal migrations (Figure 2). It is noteworthy that the phenotype of DCC/unc-40 null mutations is less severe than mutations which result in truncated DCC/unc-40 proteins. These nematode studies have been supported by work in Drosophila where the DCC homolog, frazzled, has been shown to be important primarily in ventral axonal migrations (10). Recent studies of netrin-1 (28) and DCC deficient (29)mice have demonstrated defects in commissural axon projections essentially identical to those of invertebrates. Moreover, these knockout studies have extended the effects of DCC and netrin to the developing brain by showing specific defects in the corpus callosum, hippocampal commissure and anterior commissure.

These mutational studies clearly suggest a ligand:receptor relationship for netrins and DCC and/or unc-5, and there is now biochemical evidence in support of this hypothesis. Keino-Masu et al. (9) have shown that netrins bind specifically to DCC expressing cells and that a monoclonal antibody directed to the DCC extracellular domain can abrogate netrin-mediated axonal outgrowth in vitro. Interestingly, this antibody blockade did not affect binding of netrin to the DCC expressing cells. Similar studies have also shown the ability of netrin to bind specifically to unc-5 expressing cells. A simplistic model consistent with these findings is that DCC mediates attractive responses to a netrin gradient, while unc-5 mediates a repulsive response. Refinement of this model awaits further definition of the actual receptor complex(es), since one interpretation of the above data is that DCC and unc-5 are modifiers of the actual receptor (24,25). Also, this model does not adequately explain the findings that unc-5 repulsive function requires DCC/unc-40 and the more severe phenotypes seen with truncated forms of DCC/unc-40. A heteromeric receptor in which truncated DCC/unc-40 acts in a dominant negative fashion has been hypothesized (26).

3.2. DCC guides cell migrations

While DCC has not had any apparent effects on cell proliferation in developmental models, it has been shown to provide pathway guidance for mesodermal and ectodermal cell migrations. Ventral migrations of the mesodermal male linker cell are abnormal in C. elegans DCC/unc-40 mutants, as are migrations of the ectodermal Q neuroblasts and P ectoblasts. The latter two defects are netrin-independent. The Q neuroblast defect is of particular interest because it has been linked to defects in wingless/wnt signaling in C. elegans (8). The wingless/wnt cell fate determination pathway is mediated by beta-catenin (armadillo), an oncogenic protein that is required for cadherin-mediated cell adhesion in addition to its role in signaling (30). The antagonism of beta-catenin signaling function by an interaction with the APC tumor suppressor protein further emphasizes the role of the developmentally important beta-catenin protein in cancer (31,32). The significance of the potential relationship between DCC/unc-40 mediated Q neuroblast migration and wingless/wnt signaling remains to be elucidated (8).

In DCC-/- mice the pontine nuclei at the base of the rostral midbrain are absent (29), a defect that is also seen in netrin-1 deficient mice (28). Though it has not yet been rigorously shown that this represents a mismigration, this seems likely as these nuclei arise in a manner similar to that of spinal commissural axons. They must undergo a ventral, circumferential migration of cells from the lateral recess of the IVth ventricle to their final position in the pons (33).

3.3. Summary

The DCC protein is of unequivocal importance in development, particularly of neural tissue. It functions in the guided migration of cells and cell processes as a component of the netrin response pathway. It appears that DCC may act in this signal transduction pathway as a netrin receptor or a component of the receptor complex, though a definitive receptor:ligand relationship has not yet been demonstrated. It is also clear that DCC can affect migrations in a netrin-independent manner, implying the existence of other DCC ligands.